An antibody binding il-11, antigen-binding fragments thereof and uses thereof

Abstract

The present application relates to the field of biological medicine, and specifically provides an antibody or antigen binding fragment thereof binding IL-11, comprising three heavy chain complementarity determining regions represented by HCDR1, HCDR2 and HCDR3 respectively and three light chain complementarity determining regions represented by LCDR1, LCDR2 and LCDR3 respectively, the antibody or antigen binding fragment thereof is selected from A-I, A-II, A-III or A-IV. The antibody or antigen binding fragment thereof provided by the present application has high binding capacity with IL-11 antigen, can block the binding of IL-11 antigen with its receptor, and then effectively inhibit the fibrogenic effect of IL-11, inhibit or prevent the generation or proliferation of fibrocytes, and can be effectively used for treating or preventing human fibrosis diseases, inflammation, cancer or autoimmune diseases.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of biological medicine, and particularly relates to an antibody binding to IL-11, an antigen binding fragment thereof and application. BACKGROUND

[0002] Fibrosis is a repair response of tissues after injury to protect the relative integrity of the tissue organs. Various harmful stimuli (including toxins, infectious agents, autoimmune reactions and mechanical stress) can induce the fibrotic response of cells. In response to tissue injury, fibroblasts of various origins (including resident fibroblasts, mesenchymal cells, circulating fibroblasts and transdifferentiation of other cell types) can initiate the wound healing response by remodeling the extracellular environment to restore tissue integrity and promote replacement of parenchymal cells. However, persistent injury and damage can lead to the disorder of this process, resulting in the pathological over-deposition of extracellular matrix (ECM) proteins (including collagen, laminin and fibronectin) and the up-regulation of fibroblast activity, causing a chronic inflammatory environment of macrophage and immune cell infiltration. Cytokines and growth factors are released in large quantities, including transforming growth factor-β (TGF-β) family members and Wingless / Int-1 (Wnt1) proteins, which are the main effectors of the fibrosis process. TGF-β and Wnt1 bind to their stem cell surface receptors and initiate downstream signal transduction, leading to the up-regulation of target gene expression, which further enhances fibroblast differentiation and ECM protein production and secretion.

[0003] Worldwide, fibrosis is the main cause of disability and death in many diseases, the main pathological changes are the increase of fibrous connective tissue in the organ tissue, the decrease of parenchymal cells, the continuous progress can cause the destruction of organ structure and the reduction of function, even the failure, which seriously threatens human health and life. According to the relevant statistical data of the United States, nearly 45% of the patients died of various diseases in the country can be attributed to fibrosis. At present, about 25% of the world's population suffers from non-alcoholic fatty liver disease, although non-alcoholic fatty liver disease can improve, but if not treated in time, it will lead to the deterioration of non-alcoholic hepatitis. Non-alcoholic hepatitis is characterized by liver inflammation, liver cell death and liver fibrosis, eventually leading to cirrhosis and liver cancer. Pulmonary fibrosis is a class of end-stage changes of lung diseases characterized by proliferation of fibroblasts, accumulation of a large amount of extracellular matrix, and inflammation damage and tissue structure destruction. That is, after the normal alveolar tissue is damaged, abnormal repair leads to structural abnormalities (scar formation). The incidence and mortality of pulmonary fibrosis are increasing year by year, pulmonary fibrosis is mostly sporadic, the incidence is about 3-5 / 100,000, accounting for about 65% of all interstitial lung diseases. The average survival period after diagnosis is only 2.8 years, and the mortality is higher than that of most tumors, which is called a "tumor-like disease". Therefore, the research and development of drugs for treating anti-fibrosis diseases are particularly urgent.

[0004] Mature IL-11 (interleukin-11) is a polypeptide with 178 amino acids, and the molecular weight is about 23KD. Through the study of the three-dimensional structure of IL-11, it is shown that it is a four-helix bundle structure composed of four alpha helices and loop rings connecting the alpha helices. IL-11 is a pleiotropic cytokine and a member of the IL-6 cytokine family, which shares the same signaling receptor subunit GP130. This family plays a crucial role in the occurrence, development and metastasis of tumors. The production of IL-11 is caused by known major fibrosis-promoting factors, including TGFβ, FGF, PDGF, CTGF or IL-13, and local overexpression of IL-11 can cause local fibrosis of the tissue. The activation of the IL-11 signaling pathway depends on the binding of IL-11 to the cell surface receptor. The receptor of IL-11 is composed of two glycoprotein chains of IL-11Ra and GP130. IL-11Ra has the ability to bind ligands. IL-11 first binds to IL-11Ra with low affinity to form an IL-11 / IL-11Ra heterodimer, which binds to GP130 to form a heterotrimeric protein with high affinity. The IL-11 / IL-11Ra / GP130 heterotrimer homodimerizes to form a hexamer, which further phosphorylates and activates the downstream STAT signaling pathway or the MAPK cascade. Finally, IL-11 transmits signals to the intracellular through the GP130 signaling chain, so that the cell obtains the signal of proliferation and activation. It has been found through research that blocking this signaling pathway can become an effective treatment for a variety of tumors, chronic fibrosis and inflammatory diseases, so the development of antibodies and antigen-binding fragments that bind to IL-11 has important clinical significance. SUMMARY

[0005] In order to meet the needs of patients with fibrosis diseases at home and abroad, the application has screened antibodies and antigen-binding fragments that can specifically bind to IL-11 and block the binding of IL-11 to its receptor.

[0006] The specific technical solutions of the application are as follows:

[0007] The application provides an antibody or an antigen binding fragment thereof which binds to IL-11, and comprises three heavy chain complementarity determining regions represented by HCDR1, HCDR2 and HCDR3 respectively, and three light chain complementarity determining regions represented by LCDR1, LCDR2 and LCDR3 respectively, wherein the amino acid sequence of the heavy chain complementarity determining region HCDR1 is shown as SEQ ID No: 8, the amino acid sequence of the heavy chain complementarity determining region HCDR2 is shown as SEQ ID No: 9, the amino acid sequence of the heavy chain complementarity determining region HCDR3 is shown as SEQ ID No: 10, the amino acid sequence of the light chain complementarity determining region LCDR1 is shown as SEQ ID No: 11, the amino acid sequence of the light chain complementarity determining region LCDR2 is shown as SEQ ID No: 12, and the amino acid sequence of the light chain complementarity determining region LCDR3 is shown as SEQ ID No: 13.

[0008] The antibody or the antigen binding fragment thereof provided by the application can specifically bind to IL-11, block the binding of IL-11 antigen to its receptor, and inhibit IL-11-mediated signal transduction, and the antibody provided by the application has good biological activity.

[0009] The application further comprises a heavy chain variable region and a light chain variable region, wherein the amino acid sequence of the heavy chain variable region is shown as SEQ ID No: 19, and the amino acid sequence of the light chain variable region is shown as SEQ ID No: 20.

[0010] The application further comprises a heavy chain constant region and a light chain constant region, wherein the amino acid sequence of the heavy chain constant region is one of SEQ ID No: 23, SEQ ID No: 24, SEQ ID No: 25 or SEQ ID No: 26, and the amino acid sequence of the light chain constant region is shown as SEQ ID No: 22.

[0011] Further, the antibody or the antigen binding fragment thereof is a chimeric antibody molecule, and the chimeric antibody molecule further comprises a human antibody constant region.

[0012] Preferably, the human antibody constant region comprises a human antibody heavy chain constant region and a human antibody light chain constant region, wherein the amino acid sequence of the human antibody heavy chain constant region is one of SEQ ID No: 27, SEQ ID No: 28 or SEQ ID No: 29, and the amino acid sequence of the human antibody light chain constant region is shown as SEQ ID No: 30.

[0013] The application further provides a nucleic acid molecule encoding the antibody or the antigen binding fragment thereof which binds to IL-11.

[0014] The present application also provides a recombinant DNA expression vector comprising the nucleic acid molecule.

[0015] The present application also provides a host cell transfected with the recombinant DNA expression vector, wherein the host cell is a prokaryotic cell, a yeast cell, an insect cell or a mammalian cell.

[0016] Preferably, the host cell is a mammalian cell, and the mammalian cell is a HEK293 cell, a CHO cell or an NS0 cell.

[0017] The present application also provides a medicament comprising the antibody or antigen-binding fragment thereof that binds to IL-11.

[0018] The present application also provides a combination composition comprising the antibody or antigen-binding fragment thereof that binds to IL-11, and a second composition selected from an antibody or antigen-binding fragment thereof that binds to PD-1, PD-L1, VEGF, VEGFR, TNF-α or TSLP.

[0019] The present application further provides use of the antibody or antigen-binding fragment thereof that binds to IL-11 in the preparation of a medicament for treating or preventing a fibrotic disease, an inflammatory disease, a cancer or an autoimmune disease in a human.

[0020] Preferably, the fibrotic disease is fibrosis of the heart, liver, kidney, lung, gallbladder, bladder, stomach, bone marrow, penis, breast, blood vessel, eye, pancreas, spleen, brain, intestine, muscle or skin.

[0021] Preferably, the inflammatory disease is hepatitis, myocarditis, nephritis, pneumonitis, cholecystitis, cystitis, gastritis, osteomyelitis, prostatitis, mastitis, pancreatitis, enteritis, arthritis, polymyositis, dermatomyositis or dermatitis.

[0022] Preferably, the cancer is leukemia, lung cancer, gastric cancer, esophageal cancer, ovarian cancer, head and neck cancer, melanoma, renal cancer, breast cancer, colorectal cancer, liver cancer, pancreatic cancer or bladder cancer.

[0023] Preferably, the autoimmune disease is psoriasis, Crohn's disease, primary biliary cirrhosis, systemic lupus erythematosus or multiple sclerosis.

[0024] The antibody or antigen binding fragment thereof provided by the present application has high binding capacity with IL-11 antigen, can block the binding of IL-11 antigen with its receptor, and effectively inhibit the fibrosis effect of IL-11, inhibit or prevent the generation or proliferation of fibrocytes, and can be effectively used for treating or preventing human fibrosis diseases, inflammation, cancer or autoimmune diseases, wherein the fibrosis diseases include but are not limited to fibrosis of heart, liver, kidney, lung, gallbladder, bladder, stomach, bone marrow, penis, breast, blood vessel, eye, pancreas, spleen, brain, intestine, muscle or skin; the inflammation includes but is not limited to hepatitis, myocarditis, nephritis, pneumonia, cholecystitis, cystitis, gastritis, osteomyelitis, prostatitis, mastitis, pancreatitis, enteritis, arthritis, polymyositis, dermatomyositis or dermatitis; the cancer includes but is not limited to leukemia, lung cancer, gastric cancer, esophageal cancer, ovarian cancer, head and neck cancer, melanoma, renal cancer, breast cancer, colorectal cancer, liver cancer, pancreatic cancer or bladder cancer; and the autoimmune disease includes but is not limited to psoriasis, Crohn's disease, primary biliary cirrhosis, systemic lupus erythematosus or multiple sclerosis. BRIEF DESCRIPTION OF DRAWINGS

[0025] Figure 1 It is a plasmid map of the pScFv-Disb-HS vector in the embodiment 2 of the present application;

[0026] Figure 2 It is a comparison chart of the affinity of the ELISA anti-IL-11 phage monoclonal antibody in the embodiment 3 of the present application;

[0027] Figure 3 It is a map of the vector pTSE in the embodiment 5 of the present application;

[0028] Figure 4 It is a denatured polyacrylamide gel electrophoresis chart of the murine antibody molecule in the embodiment 5 of the present application;

[0029] Figure 5 It is a comparison chart of the binding capacity of the murine antibody molecule with IL-11 in the embodiment 6 of the present application;

[0030] Figure 6 It is a comparison chart of the competitive inhibition experiment of the murine antibody with IL-11 receptor protein IL-11RA in the embodiment 7 of the present application;

[0031] Figure 7 It is a comparison chart of the inhibition of IL-11 with BaF / 3-IL-11RA cell surface IL-11RA receptor by the murine antibody in the embodiment 8 of the present application;

[0032] Figure 8 It is a comparison chart of the inhibition of the secretion of TIMP-1 by the murine antibody in the embodiment 9 of the present application;

[0033] Figure 9 Denaturing polyacrylamide gel electrophoresis chart of humanized antibody molecules in Example 14 of the present application;

[0034] Figure 10 Chart for comparison of binding ability of humanized antibody molecules in Example 18 of the present application to IL-11;

[0035] Figure 11 Chart for comparison of inhibition of IL-11 binding to IL-11RA receptor on the surface of BaF / 3-IL-11RA cells by humanized antibody molecules in Example 19 of the present application;

[0036] Figure 12 Chart for comparison of inhibition of IL-11 binding to GP130 receptor on the surface of BaF / 3-GP130 cells by humanized antibody molecules in Example 20 of the present application;

[0037] Figure 13 Chart for comparison of biological activity detection (reporter gene) of humanized antibody molecules in Example 21 of the present application;

[0038] Figure 14 Chart for comparison of inhibition of TIMP-1 secretion from embryonic lung fibroblast MRC-5 by humanized antibody molecules in Example 22 of the present application;

[0039] Figure 15 Chart for comparison of cross-binding experiments of humanized antibody molecules to IL-11 of different species in Example 23 of the present application;

[0040] Figure 16 Column chart for change in lung-to-body weight ratio in a mouse model of lung fibrosis in Example 24 of the present application;

[0041] Figure 17 Hematoxylin-eosin (HE) staining and Masson staining charts of lung tissue sections in a mouse model of lung fibrosis in Example 24 of the present application;

[0042] Figure 18 Column chart for change in heart-to-body weight ratio in a mouse model of heart fibrosis in Example 25 of the present application;

[0043] Figure 19 Hematoxylin-eosin (HE) staining and Masson staining charts of heart tissue sections in a mouse model of heart fibrosis in Example 25 of the present application;

[0044] Figure 20 Column chart for urine protein content in a mouse model of kidney fibrosis in Example 26 of the present application;

[0045] Figure 21Hematoxylin-eosin (HE) staining and Masson staining images of kidney tissue sections in the mouse kidney fibrosis model in Example 26 of the present application;

[0046] Figure 22 Bar chart of liver weight changes in the mouse liver fibrosis model in Example 27 of the present application;

[0047] Figure 23 Bar chart of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels in serum of mice in the mouse liver fibrosis model in Example 27 of the present application;

[0048] Figure 24 Hematoxylin-eosin (HE) staining and Masson staining images of kidney tissue sections in the mouse kidney fibrosis model in Example 26 of the present application;

[0049] Figure 25 Figure of heat stability evaluation of anti-IL-11 monoclonal antibody HA-I-A in Example 28 of the present application. DETAILED DESCRIPTION

[0050] In order to more easily understand the present application, before describing the examples, certain technical and scientific terms of the present application are explained as follows:

[0051] The term "antibody" used herein includes whole antibodies and any antigen-binding fragments thereof, and the antibody includes murine antibodies, humanized antibodies, bispecific antibodies or chimeric antibodies, and the antibody can also be a Fab, F(ab)2, Fv or ScFv (single-chain antibody) fragment, and the antibody can be a naturally occurring antibody or an antibody altered (e.g., mutated, deleted, replaced, etc.) as long as it shows binding to the relevant target molecule. The "antibody" herein includes fragments and derivatives thereof, including synthetic antibodies and fragments. As used herein, an antibody is a polypeptide capable of specifically binding to a relevant target molecule (i.e., an antigen to which the antibody has specificity).

[0052] The terms "variable region" and "constant region" used herein, i.e., the sequence region close to the N segment of the antibody heavy chain and light chain is the variable region (V region), and the remaining amino acid sequence close to the C segment is relatively stable, which is the constant region (C region), the variable region includes 3 complementarity determining regions (CDRs) and 4 framework regions (FRs), each light chain variable region and heavy chain variable region are composed of 3 CDR regions and 4 FR regions, the 3 CDR regions of the heavy chain are represented by HCDR1, HCDR2 and HCDR3, and the 3 CDR regions of the light chain are represented by LCDR1, LCDR2 and LCDR3.

[0053] The term "murine antibody molecule" used herein is an antibody obtained after immunizing mice with a human IL-11 antigen.

[0054] The term "chimeric antibody molecule" as used herein is an antibody in which the variable region of a murine antibody is fused to the constant region of a human antibody, which can reduce the immune response induced by the murine antibody in the human body. The chimeric antibody is obtained by inserting the light and heavy chain variable region genes of a murine monoclonal antibody into an expression vector containing the constant region of a human antibody using DNA recombination technology. The variable region of the light and heavy chains of the antibody molecule expressed in this way is of murine origin, while the constant region is of human origin. The entire antibody molecule is about 2 / 3 of human origin. The antibody produced in this way reduces the immunogenicity of the murine antibody while retaining the ability of the parent antibody to specifically bind to the antigen.

[0055] The term "humanized antibody molecule" as used herein is an antibody in which the CDR of a murine monoclonal antibody is transplanted into the variable region of a human antibody to replace the CDR of the human antibody, so that the human antibody acquires the antigen binding specificity of the murine monoclonal antibody while reducing its heterogeneity.

[0056] The term "CHO cell" is a Chinese hamster ovary cell; the term "HEK293E cell" is a human embryonic kidney 293E cell, and the term "NS0 cell" is a mouse NS0 thymoma cell.

[0057] In the present specification, "IL-11" refers to IL-11 from any species and includes isoforms, fragments, variants or homologues of IL-11 from any species. As used herein, a "fragment", "variant" or "homologue" of a protein can optionally be characterized as having at least 60%, preferably 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% sequence identity with the amino acid sequence of a reference protein. In some embodiments, the fragments, variants, isoforms and homologues of the reference protein can be characterized as being capable of having the functions possessed by the reference protein.

[0058] The present application is further described in detail below with reference to the following examples.

[0059] Example 1

[0060] The present application provides an antibody or antigen-binding fragment thereof that binds IL-11, comprising three heavy chain complementarity determining regions designated HCDR1, HCDR2 and HCDR3, respectively, and three light chain complementarity determining regions designated LCDR1, LCDR2 and LCDR3, respectively, the antibody or antigen-binding fragment thereof being selected from any one of the following.

[0061]

[0062] Example 2 Mouse antibody molecule screening

[0063] The present application immunizes mice with IL-11 antigen (the IL-11 protein, IL-11-Fc antigen, and IL-11-mFc ligand protein in subsequent experiments are all human IL-11), optimizes the immunization method, and creates a phage display library. The construction and screening and identification of the specific phage display library are as follows:

[0064] Step one: Immunize mice with IL-11 antigen

[0065] 1. Experimental animals:

[0066] Species and strain: BALB / c, female, mice;

[0067] Body weight: 18-20 g;

[0068] Supplier of experimental animals: Beijing Huafukang Biotechnology Co., Ltd.

[0069] 2. Immunization: Immunize mice with human IL-11 (synthetic gene from Nanjing Kingsway Biotech Co., Ltd., vector constructed and expressed by the company).

[0070] Step two: Construction of phage antibody library

[0071] Take the mouse spleen cells with higher titers, extract the total RNA in the mouse spleen cells using Trizol reagent (purchased from Ambion, catalog number: 15596026), obtain cDNA by RT-PCR, use the cDNA as a template, and use degenerate primers (the degenerate primers used are described in the reference Journal of Immunological Methods 233 (2000) 167-177) to perform PCR amplification, thereby obtaining the immune mouse antibody heavy chain variable region gene library (VH) and the light chain variable region gene library (VL). The pScFv-Disb-HS vector is modified using a series of gene cloning methods on the vector pComb3 vector (purchased from the China Plasmid Vector Strain Cell Strain Gene Preservation Center) to be used for the construction and expression of a phage single-chain antibody library. The modified vector is named pScFv-Disb-HS vector, and its plasmid map is shown in Figure 1 , and based on this vector, a mouse immune phage antibody library is constructed. The light and heavy chain variable region gene libraries are double-digested and ligated to the vector pScFv-Disb-HS, which has also been treated with the same steps of enzyme digestion, to construct the pScFv-Disb-HS-VH-VL gene library.

[0072] Step three: coat the immunotube with IL-11 as the antigen, the antigen coating amount is 5 μg / 500 μL / tube, coat overnight at 4°C, then block the immunotube and the immunophage antibody library with 4% skim milk / PBST respectively, block at room temperature for 1 h. After blocking, the immunophage antibody library is added into the immunotube for antigen-antibody binding, the phage input amount is about 10 9 ~10 12 , after reaction at room temperature for 1 h, the unbound phage is washed away with PBST-PBS, eluted with 0.1 M pH 2.2 Glycine-HCl, and finally the eluted phage antibody solution is neutralized to pH 7.0 or so with 1.5 M pH 8.8 Tris-HCl.

[0073] Step four: infect 10 ml of TG1 bacteria liquid grown to the logarithmic phase with the above neutralized phage, and let stand in a 37°C incubator for 30 min, then take out part of the bacterial liquid for gradient dilution, and spread on 2YTAG plates for calculating the phage output. The remaining bacterial liquid is centrifuged to discard the supernatant, and the bacterial pellet is resuspended in a small amount of culture medium, then aspirated and spread on a 2YTAG large plate for the next round of screening.

[0074] Step five: scrape the bacteria from the large plate after the above infection and spreading, inoculate into 2YTAG liquid medium, shake to the logarithmic phase, then add M13KO7 helper phage superinfection, culture overnight at 28°C and 220 rpm to prepare phage, and PEG / NaCl sedimentation is used to purify the phage for the next round of screening. A round of phage library enrichment screening is performed.

[0075] Step six: screening of IL-11 phage single-chain antibody positive clones: after one round of screening, single clone colonies with good separation are picked and inoculated into 2YTAG liquid medium in a 96-well deep well plate, and cultured at 37°C and 220 rpm until the logarithmic growth phase. About 10 10 μL of helper phage M13KO7 is added to each well, and the temperature is set to 37°C for 30 min of stationary infection. Centrifuge at 4000 rpm for 15 min, discard the supernatant, and resuspend the bacterial pellet in 2YTAK. Culture overnight at 28°C and 220 rpm. After centrifugation at 4000 rpm and 4°C for 15 min, the amplified phage supernatant is aspirated for ELISA identification. Four mouse-derived antibody molecules with high affinity are finally screened, and are named MA-I, MA-II, MA-III and MA-IV. The above obtained monoclonal antibodies are subjected to gene sequencing to determine the correct antibody sequence. After sequencing, the sequences of the four monoclonal antibodies screened above are as follows:

[0076]

[0077] Specifically, SEQ ID No: 16 (amino acid sequence of the heavy chain variable region of MA-I, MA-II):

[0078] EVKLEESGGGLVKPGGSLKLSCAASGFTFSDYYMFWVRQTPEKRLEWVATI SDGGTYTYYPDSVKGRFTISRDNAKNNLYLQMTSLKSEDTAMYYCARDGGYVS SPEAMDYWGQGTSVTVSS;

[0079] SEQ ID No: 17 (amino acid sequence of the light chain variable region of MA-I, MA-IV):

[0080] DIVLTQSTSSLSASLGDRVTISCRASQDISNYLNWYQQKPDGTVKLLIYYTSR LHSGVPSRFSGSGSGTDYSLTISNLEQEDIATYFCQQGNTLPPTFGGGTKLEIK;

[0081] SEQ ID No: 18 (amino acid sequence of the light chain variable region of MA-II):

[0082] DIVLTQSTSSLSASLGDRVTISCRASQDISNYLNWYQQKPDGTVKLLIYYTSR LHSGVPSRFSGSGSGTDYSLTISNLEQEDIATYFCQQGNTLPWTFGGGTKLEIK;

[0083] SEQ ID No: 19 (amino acid sequence of the heavy chain variable region of MA-III):

[0084] EVKLEQSGAEVVKPGALVKMSCKASGYTFTSYWMHWVKQRPGQGLEWIG VIDPSDSYTTYNQKFKGKATLTVDTSSSTGYMQLSSLTSEDSAVYYCSQYGYDVN WYFDVWGAGTTVTVSS;

[0085] SEQ ID No: 20 (amino acid sequence of the light chain variable region of MA-III):

[0086] DIVMTQTTLSLPVSLGDQASISCRSSQSIVHSNGNTYLEWYLQKPGQSPKLLI YEVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDLGVYYCFQGSHVPWTFGGGT KLEIK;

[0087] SEQ ID No:21 (amino acid sequence of the heavy chain variable region of MA-IV):

[0088] EVQLEESGGGLVKPGGSLKLSCVASGFTFSDYYMFWVRQTPEKRLEWVATI SDGGSYSYYPDSVKGRFTISRDNAKNNLYLQMSSLRSEDTAMYYCARDGGYISS PEAMDYWGQGTSVTVSS.

[0089] Example 3 Gradient dilution ELISA to compare the affinity of antibodies

[0090] The four murine antibody molecules (MA-I, MA-II, MA-III and MA-IV) obtained in Example 2 were subjected to monoclonal phage display and purification, and then subjected to phage gradient dilution ELISA to identify the affinity, and the specific method is as follows:

[0091] IL-11 antigen was coated with carbonate buffer at pH 9.6, 100 ng / well / 100 μL, and coated overnight at 4°C. After washing with PBST for three times, the four phage monoclonal antibodies screened in Example 2 were diluted with PBST at five times gradient, and 100 μl of the diluted sample was added to each well, and incubated at room temperature for 1 hour. The ELISA plate was washed with PBST, and HRP-anti-M13 (purchased from Bio-viewshine, item number: GE27-9421-01) monoclonal antibody diluted with 1% BSA-PBST was added to the ELISA plate, and incubated at room temperature for 1 h. The TMB color developing kit was developed (purchased from Kangwei Century, item number: CW0050S), and developed at room temperature for 10 minutes. After termination with 2M H2SO4, the enzyme-labeled instrument was read at 450nm / 630nm, and the corresponding EC50 value was calculated, and the specific data are as follows:

[0092]

[0093] From the above data and as shown in Figure 2 , the four different murine antibody molecules screened in Example 2 can bind to IL-11, and therefore it can be explained that the monoclonal antibodies provided by the present application have high affinity with IL-11.

[0094] Example 4

[0095] The antibody or antigen binding fragment further comprises a heavy chain constant region and a light chain constant region, the amino acid sequence of the heavy chain constant region is one of SEQ ID No: 23, SEQ ID No: 24, SEQ ID No: 25 or SEQ ID No: 26; the amino acid sequence of the light chain constant region is as shown in SEQ ID No: 22, and the specific sequence is as follows:

[0096] SEQ ID No: 22 (amino acid sequence of the light chain constant region of the mouse IgG1 type): k

[0097] ADAAPTVSIFPPSSEQLTSGGASVVCFLNNFYPKDINVKWKIDGSERQNGVLNSWTDQDSKDSTYSMSSTLTLTKDEYERHNSYTCEATHKTSTSPIVKSFNRNEC;

[0098] SEQ ID No: 23 (amino acid sequence of the heavy chain constant region of the mouse IgG1 type):

[0099] AKTTPPSVYPLAPGSAAQTNSMVTLGCLVKGYFPEPVTVTWNSGSLSSGVHTFPAVLQSDLYTLSSSVTVPSSTWPSETVTCNVAHPASSTKVDKKIVPRDCGCKPCICTVPEVSSVFIFPPKPKDVLTITLTPKVTCVVVDISKDDPEVQFSWFVDDVEVHTAQTQPREEQFNSTFRSVSELPIMHQDWLNGKEFKCRVNSAAFPAPIEKTISKTKGRPKAPQVYTIPPPKEQMAKDKVSLTCMITDFFPEDITVEWQWNGQPAENYKNTQPIMDTDGSYFVYSKLNVQKSNWEAGNTFTCSVLHEGLHNHHTEKSLSHSPG;

[0100] SEQ ID No: 24 (amino acid sequence of the heavy chain constant region of the mouse IgG2a type):

[0101] ​AKTTAPSVYPLAPVCGDTTGSSVTLGCLVKGYFPEPVTLTWNSGSLSSGVHTFPAVLQSDLYTLSSSVTVTSSTWPSQSITCNVAHPASSTKVDKKIEPRGPTIKPCPPCKCPAPNLLGGPSVFIFPPKIKDVLMISLSPIVTCVVVDVSEDDPDVQISWFVNNVEVHTAQTQTHREDYNSTLRVVSALPIQHQDWMSGKEFKCKVNNKDLPAPIERTISKPKGSVRAPQVYVLPPPEEEMTKKQVTLTCMVTDFMPEDIYVEWTNNGKTELNYKNTEPVLDSDGSYFMYSKLRVEKKNWVERNSYSCSVVHEGLHNHHTTKSFSRTPGK;

[0102] SEQ ID No: 25 (amino acid sequence of heavy chain constant region of mouse IgG2b type):

[0103] AKTTPPSVYPLAPGCGDTTGSSVTLGCLVKGYFPESVTVTWNSGSLSSSVHTFPALLQSGLYTMSSSVTVPSSTWPSQTVTCSVAHPASSTTVDKKLEPSGPISTINPCPPCKECHKCPAPNLEGGPSVFIFPPNIKDVLMISLTPKVTCVVVDVSEDDPDVQISWFVNNVEVHTAQTQTHREDYNSTIRVVSTLPIQHQDWMSGKEFKCKVNNKDLPSPIERTISKIKGLVRAPQVYILPPPAEQLSRKDVSLTCLVVGFNPGDISVEWTSNGHTEENYKDTAPVLDSDGSYFIYSKLNMKTSKWEKTDSFSCNVRHEGLKNYYLKKTISRSPGK;

[0104] SEQ ID No: 26 (amino acid sequence of heavy chain constant region of mouse IgG3 type):

[0105] ATTTAPSVYPLVPGCSDTSGSSVTLGCLVKGYFPEPVTVKWNYGALSSGVRTVSSVLQSGFYSLSSLVTVPSSTWPSQTVICNVAHPASKTELIKRIEPRIPKPSTPPGSSCPPGNILGGPSVFIFPPKPKDALMISLTPKVTCVVVDVSEDDPDVHVSWFVDNKEVHTAWTQPREAQYNSTFRVVSALPIQHQDWMRGK EFKCKVNNKALPAPIERTISKPKGRAQTPQVYTIPPPREQMSKKKVSLTCLVTNFFSEAISVEWERNGELEQDYKNTPPILDSDGTYFLYSKLTVDTDSW LQGEIFTCSVVHEALHNHHTQKNLSRSPELELNETCAEAQDGELDGLWTTITIFISLFLLSVCYSASVTLFKVKWIFSSVVQVKQTAIPDYRNMIGQGA.

[0106] Example 5: Preparation of mouse antibody molecules

[0107] Example 5 of the present invention, based on Example 4, preferably specifies that the murine antibody molecule includes the heavy chain constant region of murine IgG1 (its amino acid sequence is shown in SEQ ID No: 23) and murine C k The light chain constant region of type [type] (its amino acid sequence is shown in SEQ ID No:22). The specific antibody preparation method is as follows:

[0108] 1. The encoding genes for the heavy chain VH and light chain VL of the four antibody molecules screened in Example 2 were cloned into the vector pTSE (e.g., ...) containing the heavy chain and light chain constant region genes, respectively. Figure 3 As shown in SEQ ID No: 23), the preferred heavy chain constant region is the mouse IgG1 type constant region (amino acid sequence shown in SEQ ID No: 23), and the light chain constant region is mouse C k Chain (amino acid sequence as shown in SEQ ID No: 22), pTSE vector structure as shown Figure 3 (For the preparation process of the pTSE vector, please refer to paragraph

[0019] on page 3 of the instruction manual CN103525868A).

[0109] 2. HEK293 cells were transiently transfected (purchased from the Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences, catalog number GNHu43) for antibody expression. Four monoclonal antibodies were purified using an AKTA instrument via a protein A affinity column. Protein concentration was determined using a BCA kit (purchased from Beijing Huitian Oriental Technology Co., Ltd., catalog number BCA0020). Protein size was then identified by SDS-PAGE. The results are shown below. Figure 4 As shown, from left to right, the images represent non-reduced MA-I, MA-II, MA-III, and MA-IV, protein molecular weight Marker 1, protein molecular weight Marker 2, and reduced MA-I, MA-II, MA-III, and MA-IV murine anti-IL-11 monoclonal antibodies. The molecular weight of each band is consistent with the theoretical values.

[0110] Example 6: Binding experiment of mouse antibody molecules with IL-11

[0111] IL-11 antigen was coated with carbonate buffer (pH 9.6), 100 ng / well / 100 μL, and incubated overnight at 4°C. The cells were washed five times with 300 μL / well PBST, then blocked with 280 μL / well of 1% BSA-PBST at 37°C for 1 h. Different concentrations of MA-I, MA-II, MA-III, and MA-IV mouse antibodies were added, with an initial maximum concentration of 5 μg / mL for each antibody. Each antibody was serially diluted 5-fold, for a total of 8 dilutions, and incubated at 37°C for 1 h. The cells were then washed five times with 300 μL / well PBST, and Goat Anti-Mouse IgG-HRP (purchased from Solarbio, catalog number: SE131) diluted 1:2000 with 1% BSA-PBST was added, and the cells were incubated at 37°C for 1 h. The TMB chromogenic kit was used for color development. 100 μL / well was incubated at room temperature for 8 min, then the color development was stopped with 2 M H₂SO₄. Readings were taken at 450 nm and 630 nm using a microplate reader, and the corresponding EC50 values ​​were calculated. Specific data are as follows:

[0112]

[0113] Based on the above data and as follows Figure 5 As shown, all four different murine antibody molecules screened were able to bind to IL-11 and had high affinity.

[0114] Example 7: Competitive inhibition experiment between murine antibody and IL-11 receptor protein IL-11RA

[0115] IL-11-Fc was coated with carbonate buffer at pH 9.6, 200 ng / well / 100 μL, coated overnight at 4°C. Washed five times with 300 μL / well PBST, then added 1% BSA-PBST, 280 μL / well, blocked at 37°C for 1 h, then added IL-11RA-Fc (IgG4 type) diluted to 0.5 μg / mL with 1% BSA-PBST, 50 μL / well, then added MA-I, MA-II, MA-III and MA-IV mouse antibodies at different dilution concentrations, 50 μL / well, the starting highest concentration of the five antibodies was 100 μg / mL, each antibody was diluted by 2-fold gradient, a total of 13 gradients for each antibody, incubated at 37°C for 3 h. Washed five times with 300 μL / well PBST, then added Anti-Human IgG4-HRP Mouse monoclonal antibody (purchased from Sigma, item number: SAB4200770) diluted 1:5000 with 2% BSA-PBST, incubated at 37°C for 1 h. Color development with TMB color development kit, 100 μL / well, color development at room temperature for 15 min, then color development was terminated with 2M H2SO4. Readings were taken at 450 nm / 630 nm with a microplate reader, and the corresponding IC50 values were calculated, and the specific data are as follows:

[0116]

[0117] From the above data and as shown in Figure 6 , the four different mouse antibodies screened can all compete with the receptor protein IL-11RA, indicating that they can effectively inhibit the binding of IL-11 to the receptor protein IL-11RA.

[0118] Example 8 Inhibition of the binding of IL-11 to IL-11RA receptors on the surface of BaF / 3-IL-11RA cells by mouse antibodies

[0119] The BaF / 3-IL-11RA cell line was counted, and a certain number of cells were centrifuged and resuspended with PBS buffer. The cell density was adjusted to 1E+6 cells / mL, 100 μL / well, and added to a 96-well plate. The IL-11-mFc ligand protein was diluted with PBS to a concentration of 18 μg / mL, 50 μL / well, and added to the corresponding position of the 96-well plate containing the BaF / 3-IL-11RA cells. After mixing gently, the 96-well plate was placed at 4°C and incubated for 1 h. The four mouse antibody molecules MA-I, MA-II, MA-III, and MA-IV were gradient diluted with PBS, with an initial concentration of 800 μg / mL, 3-fold gradient dilution, a total of 10 gradients, 50 μL / well, and added to the corresponding position of the 96-well plate containing the mixture of BaF / 3-IL-11RA cells and IL-11-mFc ligand protein. After mixing, the plate was incubated at 4°C for 2 h. After incubation, the cells were centrifuged at 3000 rpm, washed once with PBS buffer, and the cell pellet was collected. Goat anti-mouse IgG Human ads-FITC antibody (purchased from SouthernBiotech, catalog number 1030-02) was added to the cell pellet, and incubated at 4°C for 30 min. After centrifugation at 3000 rpm, the cells were washed once with PBS buffer, resuspended with 100 μL of PBS buffer, and detected by flow cytometry. The fluorescence signal in the FL1-A channel was collected. The dose-effect curve was drawn, and the corresponding IC50 value was calculated. The specific data are as follows:

[0120]

[0121] From the above data and Figure 7 It can be seen that the four different mouse candidate molecules screened can effectively inhibit the binding of IL-11 ligand protein to the cell surface IL-11RA receptor.

[0122] Example 9 Inhibition of TIMP-1 Secretion from Embryonic Lung Fibroblast MRC-5 by Mouse Antibodies

[0123] Embryonic lung fibroblast MRC-5 was counted after trypsin digestion, a certain amount of cells was taken, the cells were resuspended after centrifugation with MEM complete medium (purchased from GIBCO, item number 10370-021), and the cell density was adjusted to 2E+5 cells / mL, 100 μL / well, and added to a 96-well plate. The IL-11-mFc ligand protein was diluted with MEM complete medium, and the concentration was prepared to be 16 μg / mL, 50 μL / well was added to the corresponding 96-well plate. The four mouse-derived antibody molecules MA-I, MA-II, MA-III and MA-IV were gradient diluted with MEM complete medium, and the initial concentration was prepared to be 40 μg / mL, 2-fold gradient dilution, a total of 8 gradients, 50 μL / well, added to the 96-well plate containing the cell suspension and the IL-11-mFc ligand protein suspension, mixed gently, and incubated in a 37℃ CO2 incubator overnight for about 20 h. The cell culture supernatant was detected by TIMP-1 ELISA kit (purchased from Yikexie Biotechnology Co., Ltd., item number EH021-96).

[0124] Human TIMP-1 detection kit: add cell supernatant and standard to sample wells, 100 μL / well. Immediately add biotinylated antibody working solution (1:100 dilution), 50 μL / well, cover the plate with sealing film, shake and incubate at room temperature for 2 h. After incubation, wash the plate 4 times with washing solution, add enzyme conjugate working solution (1:100 dilution) in the TIMP-1 detection kit, 100 μL / well. Cover the plate with sealing film and shake and incubate at room temperature for 1 h. After incubation, wash the plate 4 times with washing solution. Add TMB color developing solution, 100 μL / well, avoid light, incubate at room temperature for about 15 minutes, add 100 μL / well Stop solution to terminate the reaction. Read the value at 450 nm on a microplate reader, and calculate the corresponding IC50 value, and the specific data are as follows:

[0125]

[0126] From the above data and Figure 8 It can be seen that the four different mouse-derived candidate molecules screened can effectively inhibit the release of TIMP-1 from human embryonic lung fibroblast MRC-5 stimulated by IL-11 ligand protein.

[0127] Example 10

[0128] The antibody or antigen binding fragment thereof in the embodiment 10 of the present application is further limited to be a chimeric antibody molecule, and the chimeric antibody molecule further comprises a human antibody constant region, the human antibody constant region comprises a human antibody heavy chain constant region and a human antibody light chain constant region, the amino acid sequence of the human antibody heavy chain constant region is one of SEQ ID No: 27, SEQ ID No: 28 or SEQ ID No: 29; and the amino acid sequence of the human antibody light chain constant region is as shown in SEQ ID No: 30.

[0129] SEQ ID No: 27 (amino acid sequence of heavy chain constant region of human IgG1 type):

[0130] ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK;

[0131] SEQ ID No: 28 (amino acid sequence of heavy chain constant region of human IgG2 type):

[0132] ASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSNFGTQTYTCNVDHKPSNTKVDKTVERKCCVECPPCPAPPVAGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSNKGLPAPIEKTISKTKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPMLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK;

[0133] SEQ ID No: 29 (amino acid sequence of heavy chain constant region of human IgG4 type):

[0134] ASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGPPCPPCPAPEFLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGK;

[0135] SEQ ID No:30 (human C k chain constant region amino acid sequence):

[0136] RTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGE C.

[0137] Preparation of chimeric antibody molecules

[0138] The embodiment 11 of the present application further limits the humanized antibody constant region to include a human IgG1 type heavy chain constant region (the amino acid sequence of which is shown as SEQ ID No: 27) and a human C k type light chain constant region (the amino acid sequence of which is shown as SEQ ID No: 30) based on the embodiment 10.

[0139] Specific preparation method:

[0140] The heavy chain variable region VH (SEQ ID No: 16) of the murine antibody molecules MA-I and MA-II obtained by screening the phage antibody library in embodiment 2, and the light chain variable region VL gene of MA-I (SEQ ID No: 17) and the light chain variable region VL gene of MA-II (SEQ ID No: 18) are kept as murine sequences, and are respectively cloned into the vector pTSE (as shown in Figure 3 SEQ ID No: 28) carrying the heavy chain constant region and light chain constant region genes, the heavy chain constant region being human IgG1 type (the amino acid sequence being shown as SEQ ID No: 27), and the light chain constant region being human Ck Chimeric antibodies CA-I, CA-II were obtained by transient transfection of HEK293E cells (purchased from: Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences, item number: GNHu43) for antibody expression.

[0141] Example 12 Humanization of murine antibody molecules

[0142] First, the sequences of murine antibody molecules MA-I and MA-II in Example 2 were compared with the human antibody germline database (v-base) to find higher homologous human antibody light and heavy chain germlines as candidate sequences, and then the CDR sequences of murine antibody molecules MA-I and MA-II were transplanted onto the human candidate sequences for homology modeling. Then, the key framework amino acid residues that may play an important role in maintaining the CDR loop structure were designed by three-dimensional structure simulation calculation, so as to design the back mutation of humanized antibodies. The designed light and heavy chain variable region sequences of humanized antibodies containing back mutations were synthesized by Nanjing Kingsriver Biotechnology Co., Ltd., and then connected to the transient expression vector, and the combination of humanized light and heavy chains was analyzed, in which MA-I obtained the following humanized antibody molecules: HA-I-A, HA-I-B, HA-I-C, HA-I-D; MA-II obtained the following humanized antibody molecules: HA-II-A, HA-II-B, HA-II-C, HA-II-D; the sequences of the above 8 monoclonal antibodies screened are as follows:

[0143]

[0144] Specifically, SEQ ID No: 31 (amino acid sequence of the heavy chain variable region of HA-I-A, HA-I-C, HA-II-A and HA-II-B):

[0145] QVQLVESGGGLVKPGGSLRLSCAASGFTFSDYYMFWVRQAPGKGLEWVATI SDGGTYTYYPDSVKGRFTISRDNAKNSLYLQMNSLRAEDTAVYYCARDGGYVSS PEAMDYWGQGTLVTVSS;

[0146] SEQ ID No: 32 (amino acid sequence of the light chain variable region of HA-I-A):

[0147] DIVLTQSPSSLSASVGDRVTITCRASQDISNYLNWYQQKPGKAPKLLIYYTS RLHSGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQGNTLPPTFGGGTKVEIK;

[0148] SEQ ID No:33 (amino acid sequence of the heavy chain variable region of HA-I-B and HA-II-C):

[0149] QVQLVESGGGLVKPGGSLRLSCAASGFTFSDYYMFWVRQAPGKGLEWVST ISDGGTYTYYPDSVKGRFTISRDNAKNSLYLQMNSLRAEDTAVYYCARDGGYVS SPEAMDYWGQGTLVTVSS;

[0150] SEQ ID No:34 (amino acid sequence of the light chain variable region of HA-I-B):

[0151] DIVLTQSPSSLSASVGDRVTITCRASQDISNYLNWYQQKPGKAPKLLIYYTS RLHSGVPSRFSGSGSGTDYTFTISSLQPEDIATYYCQQGNTLPPTFGGGTKVEIK;

[0152] SEQ ID No:35 (amino acid sequence of the light chain variable region of HA-I-C):

[0153] DIVLTQSPSSLSASVGDRVTITCRASQDISNYLNWYQQKPGKAVKLLIYYTS RLHSGVPSRFSGSGSGTDYTFTISSLQPEDIATYFCQQGNTLPPTFGGGTKVEIK;

[0154] SEQ ID No:36 (amino acid sequence of the heavy chain variable region of HA-I-D and HA-II-D):

[0155] QVQLVESGGGLVKPGGSLRLSCAASGFTFSDYYMFWVRQAPGKGLEWVATI SDGGTYTYYPDSVKGRFTISRDNAKNSLYLQMNSLRAEDTAMYYCARDGGYVS SPEAMDYWGQGTSVTVSS;

[0156] SEQ ID No:37 (amino acid sequence of the light chain variable region of HA-I-D):

[0157] DIVLTQSPSSLSASVGDRVTITCRASQDISNYLNWYQQKPGGAVKLLIYYTS RLHSGVPSRFSGSGSGTDYTFTISSLQPEDIATYFCQQGNTLPPTFGGGTKVEIK;

[0158] SEQ ID No:38 (amino acid sequence of the light chain variable region of HA-II-A) :

[0159] DIVLTQSPSSLSASVGDRVTITCRASQDISNYLNWYQQKPGKAPKLLIYYTS RLHSGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQGNTLPWTFGGGTKVEIK;

[0160] SEQ ID No:39 (amino acid sequence of the light chain variable region of HA-II-B and HA-II-C) :

[0161] DIVLTQSPSSLSASVGDRVTITCRASQDISNYLNWYQQKPGKAPKLLIYYTS RLHSGVPSRFSGSGSGTDYTFTISSLQPEDIATYYCQQGNTLPWTFGGGTKVEIK;

[0162] SEQ ID No:40 (amino acid sequence of the light chain variable region of HA-II-D) :

[0163] DIVLTQSPSSLSASVGDRVTITCRASQDISNYLNWYQQKPGGTVKLLIYYTS RLHSGVPSRFSGSGSGTDYTFTISSLQPEDIATYFCQQGNTLPWTFGGGTKVEIK.

[0164] Embodiment 13

[0165] The embodiment 13 of the present application further limits the humanized antibody constant region to include a humanized antibody heavy chain constant region and a humanized antibody light chain constant region, the amino acid sequence of the humanized antibody heavy chain constant region is one of SEQ ID No: 27, SEQ ID No: 28 or SEQ ID No: 29; the amino acid sequence of the humanized antibody light chain constant region is shown in SEQ ID No: 30.

[0166] The specific sequence of the humanized antibody constant region is the same as that of embodiment 10.

[0167] Preparation of humanized antibody molecules

[0168] Example 14 further defines the humanized antibody molecules of the present application to include a human IgG1 type heavy chain constant region (the amino acid sequence of which is shown as SEQ ID No: 27) and a human C k type light chain constant region (the amino acid sequence of which is shown as SEQ ID No: 30).

[0169] The genes encoding the heavy chain VHand light chain VLof the 8 humanized antibody molecules HA-I-A, HA-I-B, HA-I-C, HA-I-D, HA-II-A, HA-II-B, HA-II-C, HA-II-D obtained in Example 12 were cloned into the vector pTSE (as shown in Figure 3 ) containing the heavy chain constant region and light chain constant region genes, respectively. The heavy chain constant region was human IgG1 type (the amino acid sequence of which is shown as SEQ ID NO: 27) and the light chain constant region was C k ( the amino acid sequence of which is shown as SEQ ID NO: 30).

[0170] The 2 chimeric antibodies CA-I and CA-II obtained in Example 11 and the 8 humanized antibody molecules HA-I-A, HA-I-B, HA-I-C, HA-I-D, HA-II-A, HA-II-B, HA-II-C, HA-II-D obtained in Example 12 were transiently transfected into HEK293 cells (purchased from the Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences, item number GNHu43) for antibody expression. The monoclonal antibodies were purified by protein A affinity column using AKTA instrument, and the protein concentration was determined using BCA kit (purchased from Beijing Huitian Dongfang Science and Technology Co., Ltd., item number BCA0020). The protein size was then identified by SDS-PAGE, and the results are shown in Figure 9 . From left to right, the non-reduced protein molecular weights are HA-I-A, HA-I-B, HA-I-C, HA-I-D, chimeric antibody CA-I prepared in Example 11, reduced protein molecular weight Marker, HA-II-A, HA-II-B, HA-II-C, HA-II-D, chimeric antibody CA-II. The molecular weight of each band is consistent with the theory.

[0171] Example 15

[0172] Example 15 defines the antibody or antigen-binding fragment thereof disclosed in the present application as one or a combination of Fab, F(ab)2, Fv or ScFv, which is not limited to the monoclonal antibody disclosed in the present application.

[0173] Example 16

[0174] The embodiment 16 of the present application also provides a nucleic acid molecule, which encodes the antibody or antigen-binding fragment thereof binding to IL-11.

[0175] The present application also provides a recombinant DNA expression vector, which comprises the nucleic acid molecule.

[0176] The present application also provides a host cell transfected with the recombinant DNA expression vector, which comprises a prokaryotic cell, a yeast cell, an insect cell or a mammalian cell.

[0177] Further preferably, the host cell is a mammalian cell, and the mammalian cell is a HEK293 cell, a CHO cell or an NS0 cell.

[0178] The present application also provides a medicament, which comprises the antibody or antigen-binding fragment thereof binding to IL-11.

[0179] The present application also provides a detection reagent for detecting the antibody or antigen-binding fragment thereof binding to IL-11.

[0180] The present application also provides a combination composition, which comprises the antibody or antigen-binding fragment thereof binding to IL-11, and further comprises a second composition selected from an antibody or antigen-binding fragment thereof binding to PD-1, PD-L1, VEGF, VEGFR, TNF-α or TSLP.

[0181] Preferably, the antibody or antigen-binding fragment thereof binding to PD-1 is selected from DFPD1-9, DFPD1-10, DFPD1-11, DFPD1-12, DFPD1-13, Nivolumab, Pembrolizumab, Tepezikimab, Sintilimab, Toripalimab, Camrelizumab, Pidilizumab or Sipatuzumab, wherein the heavy chain constant region and the light chain constant region sequence of DFPD1-9, DFPD1-10, DFPD1-11, DFPD1-12 and DFPD1-13 are the same as those of the monoclonal antibody against PD-1 provided in the patent application No. CN201510312910.8.

[0182] Preferably, the antibody or antigen-binding fragment thereof binding to VEGF is selected from Aflibercept, Bevacizumab or Ranibizumab.

[0183] Embodiment 17

[0184] Embodiment 17 provides use of an antibody or an antigen-binding fragment thereof that binds IL-11 in the preparation of a medicament for treating or preventing a fibrotic disease, an inflammation, a cancer, or an autoimmune disease in a human.

[0185] Preferably, the fibrotic disease comprises, but is not limited to, fibrosis of heart, liver, kidney, lung, gallbladder, bladder, stomach, bone marrow, penis, breast, blood vessel, eye, pancreas, spleen, brain, intestine, muscle, or skin.

[0186] Preferably, the inflammation comprises, but is not limited to, hepatitis, myocarditis, nephritis, pneumonitis, cholecystitis, cystitis, gastritis, osteomyelitis, prostatitis, mastitis, pancreatitis, enteritis, arthritis, polymyositis, dermatomyositis, or dermatitis.

[0187] Preferably, the cancer comprises, but is not limited to, leukemia, lung cancer, gastric cancer, esophageal cancer, ovarian cancer, head and neck cancer, melanoma, renal cancer, breast cancer, colorectal cancer, liver cancer, pancreatic cancer, or bladder cancer.

[0188] Preferably, the autoimmune disease comprises, but is not limited to, psoriasis, Crohn's disease, primary biliary cirrhosis, systemic lupus erythematosus, or multiple sclerosis.

[0189] The present application also provides a method of treating or preventing a fibrotic disease, an inflammation, a cancer, or an autoimmune disease in a subject, the method comprising administering to the subject a therapeutically or prophylactically effective amount of an antibody or an antigen-binding fragment thereof that binds IL-11.

[0190] Embodiment 18 Experiment of binding of humanized antibody molecules to IL-11

[0191] IL-11 antigen was coated with carbonate buffer at pH 9.6, 100 ng / well / 100 μL, coated overnight at 4°C. Washed five times with 300 μL / well PBST, then added 1% BSA-PBST 280 μL / well, blocked at 37°C for 1 h. The initial concentration of humanized antibodies HA-I-A, HA-I-B, HA-I-C, HA-I-D, HA-II-A, HA-II-B, HA-II-C, HA-II-D and the chimeric antibodies CA-I, CA-II prepared in Example 11, humanized antibodies were all 10 μg / mL, 5-fold gradient dilution, a total of 8 gradients, incubated at 37°C for 1 h. Washed five times with 300 μL / well PBST, then added goat anti-Human IgG Fab HRP (purchased from invitrogen, item number: 31482) diluted 1:5000 with 1% BSA-PBST, incubated at 37°C for 1 h. Color development with TMB color development kit, 100 μL / well, color development at room temperature for 5 min, then color development was terminated with 2M H2SO4. Readings were taken at 450 nm / 630 nm by a microplate reader, and the corresponding EC50values were calculated, and the specific data are as follows:

[0192]

[0193] As shown in the above data and experimental results, Figure 10 all 8 different humanized antibody molecules can bind to IL-11, and the EC50values of HA-I-A, HA-I-B, HA-I-C, HA-I-D humanized antibody molecules are close to the chimeric antibody CA-I, and the EC50values of HA-II-A, HA-II-B, HA-II-C, HA-II-D humanized antibody molecules are close to the chimeric antibody CA-II, indicating that the humanized antibody molecules retain the high binding ability of the murine parent antibody MA-I, MA-II to IL-11.

[0194] Example 19 Humanized antibody molecules inhibit the binding of IL-11 to IL-11RA receptors on the surface of BaF / 3-IL-11RA cells

[0195] The four humanized antibody molecules HA-I-A, HA-I-B, HA-I-C and HA-I-D with better activity at the protein level were selected for cell activity evaluation experiment. The BaF / 3-IL-11RA cell line was counted, a certain amount of cells were centrifuged and resuspended with PBS buffer, and the cell density was adjusted to 1E+6 cells / mL, 100 μL / well, and added to the 96-well plate. The IL-11-mFc ligand protein was diluted with PBS buffer to a concentration of 18 μg / mL, 50 μL / well, and added to the corresponding position of the 96-well plate containing the BaF / 3-IL-11RA cells. After gentle mixing, the 96-well plate was placed at 4°C for incubation for 1 h. The four humanized antibody molecules HA-I-A, HA-I-B, HA-I-C and HA-I-D were gradient diluted with PBS buffer, and the initial concentration was prepared to be 800 μg / mL, 3-fold gradient dilution, a total of 10 gradients, 50 μL / well, added to the corresponding position of the 96-well plate containing the mixture of BaF / 3-IL-11RA cells and IL-11-mFc ligand protein. After mixing, it was placed at 4°C for 2 h of incubation. After incubation, the cells were washed once by centrifugation at 3000 rpm with PBS buffer, and the cell precipitate was collected. Goat anti-mouse IgG Human ads-FITC antibody (purchased from SouthernBiotech, catalog number 1030-02) was added to the cell precipitate, and incubated at 4°C for 30 min, then centrifuged at 3000 rpm for washing once, resuspended with 100 μL of PBS, and then detected by flow cytometry. The fluorescence signal in the FL1-A channel was collected. The dose-effect curve was drawn, and the corresponding IC50 value was calculated. The specific data are as follows:

[0196]

[0197] From the above data and Figure 11 It can be seen that the four humanized candidate molecules selected can inhibit the binding of IL-11 ligand protein to the surface IL-11RA receptor of BaF / 3-IL-11RA cells.

[0198] Example 20 Humanized antibody molecules inhibit the binding of IL-11 to the surface GP130 receptor of BaF / 3-GP130 cells

[0199] The BaF / 3-GP130 cell line was counted, and a certain number of cells were centrifuged and resuspended with PBS buffer. The cell density was adjusted to 1E+6 cells / mL, 100 μL / well, and added to a 96-well plate. The IL-11-mFc ligand protein was diluted with PBS, and the concentration was adjusted to 12 μg / mL, 50 μL / well, and added to the corresponding position of the 96-well plate containing the BaF / 3-GP130 cells. After mixing gently, the 96-well plate was placed at 4°C and incubated for 1 h. The four human antibody molecules HA-I-A, HA-I-B, HA-I-C, and HA-I-D were gradient diluted with PBS buffer, and the initial concentration was adjusted to 2000 μg / mL, 2-fold gradient dilution, a total of 10 gradients, 50 μL / well, and added to the corresponding position of the 96-well plate containing the BaF / 3-GP130 cells and the IL-11-mFc ligand protein. After mixing, the 96-well plate was placed at 4°C and incubated for 2 h. After incubation, the cells were washed once with PBS buffer at 3000 rpm, and the cell precipitate was collected. Goat anti-mouse IgG Human ads-FITC antibody (purchased from SouthernBiotech, catalog number 1030-02) was added to the cell precipitate, 100 μL / well, and incubated at 4°C for 30 min. After washing once with PBS buffer at 3000 rpm, the cells were resuspended with 100 μL / well of PBS buffer, and the fluorescence signal in the FL1-A channel was detected by flow cytometry. The dose-effect curve was drawn, and the corresponding IC50 value was calculated. The specific data are as follows:

[0200]

[0201] From the above data and Figure 12 It can be seen that the four humanized candidate molecules screened can block the binding of the IL-11 ligand protein to the GP130 receptor on the surface of the BaF / 3-GP130 cells.

[0202] Example 21 Biological activity detection of humanized antibody molecules (reporter gene)

[0203] The BaF / 3-IL-11RA-GP130-STAT3-Luc engineered cell strain was counted, and the cell density was adjusted to 2E+6 cells / mL using a sample diluent (which contains 90% IMDM, 10% FBS, and 10 ng / mL mouse IL-3). After gentle mixing, 50 μL / well of the cell solution was added to a 96-well plate. Four humanized antibody molecules, HA-I-A, HA-I-B, HA-I-C, and HA-I-D, were diluted to an initial concentration of 200 μg / mL using a sample diluent, and 5-fold gradient dilution was performed, for a total of 10 gradients, 100 μL / well. The 96-well plate containing the engineered cell strain was added to the corresponding positions, and two replicate wells were set up for each sample concentration. The sample diluent was used to prepare IL-11 protein at a concentration of 10 μg / mL, 50 μL / well, and added to the 96-well plate containing the engineered cell strain and the humanized antibody molecules. The cell culture plate was gently mixed and incubated in a 37°C CO2 incubator for 6 h. The supernatant was discarded by centrifugation, and 10 μL / well of lysis solution was added to a 384-well plate. An equal amount of luciferase reaction substrate (purchased from Promega Biotechnology Co., Ltd., catalog number E2610) was added, and the reaction was performed at room temperature for 5 min. The fluorescence value was read on an enzyme marker, and the corresponding IC50 value was calculated. The specific data are as follows:

[0204]

[0205] As shown by the above data and Figure 13 , the four humanized antibody molecules screened can block the binding of IL-11 to IL-11RA and GP130 receptors and inhibit the conduction of the signal pathway.

[0206] Example 22 Inhibition of TIMP-1 Secretion from Embryonic Lung Fibroblast MRC-5 by Humanized Antibody Molecules

[0207] Embryonic lung fibroblast MRC-5 was counted after trypsin digestion, a certain amount of cells was taken, the cells were resuspended with MEM complete medium after centrifugation, and the cell density was adjusted to 2E+5 cells / mL, 100 μL / well, and added to a 96-well plate. The IL-11-mFc ligand protein was diluted with MEM complete medium, and the concentration was prepared to be 16 μg / mL, 50 μL / well, and added to the corresponding 96-well plate. The four humanized antibody molecules HA-I-A, HA-I-B, HA-I-C and HA-I-D were gradient diluted with MEM complete medium, and the initial concentration was prepared to be 40 μg / mL, 3 times gradient dilution, a total of 8 gradients, 50 μL / well, added to the 96-well plate containing the cell suspension and the IL-11-mFc ligand protein suspension, mixed gently and uniformly, incubated in a 37°C CO2 incubator overnight, about 20 h, and the cell culture supernatant was detected by TIMP-1 ELISA kit (method same as Example 9). The microplate reader was read at 450 nm, and the corresponding IC50 value was calculated, and the specific data were as follows:

[0208]

[0209] From the above data and Figure 14 It can be seen that the four humanized antibody molecules screened can effectively inhibit the release of TIMP-1 of human embryonic lung fibroblast MRC-5 stimulated by IL-11 ligand protein.

[0210] Example 23 Cross-binding experiment of humanized antibody molecules with different species of IL-11

[0211] The humanized antibody molecule HA-I-A with better protein level and function detection activity was selected for cross-binding detection with different species of IL-11. Human IL-11, mouse IL-11 (purchased from Beijing Yiqiao Godstate Technology Co., Ltd., item number: 50117-MNCE), rat IL-11 (purchased from Kanglang Biotechnology, item number: KL40001Ra), and cynomolgus monkey IL-11 (purchased from Yiqiao Godstate, item number: 90925-CNCE) were coated with 100 ng / well / 100 μL of carbonate buffer at pH 9.6 at 4°C overnight. Five washes with 300 μL / well PBST were performed, and 1% BSA-PBST was added at 280 μL / well, and incubated at 37°C for 1 h. The humanized antibody HA-I-A was diluted with 1% BSA-PBST, and the initial concentration was 50 μg / mL, and 5-fold gradient dilution was performed, a total of 9 gradients, and each gradient had two duplicate wells, 100 μL / well was added to the 96-well plate, and incubated at 37°C for 1 h. Five washes with 300 μL / well PBST were performed, and goat anti-human IgG Fab HRP (purchased from invitrogen, item number: 31482) was diluted with 1% BSA-PBST at a working concentration of 1:5000, 100 μL / well was added to the 96-well plate, and incubated at 37°C for 1 h. Five washes with 300 μL / well PBST were performed, and TMB color developing kit was used for color development, 100 μL / well, color development at room temperature for 5 min, and then color development was terminated with 2M H2SO4. The microplate reader was read at 450 nm / 630 nm, and the corresponding EC50 values were calculated, and the specific data are as follows:

[0212]

[0213] According to the above data and as shown in Figure 15 , the humanized antibody molecule HA-I-A can bind to human IL-11, mouse IL-11, rat IL-11, and cynomolgus monkey IL-11, and has high affinity.

[0214] Example 24 Therapeutic efficacy experiment of anti-IL-11 monoclonal antibody on pulmonary fibrosis

[0215] Bleomycin (bLF) was used to model to study the therapeutic effect of anti-IL-11 monoclonal antibody HA-I-A on pulmonary fibrosis.

[0216] Animal species: C57BL / 6J mice (purchased from Jiangsu Jizhuangkang Biotechnology Co., Ltd.)

[0217] Number, gender, and mouse age: 6 per group, male, 6-8 weeks;

[0218] The control group was only injected with normal saline.

[0219] The administration group was administered with the HA-I-A antibody molecule twice a week for 4 weeks.

[0220] The animal weight was measured once a week, and the animals were observed for abnormalities; organ weight detection: the lung organs were collected, the lung organ weight was measured, and the lung to body weight ratio was calculated; lung pathology detection: lung sections were made and hematoxylin-eosin (HE) and Masson staining were used to observe the degree of lung fibrosis.

[0221] As shown in the results, Figure 16 the lung to body weight ratio of the mice in the administration group was significantly smaller than that in the control group after administration of the HA-I-A antibody molecule; as shown in the results, Figure 17 the lung sections of the administration group showed significantly reduced lung fibrosis compared with the control group, thus indicating that the anti-IL-11 monoclonal antibody HA-I-A antibody molecule can effectively inhibit the generation of lung fibrosis.

[0222] Example 25 Therapeutic efficacy experiment of anti-IL-11 monoclonal antibody on cardiac fibrosis

[0223] The therapeutic effect of the anti-IL-11 monoclonal antibody HA-I-A on cardiac fibrosis was studied using isoproterenol modeling.

[0224] Animal species: C57BL / 6J mice (purchased from Jiangsu Jicui Yekang Biotechnology Co., Ltd.)

[0225] Number, gender and mouse age: 6 per group, male, 6-8 weeks;

[0226] The control group was only injected with normal saline;

[0227] The administration group was administered with the HA-I-A antibody molecule twice a week for 4 weeks.

[0228] The animal weight was measured once a week, and the animals were observed for abnormalities; organ weight detection: the heart was collected, the heart weight was measured, and the heart to body weight ratio was calculated; heart pathology detection: heart sections were made and hematoxylin-eosin (HE) and Masson staining were used to observe the degree of heart fibrosis.

[0229] As shown in the results, Figure 18 the heart to body weight ratio of the mice in the administration group was significantly smaller than that in the control group; as shown in the results, Figure 19 the heart sections of the administration group showed significantly reduced heart fibrosis compared with the control group, thus indicating that the anti-IL-11 monoclonal antibody HA-I-A antibody molecule can effectively inhibit the generation of heart fibrosis.

[0230] Example 26 Therapeutic efficacy experiment of anti-IL-11 monoclonal antibody on kidney fibrosis

[0231] To study the therapeutic effect of anti-IL-11 monoclonal antibody HA-I-A on renal fibrosis using adriamycin (dKF) modeling.

[0232] Animal species: BALB / c mice (purchased from Jiangsu Jucu Yaoke Biotechnology Co., Ltd.)

[0233] Number, gender and mouse age: 6 per group, male, 6-8 weeks;

[0234] The control group was only injected with normal saline.

[0235] The administration group was given two injections of HA-I-A antibody molecules per week for 4 weeks.

[0236] Animal body weight was measured once a week, and the animals were observed for abnormalities; organ weight detection: the heart was collected, the weight of the kidney was measured, and the urine protein content was detected; kidney pathology detection: kidney sections were stained with hematoxylin and eosin (HE) and Masson to observe the degree of renal fibrosis.

[0237] The results are shown in Figure 20 The results are shown in Figure 21 Compared with the control group, the administration group showed significantly reduced renal fibrosis in kidney sections, indicating that the anti-IL-11 monoclonal antibody HA-I-A antibody molecule can effectively inhibit the development of renal fibrosis.

[0238] Example 27 Therapeutic efficacy of anti-IL-11 monoclonal antibody on liver fibrosis

[0239] To study the therapeutic effect of anti-IL-11 monoclonal antibody HA-I-A on liver fibrosis using CCl4 modeling.

[0240] Animal species: C57BL / 6J mice (purchased from Jiangsu Jucu Yaoke Biotechnology Co., Ltd.)

[0241] Number, gender and mouse age: 6 per group, male, 6-8 weeks;

[0242] The control group was only injected with normal saline.

[0243] The administration group was given two injections of HA-I-A antibody molecules per week for 4 weeks.

[0244] Body weight monitoring: animal body weight was measured once a week, and the animals were observed for abnormalities; liver pathology detection: liver sections were stained with hematoxylin and eosin (HE) and Masson to observe the degree of liver fibrosis; organ weight detection: the kidney was collected, the weight of the liver was measured, and HE staining was performed; serum detection: serum was collected, and the alanine transaminase (ALT) and aspartate transaminase (AST) levels in the mouse serum were detected.

[0245] The results are as follows Figure 22 As shown, the liver weight of mice in the treatment group was significantly lower than that in the control group; the results are as follows. Figure 23 As shown, the serum ALT and AST levels in the treatment group were significantly lower than those in the control group; the results are as follows. Figure 24 As shown, compared with the control group, the liver slices of the treatment group showed a significant reduction in liver fibrosis, which indicates that the anti-IL-11 monoclonal antibody HA-IA antibody molecule can effectively inhibit the production of liver fibrosis.

[0246] Example 28: Thermostability Assessment of Anti-IL-11 Monoclonal Antibody HA-IA

[0247] The thermal stability of the anti-IL-11 monoclonal antibody HA-IA was assessed using a multifunctional protein thermal stability analysis system (purchased from Unchained Labs). Protein conformational changes were detected by monitoring intrinsic fluorescence as a function of temperature (starting at 25°C and increasing to 95°C at a rate of 0.3°C / min) to determine the protein melting temperature (Tm) and assess conformational stability. When samples aggregate, interference occurs in the scattered light waves, increasing the scattered light signal. The colloidal stability of the protein was determined by static light scattering (characterized using Tagg). The results are shown in the table below and appendix. Figure 25 As shown.

[0248]

[0249] The melting temperature (Tm) of the anti-IL-11 monoclonal antibody HA-IA was 79.5℃, and the average Tagg value was 75.9℃, indicating good conformational and colloidal stability.

[0250] This invention is not limited to the preferred embodiments described above. Anyone can derive other products in various forms under the guidance of this invention. However, regardless of any changes in shape or structure, any technical solution that is the same as or similar to this application falls within the protection scope of this invention.

Claims

1. An antibody or antigen-binding fragment thereof that binds IL-1 1, characterized in that, complementarity determining region HCDR1 has an amino acid sequence as set forth in SEQ ID No: 8, the heavy chain complementarity determining region HCDR2 has an amino acid sequence as set forth in SEQ ID No: 9, the heavy chain complementarity determining region HCDR3 has an amino acid sequence as set forth in SEQ ID No: 10, the light chain complementarity determining region LCDR1 has an amino acid sequence as set forth in SEQ ID No: 11, the light chain complementarity determining region LCDR2 has an amino acid sequence as set forth in SEQ ID No: 12, and the light chain complementarity determining region LCDR3 has an amino acid sequence as set forth in SEQ ID No:

13.

2. The antibody or antigen-binding fragment thereof that binds IL-1 1 of claim 1, wherein, complementarity determining region HCDR1 has an amino acid sequence as set forth in SEQ ID No: 8, the heavy chain complementarity determining region HCDR2 has an amino acid sequence as set forth in SEQ ID No: 9, the heavy chain complementarity determining region HCDR3 has an amino acid sequence as set forth in SEQ ID No: 10, the light chain complementarity determining region LCDR1 has an amino acid sequence as set forth in SEQ ID No: 11, the light chain complementarity determining region LCDR2 has an amino acid sequence as set forth in SEQ ID No: 12, and the light chain complementarity determining region LCDR3 has an amino acid sequence as set forth in SEQ ID No:

13.

3. The antibody or antigen-binding fragment thereof that binds IL-1 1 of claim 2, wherein, complementarity determining region HCDR1 has an amino acid sequence as set forth in SEQ ID No: 8, the heavy chain complementarity determining region HCDR2 has an amino acid sequence as set forth in SEQ ID No: 9, the heavy chain complementarity determining region HCDR3 has an amino acid sequence as set forth in SEQ ID No: 10, the light chain complementarity determining region LCDR1 has an amino acid sequence as set forth in SEQ ID No: 11, the light chain complementarity determining region LCDR2 has an amino acid sequence as set forth in SEQ ID No: 12, and the light chain complementarity determining region LCDR3 has an amino acid sequence as set forth in SEQ ID No:

13.

4. The antibody or antigen-binding fragment thereof that binds IL-1 1 of claim 2, wherein, The antibody or antigen-binding fragment thereof is a chimeric antibody molecule, and the chimeric antibody molecule further comprises a human antibody constant region.

5. The antibody or antigen-binding fragment thereof that binds IL-1 1 of claim 4, wherein, The human antibody constant region comprises a human antibody heavy chain constant region and a human antibody light chain constant region, wherein the amino acid sequence of the human antibody heavy chain constant region is one of SEQ ID No: 27, SEQ ID No: 28 or SEQ ID No: 29, and the amino acid sequence of the human antibody light chain constant region is as set forth in SEQ ID No:

30.

6. A nucleic acid molecule, characterized in that, The nucleic acid molecule encodes the antibody or antigen-binding fragment thereof that binds to IL-11 as claimed in claim 1.

7. A recombinant DNA expression vector, characterized by, The recombinant DNA expression vector comprises the nucleic acid molecule as claimed in claim 6.

8. A host cell transfected with the recombinant DNA expression vector of claim 7. The host cell comprises a prokaryotic cell, a yeast cell, an insect cell or a mammalian cell.

9. The host cell of claim 8, wherein, The host cell is a mammalian cell, and the mammalian cell is a HEK293 cell, a CHO cell or an NS0 cell.

10. A medicament, characterized by comprising: The medicament comprises the antibody or antigen-binding fragment thereof that binds to IL-11 as claimed in claim 1.

11. A combination drug composition, characterized in that, The combination composition comprises the antibody or antigen-binding fragment thereof that binds to IL-11 as claimed in claim 1, and further comprises a second composition selected from an antibody or antigen-binding fragment thereof that binds to PD-1, PD-L1, VEGF, VEGFR, TNF-α or TSLP.

12. Use of the antibody or antigen-binding fragment thereof that binds to IL-11 as claimed in claim 1 in the preparation of a medicament for treating a fibrotic disease, inflammation or cancer in a human. the fibrotic disease is selected from fibrosis of the heart, liver, kidney, lung, bone marrow, eye, pancreas, spleen, brain, intestinal tract, or skin; the inflammation is selected from pancreatitis, enteritis, arthritis, or dermatitis; the cancer is selected from lung cancer, gastric cancer, breast cancer, or colorectal cancer.

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