Antigen-binding protein targeting il-11 and use thereof

Abstract

The present application relates to an antigen-binding protein targeting IL-11 and a use thereof, and in particular to an isolated antigen-binding protein capable of binding to IL-11. The present application further provides a preparation method for and a use of the isolated antigen-binding protein.

Description

Antigen binding proteins targeting IL-11 and uses thereof TECHNICAL FIELD

[0001] The present application relates to the field of biological medicine, in particular to an isolated antigen binding protein targeting IL-11 and uses thereof. BACKGROUND

[0002] Interleukin-11 is a stromal cell-derived cytokine belonging to the IL-6 superfamily, which is secreted by various types of cells such as osteoblasts, synoviocytes, fibroblasts, chondrocytes, and trophoblasts. It can activate downstream JAK / STAT signaling pathway, ERK signaling pathway and MAPK signaling pathway by binding to IL-11R alpha and GP130 receptors.

[0003] More and more studies have found that IL-11 plays an extremely important role in cell protection, anti-inflammation and anti-fibrosis of organs such as lungs. IL-11 is secreted by polarized cells or fibroblasts after injury, and has autocrine and paracrine effects. In polarized cells, IL-11 causes cell dysfunction, initiates apoptotic cell death, and at the same time blocks regeneration. In stromal cells, IL-11 triggers extracellular matrix production, invasion and migration of myofibroblasts. IL-11-activated fibroblasts and myofibroblasts secrete cytokines and chemokines, which have strong pro-inflammatory effects. Inhibition of IL-11 can protect parenchyma, anti-fibrosis, and reduce stromal-driven inflammation. In tissues with regenerative capacity, inhibition of IL-11 can promote the proliferation and regeneration of damaged cells (such as hepatocytes) and organ regeneration.

[0004] In early 2020, Boehringer Ingelheim announced the acquisition of the global exclusive rights to Enleofen's preclinical IL-11 platform to develop a monoclonal antibody drug targeting the IL-11 / IL-11Ra pathway to develop a first-in-class therapy for the treatment of various fibro-inflammatory diseases. LASN01, a monoclonal antibody drug targeting IL-11Ra developed by Lassen Therapeutics, is expected to be a new treatment for fibrosis and tumor cancer, and submitted a new drug research application to the FDA in 2021 to start a phase I clinical trial. In China, the IL-11 monoclonal antibody (9MW3811) developed by Maiwei Biotechnology is in the forefront, and submitted a clinical trial application to the FDA in February 2023, and is currently in the clinical phase I stage. SUMMARY

[0005] The present application provides an isolated antigen binding protein capable of binding to IL-11, which comprises a HCDR3 comprising an amino acid sequence as set forth in SEQ ID NO: 3.

[0006] In certain embodiments, the antigen binding protein comprises a HCDR2 comprising an amino acid sequence as set forth in SEQ ID NO: 2.

[0007] In certain embodiments, the antigen binding protein comprises a HCDR1 comprising an amino acid sequence as set forth in SEQ ID NO: 1.

[0008] In certain embodiments, the antigen binding protein comprises a LCDR3 comprising an amino acid sequence as set forth in SEQ ID NO: 6.

[0009] In certain embodiments, the antigen binding protein comprises a LCDR2 comprising an amino acid sequence as set forth in SEQ ID NO: 5.

[0010] In certain embodiments, the antigen binding protein comprises a LCDR1 comprising an amino acid sequence as set forth in SEQ ID NO: 4.

[0011] In certain embodiments, the antigen binding protein comprises a HCDR1, a HCDR2, and a HCDR3, wherein the HCDR1 comprises an amino acid sequence as set forth in SEQ ID NO: 1, the HCDR2 comprises an amino acid sequence as set forth in SEQ ID NO: 2, and the HCDR3 comprises an amino acid sequence as set forth in SEQ ID NO: 3.

[0012] In certain embodiments, the antigen binding protein comprises a LCDR1, a LCDR2, and a LCDR3, wherein the LCDR1 comprises an amino acid sequence as set forth in SEQ ID NO: 4, the LCDR2 comprises an amino acid sequence as set forth in SEQ ID NO: 5, and the LCDR3 comprises an amino acid sequence as set forth in SEQ ID NO: 6.

[0013] In certain embodiments, the antigen binding protein comprises a HCDR1, a HCDR2, a HCDR3, a LCDR1, a LCDR2, and a LCDR3, wherein the HCDR1 comprises an amino acid sequence as set forth in SEQ ID NO: 1, the HCDR2 comprises an amino acid sequence as set forth in SEQ ID NO: 2, the HCDR3 comprises an amino acid sequence as set forth in SEQ ID NO: 3, the LCDR1 comprises an amino acid sequence as set forth in SEQ ID NO: 4, the LCDR2 comprises an amino acid sequence as set forth in SEQ ID NO: 5, and the LCDR3 comprises an amino acid sequence as set forth in SEQ ID NO: 6.

[0014] In certain embodiments, the antigen binding protein comprises a H-FR1, a H-FR2, a H-FR3, and a H-FR4, wherein the H-FR1 comprises an amino acid sequence as set forth in SEQ ID NO: 30, the H-FR2 comprises an amino acid sequence as set forth in SEQ ID NO: 31, the H-FR3 comprises an amino acid sequence as set forth in SEQ ID NO: 32, and the H-FR4 comprises an amino acid sequence as set forth in SEQ ID NO: 33.

[0015] In certain embodiments, the antigen binding protein comprises a H-FR1, a H-FR2, a H-FR3, and a H-FR4, wherein the H-FR1 comprises an amino acid sequence as set forth in SEQ ID NO: 7 or SEQ ID NO: 11, the H-FR2 comprises an amino acid sequence as set forth in SEQ ID NO: 8 or SEQ ID NO: 12, the H-FR3 comprises an amino acid sequence as set forth in SEQ ID NO: 9, SEQ ID NO: 13, or SEQ ID NO: 15, and the H-FR4 comprises an amino acid sequence as set forth in SEQ ID NO: 10 or SEQ ID NO: 14.

[0016] In certain embodiments, the antigen binding protein comprises a H-FR1, a H-FR2, a H-FR3, and a H-FR4, wherein: a) the H-FR1 comprises an amino acid sequence as set forth in SEQ ID NO: 7, the H-FR2 comprises an amino acid sequence as set forth in SEQ ID NO: 8, the H-FR3 comprises an amino acid sequence as set forth in SEQ ID NO: 9, and the H-FR4 comprises an amino acid sequence as set forth in SEQ ID NO: 10; b) the H-FR1 comprises an amino acid sequence as set forth in SEQ ID NO: 11, the H-FR2 comprises an amino acid sequence as set forth in SEQ ID NO: 12, the H-FR3 comprises an amino acid sequence as set forth in SEQ ID NO: 13, and the H-FR4 comprises an amino acid sequence as set forth in SEQ ID NO: 14; or c) the H-FR1 comprises an amino acid sequence as set forth in SEQ ID NO: 11, the H-FR2 comprises an amino acid sequence as set forth in SEQ ID NO: 12, the H-FR3 comprises an amino acid sequence as set forth in SEQ ID NO: 15, and the H-FR4 comprises an amino acid sequence as set forth in SEQ ID NO: 14.

[0017] In certain embodiments, the antigen binding protein comprises a heavy chain variable region VH, wherein the VH comprises an amino acid sequence as set forth in SEQ ID NO: 24, SEQ ID NO: 25, or SEQ ID NO: 26.

[0018] In certain embodiments, the antigen binding protein further comprises a heavy chain constant region. In certain embodiments, the heavy chain constant region is an IgG heavy chain constant region. In certain embodiments, the heavy chain constant region is a human IgG heavy chain constant region.

[0019] In certain embodiments, the antigen binding protein comprises L-FR1, L-FR2, L-FR3, and L-FR4, wherein the L-FR1 comprises an amino acid sequence as set forth in SEQ ID NO: 34, the L-FR2 comprises an amino acid sequence as set forth in SEQ ID NO: 17, the L-FR3 comprises an amino acid sequence as set forth in SEQ ID NO: 35, and the L-FR4 comprises an amino acid sequence as set forth in SEQ ID NO: 36.

[0020] In certain embodiments, the antigen binding protein comprises L-FR1, L-FR2, L-FR3, and L-FR4, wherein the L-FR1 comprises an amino acid sequence as set forth in SEQ ID NO: 16 or SEQ ID NO: 20, the L-FR2 comprises an amino acid sequence as set forth in SEQ ID NO: 17, the L-FR3 comprises an amino acid sequence as set forth in SEQ ID NO: 18, SEQ ID NO: 21, or SEQ ID NO: 23, and the L-FR4 comprises an amino acid sequence as set forth in SEQ ID NO: 19 or SEQ ID NO: 22.

[0021] In certain embodiments, the antigen binding protein comprises L-FR1, L-FR2, L-FR3, and L-FR4, wherein: a) the L-FR1 comprises an amino acid sequence as set forth in SEQ ID NO: 16, the L-FR2 comprises an amino acid sequence as set forth in SEQ ID NO: 17, the L-FR3 comprises an amino acid sequence as set forth in SEQ ID NO: 18, and L-FR4 comprises an amino acid sequence as set forth in SEQ ID NO: 19; b) the L-FR1 comprises an amino acid sequence as set forth in SEQ ID NO: 20, the L-FR2 comprises an amino acid sequence as set forth in SEQ ID NO: 17, the L-FR3 comprises an amino acid sequence as set forth in SEQ ID NO: 21, and L-FR4 comprises an amino acid sequence as set forth in SEQ ID NO: 22; or c) the L-FR1 comprises an amino acid sequence as set forth in SEQ ID NO: 20, the L-FR2 comprises an amino acid sequence as set forth in SEQ ID NO: 17, the L-FR3 comprises an amino acid sequence as set forth in SEQ ID NO: 23, and L-FR4 comprises an amino acid sequence as set forth in SEQ ID NO: 22.

[0022] In certain embodiments, the antigen binding protein comprises a light chain variable region VL, which comprises an amino acid sequence as set forth in SEQ ID NO: 27, SEQ ID NO: 28, or SEQ ID NO: 29.

[0023] In certain embodiments, the antigen binding protein comprises a light chain constant region. In certain embodiments, the light chain constant region is an Ig kappa light chain constant region. In certain embodiments, the light chain constant region is a human Ig kappa light chain constant region.

[0024] In certain embodiments, the antigen binding protein comprises a VH and a VL, wherein: a) the VH comprises an amino acid sequence as set forth in SEQ ID NO: 24, and the VL comprises an amino acid sequence as set forth in SEQ ID NO: 27; b) the VH comprises an amino acid sequence as set forth in SEQ ID NO: 25, and the VL comprises an amino acid sequence as set forth in SEQ ID NO: 28; c) the VH comprises an amino acid sequence as set forth in SEQ ID NO: 25, and the VL comprises an amino acid sequence as set forth in SEQ ID NO: 29; d) the VH comprises an amino acid sequence as set forth in SEQ ID NO: 26, and the VL comprises an amino acid sequence as set forth in SEQ ID NO: 28; or e) the VH comprises an amino acid sequence as set forth in SEQ ID NO: 26, and the VL comprises an amino acid sequence as set forth in SEQ ID NO: 29.

[0025] In certain embodiments, the antigen binding protein comprises an antibody or an antigen binding fragment thereof. In certain embodiments, the antibody is selected from a monoclonal antibody, a chimeric antibody, a humanized antibody, or a fully human antibody. In certain embodiments, the antigen binding fragment is a Fab, a Fab’, a F(ab)2, a Fv fragment, a F(ab’)2, a scFv, a di-scFv, a VHH, and / or a dAb.

[0026] In another aspect, the present application also provides a fusion protein comprising the antigen binding protein.

[0027] In another aspect, the present application also provides a bispecific antibody comprising a first portion and a second portion, wherein the first portion targets IL-11, and the first portion comprises the antigen binding protein.

[0028] In certain embodiments, the first targeting portion comprises the antigen binding protein.

[0029] In another aspect, the present application also provides an immunoconjugate comprising the antigen binding protein, the fusion protein, or the bispecific antibody.

[0030] In another aspect, the present application also provides one or more isolated nucleic acid molecules encoding the antigen binding protein, the fusion protein, the bispecific antibody, and / or the immunoconjugate.

[0031] In another aspect, the present application also provides a vector comprising the nucleic acid molecule.

[0032] In another aspect, the present application also provides a cell comprising and / or expressing the antigen binding protein, the fusion protein, the bispecific antibody, the immunoconjugate, the nucleic acid molecule, and / or the vector.

[0033] In another aspect, the present application also provides a pharmaceutical composition comprising the antigen binding protein, the fusion protein, the bispecific antibody, the immunoconjugate, the nucleic acid molecule, the vector, and / or the cell and optionally a pharmaceutically acceptable carrier.

[0034] In another aspect, the present application provides use of the antigen binding protein, the fusion protein, the bispecific antibody, the immunoconjugate, the nucleic acid molecule, the vector, the cell, and / or the pharmaceutical composition in the manufacture of a medicament for preventing and / or treating a disease and / or a disorder.

[0035] In certain embodiments, the disease and / or disorder comprises a disease and / or disorder associated with IL-11 expression.

[0036] In certain embodiments, the disease and / or disorder comprises an inflammatory disease or a tumor.

[0037] In certain embodiments, the disease and / or disorder comprises pulmonary fibrosis, renal fibrosis, hepatic fibrosis, myocardial fibrosis, splenic fibrosis, arthritis, ankylosing spondylitis, gastritis, pneumonitis, hepatitis, colitis, lung cancer, stomach cancer, bone cancer, colon cancer, and / or liver cancer.

[0038] In another aspect, the present application provides a method of producing the antigen binding protein, the fusion protein, and / or the bispecific antibody. The method comprises culturing the cell under conditions such that the antigen binding protein, the fusion protein, the bispecific antibody is expressed.

[0039] In another aspect, the present application provides a kit comprising the antigen binding protein, the fusion protein, the bispecific antibody, the immunoconjugate, the nucleic acid molecule, the vector, the cell, and / or the pharmaceutical composition.

[0040] In another aspect, the present application provides a method for preventing and / or treating a disease and / or a disorder in a subject in need thereof, comprising administering to the subject an effective amount of the antigen binding protein, the fusion protein, the bispecific antibody, the immunoconjugate, the nucleic acid molecule, the vector, the cell, and / or the pharmaceutical composition of claim 28.

[0041] In certain embodiments, the disease and / or disorder comprises a disease and / or disorder associated with IL-11 expression.

[0042] In certain embodiments, the disease and / or disorder comprises an inflammatory disease or a tumor.

[0043] In certain embodiments, the disease and / or disorder comprises pulmonary fibrosis, renal fibrosis, liver fibrosis, myocardial fibrosis, splenic fibrosis, arthritis, ankylosing spondylitis, gastritis, pneumonitis, hepatitis, colitis, lung cancer, stomach cancer, bone cancer, colon cancer, and / or liver cancer.

[0044] In another aspect, the present application provides the antigen binding protein, the fusion protein, the bispecific antibody, the immunoconjugate, the nucleic acid molecule, the vector, the cell, and / or the pharmaceutical composition for use in preventing and / or treating a disease and / or a disorder.

[0045] In certain embodiments, the disease and / or disorder comprises a disease and / or disorder associated with IL-11 expression.

[0046] In certain embodiments, the disease and / or disorder comprises an inflammatory disease or a tumor.

[0047] In certain embodiments, the disease and / or disorder comprises pulmonary fibrosis, renal fibrosis, liver fibrosis, myocardial fibrosis, splenic fibrosis, arthritis, ankylosing spondylitis, gastritis, pneumonitis, hepatitis, colitis, lung cancer, stomach cancer, bone cancer, colon cancer, and / or liver cancer.

[0048] The antigen binding protein described in the present application has the following advantages: (1) has very strong affinity to human, murine and monkey IL-11, (2) can block IL-11 mediated activation of hIL11 Effector Reporter Cell, (3) blocks the formation of IL-11 / IL-11Ra / GP130 ternary complex, (4) can inhibit TGFβ-induced fibrosis of NICH0018 cells in vitro, and (5) inhibits renal fibrosis in chronic kidney injury mice.

[0049] Other aspects and advantages of the present application will become apparent from the following detailed description, taken in conjunction with the accompanying drawings. The detailed description merely illustrates exemplary embodiments of the present application. It will be readily apparent to those skilled in the art that modifications can be made to the specific embodiments disclosed without departing from the spirit and scope of the application. Accordingly, the drawings and descriptions are to be regarded as illustrative in nature, and not as restrictive. BRIEF DESCRIPTION OF DRAWINGS

[0050] The specific features of the application involved in the present application are shown in the appended claims. The features and advantages of the application involved in the present application can be better understood by referring to the detailed description of exemplary embodiments and the accompanying drawings. The drawings are briefly described as follows:

[0051] Figure 1 shows the IL-11 antibody described in the present application inhibits the activation effect curve of hIL-11 Effector Reporter Cell mediated by IL-11.

[0052] Figure 2 shows the blocking effect curve of the IL-11 antibody described in the present application inhibiting the formation of IL-11 / IL-11Ra / GP130 ternary complex.

[0053] Figure 3 shows the PCR results of the IL-11 antibody described in the present application detecting the ability of NICH0018 (healthy human skin fibroblasts) to inhibit TGFβ-induced fibrosis.

[0054] Figure 4 shows the immunofluorescence results of the IL-11 antibody described in the present application detecting the ability of NICH0018 (healthy human skin fibroblasts) to inhibit TGFβ-induced fibrosis.

[0055] Figure 5 shows the staining quantification results of the IL-11 antibody described in the present application detecting the ability of the IL-11 antibody to inhibit renal fibrosis in mice with chronic kidney injury in vivo. DETAILED DESCRIPTION

[0056] The embodiments of the present application are illustrated by the following specific examples, and other advantages and effects of the present application can be easily understood by those skilled in the art from the disclosure of the present specification.

[0057] TERMS

[0058] In the present application, the term "IL-11" generally refers to a stromal cell-derived cytokine belonging to the IL-6 superfamily. The "IL-11" encompasses "full-length", unprocessed IL-11 and any form of IL-11 produced by cellular processing. In the present application, the term "IL-11" includes mutants, fragments, variants, isoforms and homologues thereof.

[0059] The term "disease and / or disorder associated with IL-11 expression" refers to a disease and / or disorder caused by the activation of JAK / STAT and / or ERK signaling pathway after IL-11 expression, which is manifested as inflammatory disease and / or tumor cancer. The inflammatory disease includes inflammation, fibrotic process, and autoimmune disease. The fibrotic process refers to a pathological process in which functional cells of an organ are necrotic. The organ includes heart, lung, liver, spleen, kidney, pancreas, eye, etc. The tumor cancer includes solid tumor and blood tumor. The solid tumor includes gastric cancer, colorectal cancer, breast cancer, bone cancer, gastrointestinal tumor, etc. The blood tumor includes leukemia, aplastic anemia, thrombocytopenia, etc.

[0060] The term "isolated" generally refers to an artificially obtained state. If a certain "isolated" substance or component appears in nature, it may be that the natural environment has changed, or the substance has been separated from the natural environment, or both. For example, a certain polynucleotide or polypeptide naturally exists in a certain living animal without being isolated, and a high-purity same polynucleotide or polypeptide separated from such a natural state is called isolated. The term "isolated" does not exclude the mixing of artificial or synthetic substances, nor does it exclude the presence of other impurities that do not affect the activity of the substance.

[0061] The term "isolated antigen binding protein" generally refers to a protein with antigen binding ability obtained by artificial means from the natural state. The "isolated antigen binding protein" can include an antigen binding portion and, optionally, a framework or framework portion that allows the antigen binding portion to adopt a conformation that facilitates binding of the antigen binding portion to an antigen. The antigen binding protein can include, for example, an antibody-derived protein framework region (FR) or an alternative protein framework region or an artificial framework region with grafted CDRs or CDR derivatives. Such frameworks include, but are not limited to, antibody-derived framework regions comprising mutations introduced, for example, to stabilize the three-dimensional structure of the antigen binding protein, and completely synthetic framework regions comprising, for example, biocompatible polymers. Examples of antigen binding proteins include, but are not limited to: human antibodies, humanized antibodies; chimeric antibodies; recombinant antibodies; single-chain antibodies; bifunctional antibodies; trifunctional antibodies; tetrafunctional antibodies; Fab, Fab', Fv fragments, F(ab')2, F(ab)2, scFv, di-scFv, dAb, IgD antibodies; IgE antibodies; IgM antibodies; IgG1 antibodies; IgG2 antibodies; IgG3 antibodies; or IgG4 antibodies and fragments thereof. The antigen binding proteins described herein can also include, for example, antibody fusion proteins having an antigen recognition site that binds IL-11, for example, the antigen recognition site can include CDRs. The antigen binding proteins described herein can also include chimeric antigen receptors, which can have an extracellular structure and an intracellular structure, the extracellular structure can recognize IL-11, for example, can include CDRs.

[0062] The term "CDR" also known as "complementarity determining region" generally refers to a region in an antibody variable domain whose sequence is highly variable and / or forms a structurally defining loop. Typically, an antibody includes six CDRs: three in VH (HCDR1, HCDR2, HCDR3), and three in VL (LCDR1, LCDR2, LCDR3). In certain embodiments, naturally occurring camelid antibodies consisting of only heavy chains are able to function normally and stably in the absence of light chains. Antibody CDRs can be determined by various encoding systems, such as CCG, Kabat, Chothia, IMGT, integrated consideration of Kabat / Chothia, etc. These encoding systems are known in the art. For example, the amino acid sequence numbering of the antigen binding protein can be divided according to the IMGT numbering system. For example, the CDRs of the antigen binding protein can be divided according to the Kabat numbering system.

[0063] The term "FR" generally refers to a more highly conserved portion of an antibody variable domain, which is referred to as a framework region. Typically, the variable domains of the heavy and light chains of a native antibody each comprise four FR regions, i.e., four in VH (H-FR1, H-FR2, H-FR3 and H-FR4), and four in VL (L-FR1, L-FR2, L-FR3 and L-FR4).

[0064] The term "fusion protein" generally refers to a polypeptide having at least two moieties covalently linked together. Each of the moieties can be a polypeptide having different properties. The property can be a biological property, such as an activity in vitro or in vivo. The property can also be a simple chemical or physical property, such as binding to a target molecule, catalysis of a reaction, etc. The two moieties can be linked directly by a single peptide bond or through a peptide linker.

[0065] The term "bispecific antibody" generally refers to an antibody that is capable of having binding specificity to at least two antigens, which can comprise two or more antibodies, antigen binding fragments (such as scFv), derivatives thereof, or antigen binding regions linked together. The binding specificity comprises a binding specificity to an Fc receptor on an effector cell and a binding specificity to an antigen or epitope on a target cell, such as a tumor cell. The bispecific antibodies described herein can specifically bind to IL-11 and a target associated with inflammatory diseases, such as TSLP, IL-36R, IL-4R, or IL-17.

[0066] The term "nucleic acid molecule" generally refers to an isolated form of nucleotide, deoxyribonucleotide or ribonucleotide of any length, or an analog thereof, either in isolation or in a natural environment.

[0067] The term "vector" generally refers to a nucleic acid vehicle into which a polynucleotide encoding a protein can be inserted and expressed. The vector can express the genetic material elements it carries by transforming, transducing or transfecting a host cell, so that the expression of the genetic material elements carried by the vector is enabled in the host cell. For example, the vector can include a plasmid; a phagemid; a cosmid; an artificial chromosome such as a yeast artificial chromosome (YAC), a bacterial artificial chromosome (BAC) or a P1 -derived artificial chromosome (PAC); a bacteriophage such as a lambda phage or a M13 phage; and an animal virus, etc. The types of animal viruses that can be used as vectors can include a retrovirus (including a lentivirus), an adenovirus, an adeno-associated virus, a herpes virus (such as a herpes simplex virus), a poxvirus, a baculovirus, a papillomavirus, a papovavirus (such as SV40). A vector can contain various elements for controlling expression, including a promoter sequence, a transcription initiation sequence, an enhancer sequence, a selection element, and a reporter gene. In addition, the vector can contain a replication initiation site. The vector can also include components that facilitate its entry into a cell, such as a viral particle, a liposome, or a protein coat, but not only these.

[0068] The term "cell" generally refers to an individual cell, a cell line, or a cell culture that can be or has been the recipient of a plasmid or vector of a subject, which includes a nucleic acid molecule of the present application or a vector of the present application. The cell can include the progeny of the subject individual cell. The progeny can not necessarily be completely identical (in morphology or in genomic

[0069] The term "immunoconjugate" generally refers to a conjugate formed by conjugating (e.g., covalently linking via a linking molecule) the other agent (e.g., a chemotherapeutic agent, a radioactive element, a cytostatic and a cytotoxic agent) to the antibody or antigen-binding fragment thereof, which can deliver the other agent to a target cell (e.g., a tumor cell) via the specific binding of the antibody or antigen-binding fragment thereof to an antigen on the target cell.

[0070] The term "pharmaceutical composition" generally refers to a composition for preventing / treating a disease or a disorder. The pharmaceutical composition can comprise an isolated antigen binding protein of the present application, a bispecific antibody of the present application, a fusion protein of the present application, an immunoconjugate of the present application, a nucleic acid molecule of the present application, a vector of the present application, and / or a cell of the present application, and optionally a pharmaceutically acceptable adjuvant. In addition, the pharmaceutical composition can further comprise a suitable formulation of one or more (pharmaceutically effective) carriers, stabilizers, excipients, diluents, solubilizers, surfactants, emulsifiers, and / or preservatives. Acceptable ingredients of the composition are preferably non-toxic to the recipient at the doses and concentrations employed. Pharmaceutical compositions of the present application include, but are not limited to, liquid, frozen, and lyophilized compositions.

[0071] The term "pharmaceutically acceptable carrier" generally includes a pharmaceutically acceptable carrier, excipient, or stabilizer, which is nontoxic to the cells or mammals being exposed thereto at the dosages and concentrations employed. Physiologically acceptable carriers can include, for example, buffers, antioxidants, low molecular weight (less than about 10 residues) polypeptides, proteins, hydrophilic polymers, amino acids, monosaccharides, disaccharides and other carbohydrates, chelating agents, sugar alcohols, salt-forming counterions, such as sodium; and / or nonionic surfactants.

[0072] The term "subject" generally refers to a human or non-human animal, including, but not limited to, a cat, dog, horse, pig, cow, sheep, rabbit, mouse, rat, or monkey.

[0073] In the present application, the protein, polypeptide, and / or amino acid sequence referred to should also be understood to encompass at least variants or homologues of the protein or polypeptide having the same or similar function.

[0074] In the present application, the variant can be, for example, a protein or polypeptide having one or more amino acids substituted, deleted, or added in the amino acid sequence of the protein and / or the polypeptide (e.g., an antibody or fragment thereof that specifically binds to an IL-11 protein). For example, the functional variant can include a protein or polypeptide having an amino acid change by at least 1, for example, 1-30, 1-20, or 1-10, and further, for example, 1, 2, 3, 4, or 5 amino acid substitutions, deletions, and / or insertions. The functional variant can substantially maintain the biological properties of the protein or the polypeptide before the change (e.g., substitution, deletion, or addition). For example, the functional variant can maintain at least 60%, 70%, 80%, 90%, or 100% of the biological activity (e.g., antigen binding ability) of the protein or the polypeptide before the change. For example, the substitution can be a conservative substitution.

[0075] The homologue can be a protein or polypeptide having at least about 85% (e.g., at least about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or more) sequence homology to the amino acid sequence of the protein and / or the polypeptide (e.g., an antibody or fragment thereof that specifically binds to an IL-11 protein).

[0076] In the present application, the homology generally refers to similarity, analogy, or association between two or more sequences. The "percent sequence homology" can be calculated by comparing two sequences to be aligned in a comparison window, determining the number of positions in the two sequences in which the same nucleic acid base (e.g., A, T, C, G, I) or the same amino acid residue (e.g., Ala, Pro, Ser, Thr, Gly, Val, Leu, Ile, Phe, Tyr, Trp, Lys, Arg, His, Asp, Glu, Asn, Gin, Cys, or Met) is present to obtain the number of matching positions, dividing the number of matching positions by the total number of positions in the comparison window (i.e., window size), and multiplying the result by 100 to produce the percent sequence homology. The alignment for determining the percent sequence homology can be achieved in various ways known in the art.

[0077] In the present application, the term "comprising" generally means the inclusion without numerical limits of expressly listed items, as well as items which are generally equivalent to the expressly listed items. In certain instances, "comprising" will also mean "consisting of" or "consisting essentially of" in some embodiments.

[0078] In the present application, the term "about" generally means a range of plus or minus 0.5-10% from a specified value, for example, a range of plus or minus 0.5%, 1%, 1.5%, 2%, 2.5%, 3%, 3.5%, 4%, 4.5%, 5%, 5.5%, 6%, 6.5%, 7%, 7.5%, 8%, 8.5%, 9%, 9.5%, or 10% from the specified value.

[0079] DETAILED DESCRIPTION

[0080] A. Isolated antigen binding protein

[0081] In one aspect, the present application provides an isolated antigen binding protein that can bind IL-11, which can include an antibody or an antigen binding fragment thereof. The antigen binding fragment can include a Fab, Fab', F(ab)2, Fv fragment, F(ab')2, scFv, di-scFv, VHH, and / or dAb. The antibody can include a monoclonal antibody, a chimeric antibody, a humanized antibody, and a fully human antibody.

[0082] A1. CDR

[0083] The CDRs of an antibody, also known as complementarity determining regions, are portions of the variable region. The amino acid residues of this region can contact the antigen or epitope. Antibody CDRs can be determined by various numbering systems, such as CCG, Kabat, Chothia, IMGT, AbM, integrated consideration of Kabat / Chothia, etc. These numbering systems are known in the art, and can be found, for example, at http: / / www.bioinf.org.uk / abs / index.html#kabatnum. One of skill in the art can determine the CDR regions using different numbering systems based on the sequence and structure of the antibody. The CDR regions can differ using different numbering systems. In the present application, the CDRs encompass the CDR sequences determined according to any CDR partitioning scheme, and the CDRs also encompass variants thereof. The variants include substitution, deletion, and / or addition of one or more amino acids of the CDR. For example, 1-30, 1-20, or 1-10 amino acid substitutions, deletions, and / or insertions; also, for example, 1, 2, 3, 4, 5, 6, 7, 8, or 9 amino acid substitutions, deletions, and / or insertions; and also encompass homologues thereof, which can be an amino acid sequence having at least about 85% (e.g., at least about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or more) sequence homology to the amino acid sequence of the CDR. In the present application, the isolated antigen binding protein is defined by the IMGT numbering system.

[0084] In the present application, the antigen binding protein can comprise at least one CDR in the VH, which comprises an amino acid sequence as set forth in SEQ ID NO: 24, SEQ ID NO: 25, or SEQ ID NO: 26.

[0085] In the present application, the antigen binding protein can comprise a HCDR3, which comprises an amino acid sequence as set forth in SEQ ID NO: 3.

[0086] In the present application, the antigen binding protein can comprise a HCDR2, which comprises an amino acid sequence as set forth in SEQ ID NO: 2.

[0087] In the present application, the antigen binding protein can comprise a HCDR1, which comprises an amino acid sequence as set forth in SEQ ID NO: 1.

[0088] In the present application, the antigen binding protein can comprise a LCDR3, which comprises an amino acid sequence as set forth in SEQ ID NO: 6.

[0089] In the present application, the antigen binding protein can comprise a LCDR2 comprising an amino acid sequence as set forth in SEQ ID NO: 5.

[0090] In the present application, the antigen binding protein can comprise a LCDR1 comprising an amino acid sequence as set forth in SEQ ID NO: 4.

[0091] In the present application, the antigen binding protein can comprise a HCDR1 comprising an amino acid sequence as set forth in SEQ ID NO: 1, a HCDR2 comprising an amino acid sequence as set forth in SEQ ID NO: 2, and a HCDR3 comprising an amino acid sequence as set forth in SEQ ID NO: 3.

[0092] In the present application, the antigen binding protein can comprise a LCDR1 comprising an amino acid sequence as set forth in SEQ ID NO: 4, a LCDR2 comprising an amino acid sequence as set forth in SEQ ID NO: 5, and a LCDR3 comprising an amino acid sequence as set forth in SEQ ID NO: 6.

[0093] In the present application, the antigen binding protein can comprise a HCDR1 comprising an amino acid sequence as set forth in SEQ ID NO: 1, a HCDR2 comprising an amino acid sequence as set forth in SEQ ID NO: 2, a HCDR3 comprising an amino acid sequence as set forth in SEQ ID NO: 3, a LCDR1 comprising an amino acid sequence as set forth in SEQ ID NO: 4, a LCDR2 comprising an amino acid sequence as set forth in SEQ ID NO: 5, and a LCDR3 comprising an amino acid sequence as set forth in SEQ ID NO: 6.

[0094] A2.FR

[0095] In the present application, antibody framework regions FR refer to the portions of an antibody variable region that are present between the more diverse (i.e., hypervariable) CDRs. Such framework regions are typically referred to as framework 1 through 4 (FR1, FR2, FR3, and FR4) and provide a scaffold for presenting the six CDRs (three from the heavy chain and three from the light chain) in three-dimensional space to form an antigen binding surface.

[0096] In the present application, the isolated antigen binding protein can comprise a H-FR1, the C-terminus of which is directly or indirectly connected to the N-terminus of the HCDR1, and the H-FR1 comprises an amino acid sequence as set forth in SEQ ID NO: 30.

[0097] In the present application, the isolated antigen binding protein can comprise an H-FR2, which is located between the HCDR1 and the HCDR2, and the H-FR2 comprises an amino acid sequence as set forth in SEQ ID NO: 31.

[0098] In the present application, the isolated antigen binding protein can comprise an H-FR3, which is located between the HCDR2 and the HCDR3, and the H-FR3 comprises an amino acid sequence as set forth in SEQ ID NO: 32.

[0099] In the present application, the isolated antigen binding protein can comprise an H-FR4, which is located between the HCDR3 and the HCDR4, and the H-FR4 comprises an amino acid sequence as set forth in SEQ ID NO: 33.

[0100] In the present application, the isolated antigen binding protein can comprise an H-FR1, an H-FR2, an H-FR3 and an H-FR4. Wherein the H-FR1 comprises an amino acid sequence as set forth in SEQ ID NO: 30, the H-FR2 comprises an amino acid sequence as set forth in SEQ ID NO: 31, the H-FR3 comprises an amino acid sequence as set forth in SEQ ID NO: 32, and the H-FR4 comprises an amino acid sequence as set forth in SEQ ID NO: 33.

[0101] In the present application, the isolated antigen binding protein can comprise an H-FR1, which is located between the HCDR1 and the HCDR2, and the H-FR1 comprises an amino acid sequence as set forth in SEQ ID NO: 7 or SEQ ID NO: 11.

[0102] In the present application, the isolated antigen binding protein can comprise an H-FR2, which is located between the HCDR1 and the HCDR2, and the H-FR2 comprises an amino acid sequence as set forth in SEQ ID NO: 8 or SEQ ID NO: 12.

[0103] In the present application, the isolated antigen binding protein can comprise an H-FR3, which is located between the HCDR2 and the HCDR3, and the H-FR3 comprises an amino acid sequence as set forth in SEQ ID NO: 9, SEQ ID NO: 13 or SEQ ID NO: 15.

[0104] In the present application, the isolated antigen binding protein can comprise an H-FR4, the N-terminus of which is directly or indirectly connected to the C-terminus of the HCDR3, and the H-FR4 comprises an amino acid sequence as set forth in SEQ ID NO: 10 or SEQ ID NO: 14.

[0105] In the present application, the isolated antigen binding protein can comprise an H-FR1, an H-FR2, an H-FR3 and an H-FR4, wherein the H-FR1 comprises an amino acid sequence as set forth in SEQ ID NO: 7 or SEQ ID NO: 11, the H-FR2 comprises an amino acid sequence as set forth in SEQ ID NO: 8 or SEQ ID NO: 12, the H-FR3 comprises an amino acid sequence as set forth in SEQ ID NO: 9, SEQ ID NO: 13 or SEQ ID NO: 15, and the H-FR4 comprises an amino acid sequence as set forth in SEQ ID NO: 10 or SEQ ID NO: 14.

[0106] In the present application, the isolated antigen binding protein can comprise an H-FR1, an H-FR2, an H-FR3 and an H-FR4, wherein: a) the H-FR1 comprises an amino acid sequence as set forth in SEQ ID NO: 7, the H-FR2 comprises an amino acid sequence as set forth in SEQ ID NO: 8, the H-FR3 comprises an amino acid sequence as set forth in SEQ ID NO: 9, and the H-FR4 comprises an amino acid sequence as set forth in SEQ ID NO: 10; b) the H-FR1 comprises an amino acid sequence as set forth in SEQ ID NO: 11, the H-FR2 comprises an amino acid sequence as set forth in SEQ ID NO: 12, the H-FR3 comprises an amino acid sequence as set forth in SEQ ID NO: 13, and the H-FR4 comprises an amino acid sequence as set forth in SEQ ID NO: 14; or c) the H-FR1 comprises an amino acid sequence as set forth in SEQ ID NO: 11, the H-FR2 comprises an amino acid sequence as set forth in SEQ ID NO: 12, the H-FR3 comprises an amino acid sequence as set forth in SEQ ID NO: 15, and the H-FR4 comprises an amino acid sequence as set forth in SEQ ID NO: 14.

[0107] In the present application, the isolated antigen binding protein can comprise an L-FR1, the C-terminus of which is directly or indirectly connected to the N-terminus of the LCDR1, and the L-FR1 comprises an amino acid sequence as set forth in SEQ ID NO: 34.

[0108] In the present application, the isolated antigen binding protein can comprise an L-FR2 between the LCDR1 and the LCDR2, and the L-FR2 comprises an amino acid sequence as set forth in SEQ ID NO: 17.

[0109] In the present application, the isolated antigen binding protein can comprise an L-FR3 between the LCDR2 and the LCDR3, and the L-FR3 comprises an amino acid sequence as set forth in SEQ ID NO: 35.

[0110] In the present application, the isolated antigen binding protein can comprise an L-FR4, the N-terminus of which is directly or indirectly connected to the C-terminus of the HCDR3, and the L-FR4 comprises an amino acid sequence as set forth in SEQ ID NO: 36.

[0111] In the present application, the isolated antigen binding protein can comprise an L-FR1, an L-FR2, an L-FR3 and an L-FR4. The L-FR1 comprises an amino acid sequence as set forth in SEQ ID NO: 34, the L-FR2 comprises an amino acid sequence as set forth in SEQ ID NO: 17, the L-FR3 comprises an amino acid sequence as set forth in SEQ ID NO: 35, and the L-FR4 comprises an amino acid sequence as set forth in SEQ ID NO: 36.

[0112] In the present application, the isolated antigen binding protein can comprise an L-FR1, the C-terminus of which is directly or indirectly connected to the N-terminus of the LCDR1, and the L-FR1 comprises an amino acid sequence as set forth in SEQ ID NO: 16 or SEQ ID NO: 20.

[0113] In the present application, the isolated antigen binding protein can comprise an L-FR2 between the LCDR1 and the LCDR2, and the L-FR2 comprises an amino acid sequence as set forth in SEQ ID NO: 17.

[0114] In the present application, the isolated antigen binding protein can comprise an L-FR3 between the LCDR2 and the LCDR3, and the L-FR3 comprises an amino acid sequence as set forth in SEQ ID NO: 18, SEQ ID NO: 21 or SEQ ID NO: 23.

[0115] In the present application, the isolated antigen binding protein can comprise an L-FR4, the N-terminus of which is directly or indirectly connected to the C-terminus of the HCDR3, and the L-FR4 comprises an amino acid sequence as set forth in SEQ ID NO: 19 or SEQ ID NO: 22.

[0116] In the present application, the isolated antigen binding protein can comprise an L-FR1, an L-FR2, an L-FR3, and an L-FR4. Wherein the L-FR1 comprises an amino acid sequence as set forth in SEQ ID NO: 16 or SEQ ID NO: 20, the L-FR2 comprises an amino acid sequence as set forth in SEQ ID NO: 17, the L-FR3 comprises an amino acid sequence as set forth in SEQ ID NO: 18, SEQ ID NO: 21, or SEQ ID NO: 23, and the L-FR4 comprises an amino acid sequence as set forth in SEQ ID NO: 19 or SEQ ID NO: 22.

[0117] In the present application, the isolated antigen binding protein can comprise an L-FR1, an L-FR2, an L-FR3, and an L-FR4, wherein: a) the L-FR1 comprises an amino acid sequence as set forth in SEQ ID NO: 16, the L-FR2 comprises an amino acid sequence as set forth in SEQ ID NO: 17, the L-FR3 comprises an amino acid sequence as set forth in SEQ ID NO: 18, and L-FR4 comprises an amino acid sequence as set forth in SEQ ID NO: 19; b) the L-FR1 comprises an amino acid sequence as set forth in SEQ ID NO: 20, the L-FR2 comprises an amino acid sequence as set forth in SEQ ID NO: 17, the L-FR3 comprises an amino acid sequence as set forth in SEQ ID NO: 21, and L-FR4 comprises an amino acid sequence as set forth in SEQ ID NO: 22; or c) the L-FR1 comprises an amino acid sequence as set forth in SEQ ID NO: 20, the L-FR2 comprises an amino acid sequence as set forth in SEQ ID NO: 17, the L-FR3 comprises an amino acid sequence as set forth in SEQ ID NO: 23, and L-FR4 comprises an amino acid sequence as set forth in SEQ ID NO: 22.

[0118] A3. Antibody

[0119] In the present application, the isolated antigen binding protein can comprise a VH, the amino acid sequence of which comprises an amino acid sequence as set forth in SEQ ID NO: 24, SEQ ID NO: 25, or SEQ ID NO: 26.

[0120] In the present application, the isolated antigen binding protein can comprise a VL comprising an amino acid sequence as set forth in SEQ ID NO: 27, SEQ ID NO: 28, or SEQ ID NO: 29.

[0121] In the present application, the isolated antigen binding protein can comprise a VH and a VL, wherein: a) the VH comprises an amino acid sequence as set forth in SEQ ID NO: 24 and the VL comprises an amino acid sequence as set forth in SEQ ID NO: 27; b) the VH comprises an amino acid sequence as set forth in SEQ ID NO: 25 and the VL comprises an amino acid sequence as set forth in SEQ ID NO: 28; c) the VH comprises an amino acid sequence as set forth in SEQ ID NO: 25 and the VL comprises an amino acid sequence as set forth in SEQ ID NO: 29; d) the VH comprises an amino acid sequence as set forth in SEQ ID NO: 26 and the VL comprises an amino acid sequence as set forth in SEQ ID NO: 28; or e) the VH comprises an amino acid sequence as set forth in SEQ ID NO: 26 and the VL comprises an amino acid sequence as set forth in SEQ ID NO: 29.

[0122] In the present application, the isolated antigen binding protein can comprise an antibody or an antigen binding fragment thereof.

[0123] For example, the antigen binding fragment comprises a Fab, a Fab', a F(ab)2, a Fv fragment, a F(ab')2, a scFv, a di-scFv, a VHH, and / or a dAb.

[0124] For example, the antibody is selected from the group consisting of a monoclonal antibody, a chimeric antibody, a humanized antibody, and a fully human antibody.

[0125] In the present application, the isolated antigen binding protein can comprise an antibody heavy chain constant region.

[0126] For example, the heavy chain constant region can comprise an IgG heavy chain constant region. For example, the heavy chain constant region can comprise an IgA heavy chain constant region. For example, the heavy chain constant region can comprise an IgD heavy chain constant region. For example, the heavy chain constant region can comprise an IgM heavy chain constant region. For example, the heavy chain constant region can comprise an IgE heavy chain constant region.

[0127] For example, the heavy chain constant region can comprise an IgG1 heavy chain constant region. For example, the heavy chain constant region can comprise an IgG2 heavy chain constant region. For example, the heavy chain constant region can comprise an IgG3 heavy chain constant region. For example, the heavy chain constant region can comprise an IgG4 heavy chain constant region.

[0128] For example, the heavy chain constant region can be a human IgGl heavy chain constant region.

[0129] In the present application, the isolated antigen binding protein can comprise an antibody light chain constant region.

[0130] For example, the light chain constant region can comprise an IgK light chain constant region. For example, the light chain constant region can comprise an IgA light chain constant region

[0131] For example, the light chain constant region is a human IgK light chain constant region.

[0132] Furthermore, it is noted that the isolated antigen binding protein of the present application can comprise heavy and / or light chain sequences with one or more conservative sequence modifications thereto. By "conservative sequence modification" is meant a modification of an antibody which does not significantly affect or alter its binding characteristics. Such conservative modifications are well known in the art and include, for example, the use of one amino acid for another having a similar side chain. Modifications can be introduced into the isolated antigen binding protein of the present application by standard techniques known in the art, such as site-directed mutagenesis and PCR-mediated mutagenesis. Conservative amino acid substitutions are those substitutions that do not significantly affect or alter the binding characteristics of the antibody. Groups of amino acids that have side chains having similar chemical properties are well known in the art. These groups include amino acids with basic side chains (e.g., lysine, arginine, histidine), acidic side chains (e.g., aspartic acid, glutamic acid), uncharged polar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine, tryptophan, cysteine), nonpolar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine), beta-branched side chains (e.g., threonine, valine, isoleucine) and aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, histidine). In certain embodiments, one or more amino acid residues in the CDR regions of the isolated antigen binding protein of the present application can be replaced with other amino acid residues from the same side chain group. Those skilled in the art will know that some conservative sequence modifications do not render the antigen binding property lost.

[0133] B. Bispecific antibodies

[0134] In another aspect, the present application also provides a bispecific antibody, which can comprise at least a first portion and a second portion, the first portion can target IL-11.

[0135] For example, the second portion can target TSLP, can target IL-17, can target IL-36R, can target IL-4R.

[0136] In the present application, the two portions of the bispecific antibody comprise an antibody or an antigen binding fragment thereof.

[0137] For example, the antibody can be a monoclonal antibody, a chimeric antibody, a humanized antibody, or a fully human antibody.

[0138] For example, the antigen binding fragment is a Fab, a Fab', a F(ab)2, a Fv fragment, a F(ab')2, a scFv, a di-scFv, a VHH, and / or a dAb.

[0139] In the present application, the bispecific antibody can comprise at least a first targeting moiety and a second targeting moiety.

[0140] For example, the first targeting moiety can be a polypeptide that binds to IL-11, can be an antigen binding protein that binds to IL-11.

[0141] For example, the first targeting moiety can be an antigen binding protein as described in the present application.

[0142] In the present application, the bispecific antibody can further comprise a protein scaffold.

[0143] For example, the protein scaffold can be an IgG protein, can be an IgM protein, can be an IgA protein, can be an IgD protein, can be an IgE protein.

[0144] In the present application, the bispecific antigen binding construct can further comprise a linker connecting the antigen binding domain and the protein scaffold.

[0145] For example, the linker comprises 1-150 amino acids.

[0146] C. Fusion proteins, immunoconjugates

[0147] In another aspect, the present application also provides a fusion protein, which can comprise a targeting moiety that binds to IL-11.

[0148] For example, the fusion protein can comprise an antigen binding protein as described in the present application.

[0149] For example, the fusion protein can comprise a bispecific antibody as described in the present application.

[0150] In the present application, the fusion protein can further comprise a polypeptide molecule.

[0151] For example, the fusion protein can be a polypeptide molecule.

[0152] For example, the fusion protein can be a chimeric antigen receptor.

[0153] For example, the fusion protein can be an antibody drug conjugate.

[0154] In another aspect, the present application also provides immunoconjugates, which can comprise the antigen binding proteins described herein, the bispecific antibodies described herein, and / or the fusion proteins described herein.

[0155] D. Nucleic acid molecules, vectors, cells, pharmaceutical compositions

[0156] In another aspect, the present application provides isolated nucleic acid molecules, which can encode the isolated antigen binding proteins described herein. For example, they can be produced or synthesized by (1) amplification in vitro, such as by polymerase chain reaction (PCR); (2) recombinantly produced by cloning; (3) purified, such as by enzymatic cleavage and fractionation by gel electrophoresis; or (4) synthesized, such as by chemical synthesis.

[0157] In another aspect, the present application provides a vector, which can comprise the nucleic acid molecules described herein. In addition, the vector can further comprise other genes, such as marker genes that allow for selection of the vector in appropriate host cells and under appropriate conditions. In addition, the vector can further comprise expression control elements that allow for proper expression of the coding region in the appropriate host. Such control elements are well known to those skilled in the art, and can include, for example, promoters, ribosome binding sites, enhancers, and other control elements that modulate transcription or translation of mRNA. The vector can be introduced into a host cell by transformation, transduction, or transfection, such that the genetic material elements carried by the vector are expressed within the host cell. The vector can include, for example, a plasmid, cosmid, virus, bacteriophage, or other vector commonly used in, for example, genetic engineering. For example, the vector is an expression vector. In addition, the vector can further comprise components that aid in its entry into a cell, such as a viral particle, liposome, or protein coat, but not limited to these.

[0158] In another aspect, the present application provides a cell, which can comprise a nucleic acid molecule described herein or a vector described herein. In certain embodiments, each or every host cell can comprise one or a nucleic acid molecule or vector described herein. In certain embodiments, each or every host cell can comprise a plurality (e.g., 2 or more) or plurality (e.g., 2 or more) of nucleic acid molecules or vectors described herein. For example, a vector described herein can be introduced into the host cell, e.g., a eukaryotic cell, such as a cell from a plant, a fungal or yeast cell, etc. In certain embodiments, the cell can be a bacterial cell (e.g., E. coli), a yeast cell, or other eukaryotic cell, e.g., a COS cell, a Chinese hamster ovary (CHO) cell, a CHO-K1 cell, a LNCAP cell, a HeLa cell, a 293T cell, a COS-1 cell, a SP2 / 0 cell, a NS0 cell, or a myeloma cell. The vector described herein can be introduced into the host cell by methods known in the art, e.g., electroporation, lipofectine transfection, lipofectamin transfection, etc.

[0159] In another aspect, the present application also provides a pharmaceutical composition, which can comprise an isolated antigen binding protein described herein, a bispecific antibody described herein, a fusion protein described herein, an immunoconjugate described herein, a nucleic acid molecule described herein, a vector described herein, and / or a cell described herein, and optionally a pharmaceutically acceptable carrier.

[0160] In certain embodiments, the pharmaceutical composition can further comprise one or more suitable formulations of (pharmaceutically effective) adjuvants, stabilizers, excipients, diluents, solubilizers, surfactants, emulsifiers, and / or preservatives. Acceptable ingredients of the composition are preferably non-toxic to the recipient at the doses and concentrations employed. Pharmaceutical compositions of the present application include, but are not limited to, liquid, frozen, and lyophilized compositions.

[0161] In certain embodiments, the pharmaceutical composition can be a bispecific or multispecific molecule comprising an isolated antigen binding protein described herein. In certain embodiments, in the bispecific or multispecific molecule, the other molecule(s) binding to a target can be unrelated to IL-11.

[0162] In certain embodiments, the pharmaceutical composition can further contain more than one active compound, typically those having complementary activities which do not adversely affect each other. The type and effective amount of such agents can depend, for example, on the amount and type of antagonist present in the formulation, as well as the clinical parameters of the subject.

[0163] In certain embodiments, the pharmaceutically acceptable carrier can include any and all solvents, dispersion media, coatings, isotonic and absorption delaying agents, and the like, that are physiologically compatible and generally safe, non-toxic.

[0164] E. Methods of Preparation

[0165] In another aspect, the present application provides methods of preparing the antigen binding proteins described herein. The methods can comprise culturing the host cells described herein under conditions such that the antigen binding proteins are expressed. For example, the methods can be performed by using appropriate media, appropriate temperatures, and culturing times, as are known to those of ordinary skill in the art.

[0166] Any method suitable for producing monoclonal antibodies can be used to produce the antigen binding proteins of the present application. For example, an animal can be immunized with a TSLP or fragment thereof, either linked or naturally occurring. Suitable immunization protocols can be used, including adjuvants, immunostimulants, repeated booster immunizations, and one or more routes of administration. For example, a hybridoma preparation method can be used, in which spleen cells from an immunized mouse are fused with SP2 / 0 myeloma cells, and hybridoma cell lines are selected by HAT.

[0167] Any suitable form of IL-11 can be used as an immunogen (antigen) for producing non-human antibodies specific for IL-11, and for screening the antibodies for biological activity. For example, the immunogen can be full-length, mature human IL-11, including the native homodimer, or a peptide containing one or more epitopes. The immunogen can be used alone or in combination with one or more immunogenicity enhancers known in the art.

[0168] Chimeric humanized antibodies can be selected from any class of immunoglobulin, including IgM, IgD, IgG, IgA, and IgE. In the present application, the antibodies can be IgG antibodies, and the IgG1 subtype can be used. Optimization of the necessary constant domain sequences to produce the desired biological activity can be achieved by screening the antibodies using the biological assays described in the Examples below. Likewise, any class of light chain can be used in the compounds and methods of the present application. For example, kappa chains or variants thereof can be used in the compounds and methods of the present application.

[0169] F. Methods and Uses

[0170] In another aspect, the present application provides the use of the isolated antigen binding proteins, the bispecific antibodies, the fusion proteins, the nucleic acid molecules, the vectors, the cells, the immunoconjugates, and / or the pharmaceutical compositions described herein for the preparation of a medicament for the prevention and / or treatment of a disease and / or a disorder.

[0171] In another aspect, the present application also provides a method of preventing and / or treating a disease and / or a disorder, which can comprise administering to a subject in need thereof the isolated antigen binding protein, the bispecific antibody, the fusion protein, the nucleic acid molecule, the vector, the cell, the immunoconjugate and / or the pharmaceutical composition described herein.

[0172] In another aspect, the isolated antigen binding protein, the bispecific antibody, the fusion protein, the nucleic acid molecule, the vector, the cell, the immunoconjugate and / or the pharmaceutical composition described herein can be used for preventing and / or treating a disease and / or a disorder.

[0173] In the present application, the disease and / or disorder can be an IL-11 related disease and / or disorder.

[0174] In the present application, the disease and / or disorder can be an inflammatory disease or a tumor.

[0175] In the present application, the disease and / or disorder includes pulmonary fibrosis, renal fibrosis, hepatic fibrosis, myocardial fibrosis, splenic fibrosis, arthritis, ankylosing spondylitis, gastritis, pneumonia, hepatitis, colitis, lung cancer, stomach cancer, bone cancer, colon cancer, and / or liver cancer.

[0176] In another aspect, the present application also provides a method of detecting IL-11 in a sample, which comprises administering the isolated antigen binding protein, the bispecific antibody, the fusion protein, the nucleic acid molecule, the vector, the cell, the immunoconjugate and / or the pharmaceutical composition described herein.

[0177] In certain embodiments, the method of detecting IL-11 in a sample is an in vitro method. In certain embodiments, the method of detecting IL-11 in a sample is not for therapeutic purposes. In certain embodiments, the method of detecting IL-11 in a sample is not a diagnostic method.

[0178] In another aspect, the present application also provides a reagent or a kit for detecting IL-11 in a sample, which comprises the isolated antigen binding protein, the bispecific antibody, the fusion protein, the nucleic acid molecule, the vector, the cell, the immunoconjugate and / or the pharmaceutical composition described herein.

[0179] In another aspect, the present application also provides a kit comprising the isolated antigen binding protein, the bispecific antibody, the fusion protein, the nucleic acid molecule, the vector, the cell, the immunoconjugate and / or the pharmaceutical composition described herein, which is used for detecting the presence and / or the amount of IL-11 in a sample.

[0180] Without wishing to be bound by any theory, the examples below are merely intended to illustrate the antigen binding proteins, methods of making, and uses of the present application, and are not intended to limit the scope of the present application.

[0181] Examples

[0182] Example 1 Preparation of hybridoma cells producing murine monoclonal antibodies

[0183] Balb / C, CD1, and other strains of mice (purchased from Shanghai Lingchang Biotechnology Co., Ltd., and raised by Shanghai Nanfang Model Organism Technology Co., Ltd.) were immunized subcutaneously in multiple sites with recombinant human IL11 (ACRO Biosystems, Cat No: IL1-H5243) / recombinant monkey IL11 protein (Sino Biological Inc., Cat No: 90925-CNCE) as antigens, fully emulsified with Freund's adjuvant (Sigma, Cat No: F5881 / F5506), and cross-immunized with recombinant human IL11 / recombinant monkey IL11 protein, respectively. After three immunizations, blood was collected, and the serum titer of the mice was measured using ELISA. When the serum titer reached 10E5 or more, a pre-fusion immunization was performed. The spleen cells of the mouse with the highest titer were selected to fuse with SP2 / 0 myeloma cells (purchased from ATCC, Cat No: CRL-8287). Hybridoma cell lines were selected using culture medium containing HAT, and the supernatant of the hybridoma cells was screened by ELISA. Hybridoma clones that bound to human IL11 were selected. ELISA was then used to detect the specific binding of the supernatant of the hybridoma to monkey and mouse IL11, and hybridoma cell lines that specifically bound to human, mouse, and monkey IL11 were obtained. The selected hybridoma cells were subjected to supernatant antibody affinity detection. Finally, sequence analysis was performed on the hybridoma cell lines expressing Hu-IL11 antibodies.

[0184] Example 2 Antibody affinity screening of hybridoma cell culture supernatant

[0185] The sample cell and flow cell temperature of the Biacore 8K were set to 25°C and the data collection frequency was 10 Hz. Anti-mouse antibody was amino coupled to a CM5 chip according to the use of mouse anti-capture kit instructions. Antibodies in the culture supernatant were captured as ligand using the anti-mouse antibody coupled CM5 chip. Human, monkey and mouse IL-11 antigens were diluted to 50 nM in running buffer (1 mg / mL CM-Dextran sodium salt, 1 mg / mL Bovine Serum Albumin, HBS-EP+, pH 7.4 buffer) as analyte and running buffer as 0 concentration for background subtraction. The Kinetic screening method was used with an analyte flow rate of 30 μL / min, an association time of 120 s, a dissociation time of 180 s, 10 mM glycine (pH 1.5) 50 μL / min regeneration for 60 s. The data was analyzed using a 1:1 binding model, Fit Local Kinetics model. The test materials and instruments are shown in Table 1.

[0186] Table 1. Kinetic screening test materials and instruments

[0187] The results of the kinetic screening are shown in Table 2.

[0188] Table 2. Hybridoma affinity screening data listing

[0189] Example 3 Variable region gene cloning of anti-human IL-11 antibodies

[0190] Based on the principle of TAKARA 5'RACE technology, the cDNA sequence of the variable region of mouse antibody expressed by hybridoma cell line was cloned. Briefly, the SMARTer 5'RACE Synthesis Kit (TAKARA, Cat. No. 634859) was used to synthesize the heavy chain and light chain variable region gene-specific cDNA according to the instructions. The 5' and 3' ends of the cDNA sequence were modified with PCR primers designed to add appropriate leader sequences to the heavy chain and light chain variable region cDNA, respectively, so that the resulting PCR products could be cloned into the existing recombinant antibody expression heavy chain vector pHB-Fc and light chain vector pHB-Ck by the method of seamless cloning. The pHB-Fc expression vector contains the human IgG1 heavy chain constant region gene sequence, with L234A and L235A (Eu numbering) mutations in CH2 to weaken the ADCC effect of the antibody; the pHB-Ck vector contains the human kappa light chain constant region gene sequence. The heavy chain and light chain variable region PCR amplification products were cloned into the expression vector by In-fusion cloning reagent (TAKARA, Cat. No. 639650) to obtain a human-mouse chimeric antibody expression vector, and transformed into Stellar TM Competent Cells E. coli competent cells (TAKARA, Cat. No. 636763). Single colonies were selected for Sanger sequencing, and the antibody variable region sequence was analyzed. The antibody 900917 was obtained, and the CDR sequence is shown in Table 3.

[0191] Table 3. 900917 antibody CDR sequence (IMGT definition)

[0192] Example 4 Expression of human-mouse chimeric antibody against human IL-11

[0193] The expression vector obtained in Example 2 was amplified in E. coli, and a sufficient amount of plasmid was prepared using an endotoxin-free plasmid extraction kit (Tiangen Biotech (Beijing) Co., Ltd., Cat. No. DP117) for transient transfection of chimeric antibody expression. The host cells used for expression were CHO-S cells (Thermo, Cat. No. R80007). The prepared two heavy chain vectors were mixed with the light chain vector to form a liposome complex with polyetherimide (PEI, Polysciences, Cat. No. 24765-1), and then transfected into CHO-S cells, which were cultured in a carbon dioxide incubator for 7 days. The cell culture supernatant was collected by centrifugation, and the human-mouse chimeric antibody was purified by Protein A affinity chromatography column.

[0194] Example 5 Affinity detection of 900917 antibody

[0195] The SPR method was used to determine the binding kinetic constants of human, mouse and monkey IL-11 with the antibody, and the test materials and methods are shown in Table 4.

[0196] Table 4. Affinity detection test materials and methods

[0197] Example 5.1 Affinity detection with human and mouse IL-11

[0198] The sample cell and flow cell temperature of Biacore 8K were set to 25°C, and the data collection frequency was 10 Hz. The Series S Protein A chip was used to capture antibody samples to 300 RU as ligands, respectively. Human and mouse IL-11 antigens were diluted with HBS-EP+ pH 7.4 buffer to a series of gradient concentrations of 50 nM, 25 nM, 12.5 nM, 6.25 nM, 3.125 nM, 1.56 nM, 0.78 nM and 0.39 nM as analytes, and HBS-EP+ pH 7.4 buffer was used as 0 concentration for background subtraction. The multi-cycle kinetics method was used, with an analyte flow rate of 30 μL / min, an association time of 120 s, a dissociation time of 600 s, 10 mM glycine (pH 1.5) 50 μL / min regeneration for 60 s. The data was analyzed in the 1:1 binding mode, Fit Local Kinetics mode.

[0199] Example 5.2 Affinity detection with monkey IL-11

[0200] The sample cell and flow cell temperature of Biacore 8K were set to 25°C, and the data collection frequency was 10 Hz. The Series S Sensor Chip CM5 chip was used for amino coupling to immobilize Cynomolgus IL-11 as ligands. The samples were diluted with HBS-EP+ pH 7.4 buffer to a series of gradient concentrations as analytes, and HBS-EP+ pH 7.4 buffer was used as 0 concentration for background subtraction. The single-cycle kinetics method was used, with an analyte flow rate of 30 μL / min, an association time of 120 s, a dissociation time of 600 s, 10 mM glycine (pH 1.5) 50 μL / min regeneration for 60 s. The data was analyzed in the 1:1 binding mode, Fit Local Kinetics mode.

[0201] Example 5.3 Affinity detection results of multiple species

[0202] The IL-11 antibody was artificially synthesized according to the sequence disclosed in WO2023 / 143556A1, as a positive control antibody, and the experimental method was the same as 5.1 and 5.2, and the affinity detection results were shown in 5.

[0203] Table 5. Affinity detection results Note: NA means not binding.

[0204] By the affinity constant (KD / M) results of various genera, compared with the positive control antibody which can only bind human and mouse IL-11, 900917 has good target affinity for human, monkey and mouse IL-11, and the target affinity for monkey IL-11 is 10E-09M order of magnitude, and the target affinity for human and mouse IL-11 is 10E-11M order of magnitude.

[0205] Example 6 900917 antibody blocks IL-11 mediated activation of hIL11 Effector Reporter Cell

[0206] Collect well-grown hIL11 Effector Report (Nanjing Kebai, cat#CBP74114) and count, resuspend in medium (ingredients: DMEM + 10% FBS) to 1.2E6 cells / mL cell suspension, 50μL / well into 96-well white plate (Jing'an Biotechnology, cat#J09601); Antibody 901077 is diluted with medium to 2.15nM, and then diluted by 3 times gradient to 4 concentrations, and then diluted by 5 times gradient to 4 concentrations, a total of 9 concentration gradients; The diluted antibody is added to the 96-well white plate at 25μL / well; IL11 protein (Jin'an, cat#C006) is diluted to 0.2ng / mL, and 25μL / well is added to the 96-well white plate; The edge hole is sealed with PBS liquid, 100μL / well, and incubated at 37℃, 5% CO2 for 6 hours; After incubation, the plate is taken out and balanced for 15 minutes, 100μL / well is added Ultra Luciferase Detection Kit reagent (Yaoke Yuan, cat#A2000301N), reaction for 5 minutes, full wavelength detection of microplate reader. Using GraphPad Prism 8 software, the sample working concentration-bioluminescence reading is used to analyze the data by four-parameter equation to obtain the sample dose-effect curve.

[0207] The experimental results are shown in Figure 1, which shows that the anti-IL-11 antibody 900917 of the application can block the activation of IL-11 mediated hIL11 Effector Reporter Cell.

[0208] Example 7 900917 antibody blocks IL-11 / IL-11Ra / GP130 ternary complex formation

[0209] The blocking activity of the anti-IL-11 antibody on the formation of the IL-11 / IL-11Ra / GP130 ternary complex was analyzed by a competitive ELISA method. Human IL-11Ra (Sino Biological Inc. Cat#: 10252-H08H) was added to the enzyme-linked plate and coated overnight; the next day, the sample was prepared, first 0.4 μg / mL human IL-11 (Sino Biological Inc. Cat#: 12225-HNCE) was prepared, then mixed with gradient-diluted anti-IL-11 antibody (dilution concentration: 1000, 62.5, 15.625, 3.906, 0.977, 0.244, 0.0610 nM) in equal volume 1:1, and then added to the IL-11Ra-coated enzyme-linked plate for co-incubation, and finally 2 μg / mL of GP130 (Sino Biological Inc. Cat#: 10974-H05H) was added for co-incubation. After one hour, HRP-labeled anti-mouse Fc secondary antibody was added after washing, and color development and OD450 detection of absorbance were performed.

[0210] The experimental results are shown in Figure 2, which shows that 900917 has good blocking activity for blocking the formation of the IL-11 / IL-11Ra / GP130 ternary complex, with an IC50 of 3.3 nM.

[0211] Example 8 Humanization of murine anti-human IL-11 antibody

[0212] By comparing the variable region sequences of the chimeric antibody (900917) with the available sequences in the NCBI IgBlast database, and by identification and analysis, the human framework regions (FR regions) suitable for constructing CDR-grafted heavy and light chains were finally determined.

[0213] During the modification, according to the conserved amino acid residues of the human antibody FR region and the important amino acid residues in the antibody FR region, the modification sites were designed, and the variable regions of the heavy and light chains of the chimeric antibody were respectively subjected to humanization mutation design. The humanization point mutation antibody expression plasmid was amplified and constructed by using PCR technology. The humanization point mutation antibody expression plasmid was expressed in CHO-S cells, and the humanized antibody protein was obtained after purification. Humanized antibodies 901069, 901072, 901074, 901077, etc. were obtained.

[0214] Example 9 Affinity and blocking activity detection of humanized IL-11 antibody

[0215] The affinity and blocking activity of the obtained humanized molecules were detected. The affinity detection method was the same as that in Examples 5.1 and 5.2, and the blocking activity detection method was the same as that in Example 6. Taking humanized antibodies 901069, 901072, 901074, and 901077 as examples, the detection results are shown in Table 6.

[0216] Table 6. Affinity and blocking activity of humanized molecules of anti-human IL-11 human-murine chimeric antibody 900917

[0217] Example 10 In vitro inhibitory ability of anti-IL-11 antibodies on fibrosis

[0218] The inhibitory ability of anti-IL-11 antibodies on TGFβ (transforming growth factor β)-induced fibrosis in NICH0018 (human skin fibroblasts) was detected using 900917 as an example.

[0219] qPCR detection: After the medium (89% DMEM (Dulbecco's medium) + 10% FBS (fetal bovine serum) + 1% P / S (penicillin, streptomycin)) of NICH0018 cells in the logarithmic growth phase in the culture bottle was removed, the cells were washed twice with 10 mL of PBS, and then 2 mL of 0.25% trypsin was added for digestion in the incubator (37°C, 5% carbon dioxide) for 3-5 minutes. After the digestion was terminated by adding the medium, a cell suspension was obtained, centrifuged (500 g x 5 min) to discard the culture medium, resuspended to 1 mL with the medium, and then counted on a counting instrument. 5 x 105 cells per well were plated in a six-well plate, the medium was supplemented to 2 mL per well, and then incubated in the incubator for 8 h to adhere to the wall. The complete medium (DMEM without FBS) was replaced for 16 h of starvation treatment. The medium was prepared according to a final concentration of 10 ng / mL TGFβ or 15 μg / mL 900917, 2 mL of the original medium was added according to the blank group after the original medium was removed, 2 mL of the medium containing 10 ng / mL TGF-beta was added to the stimulation group, and 2 mL of the medium containing 10 ng / mL TGF-beta and 15 μg / mL 900917 was added to the antibody group, with three repeats in each group. After incubation in the incubator for 24 h, the cells were collected in an EP tube without RNase by washing twice with PBS and using a cell scraper, and then centrifuged (500 g x 5 min) to obtain a cell pellet. 5

[0220] ​RT-PCR was used to detect the expression of fibrosis-related genes IL-11 (encoding IL-11 protein) and COL IV (encoding collagen IV) in the cells. 350 μL Buffer RLT was added to the cell pellet to mix and lyse the cells, and 350 μL of 70% ethanol was added to the lysate and mixed evenly. The previous lysate was transferred to a 2 mL collection tube containing a RNeasy MinElute spin column, centrifuged at 8000 g for 15 s, and the filtrate was discarded. 350 uL Buffer RW1 was added to the RNeasy MinElute spin column, centrifuged at 8000 g for 15 s, and the filtrate was discarded. The RNeasy MinElute spin column was placed in a new 2 mL collection tube, 500 μL Buffer RPE was added, centrifuged at 8000 g for 15 s, and the filtrate was discarded. 500 μL of 80% ethanol was added to the RNeasy MinElute spin column, centrifuged at 8000 g for 2 min, and the collection tube was discarded. The RNeasy MinElute spin column was placed in a new 1.5 mL collection tube, and centrifuged at the highest speed of the centrifuge for 5 min with the lid open, and the filtrate and collection tube were discarded. The RNeasy MinElute spin column was placed in a new 1.5 mL collection tube, and 14 μL of RNase-free water was added to the center of the adsorption column, and centrifuged at the highest speed of the centrifuge for 1 min. The RNA concentration was detected after eluting the RNA. The reaction system and reaction conditions for removing the genomic reaction are as follows:

[0221] After obtaining the reaction solution of the removal reaction, the reverse transcription reaction was performed, and the reaction system and reaction conditions are as follows:

[0222] After obtaining the cDNA, quantitative PCR was performed:

[0223] The reaction solution was prepared according to the following table, and the mixed solution was added to the corresponding reaction tube, 5.6 μL each. And 1 μL of sample cDNA was mixed with 3.4 μL of DEPC H2O to form Mix, and was added to the corresponding hole. At the same time, a negative control hole was set, and the template was sterile water.

[0224] Program setting: comparative CT (ΔΔCT) method was used to set up the software running program. PCR amplification program: denaturation 95 degrees for 10 min, 1 cycle; denaturation 95 degrees for 15 s, extension 60 degrees for 30 s, 40 cycles; denaturation 95 degrees for 15 s, extension 60 degrees for 1 min, 0.15 degrees gradient rise to 95 degrees. Running quantitative PCR.

[0225] The gene detection results are shown in Figure 3, and the results show that the anti-IL-11 hybridoma antibody 900917 can inhibit the increase of the fibrosis-related genes IL-11 and COLIV induced by TGFβ. It is proved that 900917 can inhibit the fibrosis of NICH0018 cells induced by TGFβ.

[0226] Example 11 Immunofluorescence detection of anti-IL-11 antibody

[0227] Taking the 900917 antibody as an example, the in-vitro fibrosis inhibition ability of the antibody of the application is further detected by immunofluorescence.

[0228] NICH0018 cells in log phase in culture flask were removed medium (89% DMEM (Dulbecco's medium) + 10% FBS (fetal bovine serum) + 1% P / S (penicillin, streptomycin)) after, 10 mL PBS wash twice, after adding 2 mL 0.25% trypsin, in the incubator (37℃, 5% carbon dioxide) digestion 3-5 min. After adding medium to stop digestion, obtain cell suspension, centrifugation (500g x 5 min) to discard the culture medium, resuspended to 1 mL after counting in the counting instrument. Poured into a twelve-hole plate with pre-placed treated coverslips with 1x105 cells per well, make up medium to 2 mL per well, incubated in the incubator for 8 h after adhering, replace the complete medium (DMEM without FBS) for 16 h. According to the final concentration of 10 ng / mL TGF beta or 15 μg / mL 900917 configuration medium, after removing the original medium, according to the blank group, add 2 mL of ordinary medium, the stimulation group adds 2 mL of 10 ng / mL TGF-beta medium, the antibody group adds 2 mL of 10 ng / mL TGF-beta and 15 μg / mL 900917 medium, three repeats in each group. After incubation in the incubator for 24 h, the culture plate was taken out, the medium was discarded, and the PBS was washed 3 times. The coverslips were fixed with 4% paraformaldehyde at room temperature for 15 min, and the coverslips were washed with PBS for 3 times. Incubate with Biyun Tian immunostaining blocking solution at room temperature for 10 min, wash the coverslips with PBS for 3 times, dilute Fibronectin-AF488 (1:200) and TRITC-Phalloidin (1:1000) with Biyun Tian immunostaining primary antibody diluent, and incubate at 4℃ overnight. Wash with Biyun Tian immunostaining washing solution for 3 times, each for 3 min. Add one drop of Thermo Fisher ProLongTM Glass Anti-Quenching Mountant (containing NucBlue TM stain) on the glass slide, and mount the treated cell coverslips with the cell side down. Use Olympus CKX53 fluorescence inverted microscope to collect 420 nm fluorescence signal (nucleus) excited by 340-390 nm fluorescence, 510 nm fluorescence signal (Fibronectin) excited by 460-495 nm fluorescence, and 575 nm fluorescence signal (Phalloidin) excited by 530-550 nm fluorescence, respectively. Merge the channel images using ImageJ software.

[0229] The results of immunofluorescence staining detection are shown in Figure 4. The results show that the anti-IL-11 hybridoma antibody 900917 can inhibit the expression of fibrosis-related protein FN induced by TGF beta. It is proved that 900917 can inhibit the fibrosis of NICH0018 cells induced by TGF beta.

[0230] Example 12 Pharmacodynamic evaluation of anti-IL-11 antibody in unilateral ureteral ligation (UUO) chronic kidney injury model

[0231] The anti-IL-11 antibody was evaluated for pharmacodynamics in a UUO chronic kidney fibrosis model. Mice were randomly divided into 4 groups, 6 in each group. Among them, 6 in the normal control group, 6 in the IgG control group (given control IgG antibody, 30 mg / kg), 6 in the positive control group (valsartan, 30 mg / kg), and 6 in the 900917 group (30 mg / kg). On the day of surgery, the left ureter of the mice was ligated using inhalation anesthesia. One day after UUO surgery, dosing began, twice a week for 3 weeks (6 doses in total). Before sampling, the animal body weight was measured. Blood was taken, and serum was aliquoted and stored at -80°C. The surgical kidney was weighed; part was preserved in formalin (for tissue staining). The indicators detected include: serum creatinine content, serum urea nitrogen content, kidney tissue PAS staining, Masson-trichrome and FN staining. Data analysis was performed using GraphPad Prism 8.0. The results are shown in Figure 5.

[0232] The analysis results show that no obvious kidney fibrosis and impaired kidney function were observed in the normal control group. In the model group (IgG control group), obvious tissue fibrosis occurred. The control drug and anti-IL-11 antibody drugs showed different degrees of inhibition of fibrosis, manifested as a decrease in the fibrosis area in fibrosis-related tissue staining.

Claims

1. An isolated antigen-binding protein capable of binding IL-11, said antigen-binding protein comprising HCDR1, HCDR2 and HCDR3, wherein said HCDR1 comprises the amino acid sequence shown in SEQ ID NO:1, said HCDR2 comprises the amino acid sequence shown in SEQ ID NO:2 and said HCDR3 comprises the amino acid sequence shown in SEQ ID NO:

3.

2. The antigen-binding protein according to claim 1, wherein the antigen-binding protein further comprises LCDR1, LCDR2 and LCDR3, wherein LCDR1 comprises the amino acid sequence shown in SEQ ID NO:4, LCDR2 comprises the amino acid sequence shown in SEQ ID NO:5, and LCDR3 comprises the amino acid sequence shown in SEQ ID NO:

6.

3. The antigen-binding protein according to any one of claims 1-2, wherein the antigen-binding proteins HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3, wherein HCDR1 comprises the amino acid sequence shown in SEQ ID NO:1, HCDR2 comprises the amino acid sequence shown in SEQ ID NO:2, HCDR3 comprises the amino acid sequence shown in SEQ ID NO:3, LCDR1 comprises the amino acid sequence shown in SEQ ID NO:4, LCDR2 comprises the amino acid sequence shown in SEQ ID NO:5, and LCDR3 comprises the amino acid sequence shown in SEQ ID NO:

6.

4. The antigen-binding protein according to any one of claims 1-3, comprising H-FR1, H-FR2, H-FR3, and H-FR4, wherein: a) The H-FR1 contains the amino acid sequence shown in SEQ ID NO:30; b) The H-FR2 contains the amino acid sequence shown in SEQ ID NO:31; c) The H-FR3 comprises the amino acid sequence shown in SEQ ID NO:32; and d) The H-FR4 contains the amino acid sequence shown in SEQ ID NO:

33.

5. The antigen-binding protein according to any one of claims 1-4, comprising H-FR1, H-FR2, H-FR3, and H-FR4, wherein: a) The H-FR1 contains an amino acid sequence as shown in SEQ ID NO:7 or SEQ ID NO:11; b) The H-FR2 contains an amino acid sequence as shown in SEQ ID NO:8 or SEQ ID NO:12; c) The H-FR3 comprises an amino acid sequence as shown in SEQ ID NO:9, SEQ ID NO:13, or SEQ ID NO:15; and d) The H-FR4 contains an amino acid sequence as shown in SEQ ID NO:10 or SEQ ID NO:

14.

6. The antigen-binding protein according to any one of claims 1-5, comprising H-FR1, H-FR2, H-FR3 and H-FR4, wherein: a) H-FR1 contains the amino acid sequence shown in SEQ ID NO:7, H-FR2 contains the amino acid sequence shown in SEQ ID NO:8, H-FR3 contains the amino acid sequence shown in SEQ ID NO:9, and H-FR4 contains the amino acid sequence shown in SEQ ID NO:

10. b) H-FR1 contains the amino acid sequence shown in SEQ ID NO:11, H-FR2 contains the amino acid sequence shown in SEQ ID NO:12, H-FR3 contains the amino acid sequence shown in SEQ ID NO:13, and H-FR4 contains the amino acid sequence shown in SEQ ID NO:14; or c) H-FR1 contains the amino acid sequence shown in SEQ ID NO:11, H-FR2 contains the amino acid sequence shown in SEQ ID NO:12, H-FR3 contains the amino acid sequence shown in SEQ ID NO:15, and H-FR4 contains the amino acid sequence shown in SEQ ID NO:

14.

7. The antigen-binding protein according to any one of claims 1-6, comprising a heavy chain variable region VH, wherein the VH comprises the amino acid sequence as described in SEQ ID NO:24, SEQ ID NO:25 or SEQ ID NO:

26.

8. The antigen-binding protein according to any one of claims 1-7, further comprising a heavy chain constant region.

9. The antigen-binding protein according to claim 8, wherein the heavy chain constant region is the IgG heavy chain constant region.

10. The antigen-binding protein according to any one of claims 8-9, wherein the heavy chain constant region is the human IgG heavy chain constant region.

11. The antigen-binding protein according to any one of claims 1-10, comprising L-FR1, L-FR2, L-FR3, and L-FR4, wherein: a) The L-FR1 contains the amino acid sequence shown in SEQ ID NO:34; b) The L-FR2 contains the amino acid sequence shown in SEQ ID NO:17; c) The L-FR3 comprises the amino acid sequence shown in SEQ ID NO:35; and d) The L-FR4 contains the amino acid sequence shown in SEQ ID NO:

36.

12. The antigen-binding protein according to any one of claims 1-11, comprising L-FR1, L-FR2, L-FR3, and L-FR4, wherein: a) The L-FR1 contains an amino acid sequence as shown in SEQ ID NO:16 or SEQ ID NO:20; b) The L-FR2 contains the amino acid sequence shown in SEQ ID NO:17; c) The L-FR3 comprises the amino acid sequence shown in SEQ ID NO:18, SEQ ID NO:21 or SEQ ID NO:23; and d) The L-FR4 contains an amino acid sequence as shown in SEQ ID NO:19 or SEQ ID NO:

22.

13. The antigen-binding protein according to any one of claims 1-12, comprising L-FR1, L-FR2, L-FR3, and L-FR4, wherein: a) The L-FR1 contains the amino acid sequence shown in SEQ ID NO:16, the L-FR2 contains the amino acid sequence shown in SEQ ID NO:17, the L-FR3 contains the amino acid sequence shown in SEQ ID NO:18, and the L-FR4 contains the amino acid sequence shown in SEQ ID NO:

19. b) The L-FR1 contains the amino acid sequence shown in SEQ ID NO:20, the L-FR2 contains the amino acid sequence shown in SEQ ID NO:17, the L-FR3 contains the amino acid sequence shown in SEQ ID NO:21, and the L-FR4 contains the amino acid sequence shown in SEQ ID NO:22; or c) The L-FR1 contains the amino acid sequence shown in SEQ ID NO:20, the L-FR2 contains the amino acid sequence shown in SEQ ID NO:17, the L-FR3 contains the amino acid sequence shown in SEQ ID NO:23, and the L-FR4 contains the amino acid sequence shown in SEQ ID NO:

22.

14. The antigen-binding protein according to any one of claims 1-13, comprising a light chain variable region VL, wherein the VL comprises the amino acid sequence as described in SEQ ID NO:27, SEQ ID NO:28 or SEQ ID NO:

29.

15. The antigen-binding protein according to any one of claims 1-14, further comprising a light chain constant region.

16. The antigen-binding protein according to claim 15, wherein the light chain constant region is the Igκ light chain constant region.

17. The antigen-binding protein according to any one of claims 15-16, wherein the light chain constant region is the human Igκ light chain constant region.

18. The antigen-binding protein according to any one of claims 1-17, comprising a heavy chain variable region VH and a light chain variable region VL, wherein: a) The VH contains the amino acid sequence shown in SEQ ID NO:24, and the VL contains the amino acid sequence shown in SEQ ID NO:27; b) The VH contains the amino acid sequence shown in SEQ ID NO:25, and the VL contains the amino acid sequence shown in SEQ ID NO:28; c) The VH contains the amino acid sequence shown in SEQ ID NO:25, and the VL contains the amino acid sequence shown in SEQ ID NO:29; d) The VH contains the amino acid sequence shown in SEQ ID NO:26, and the VL contains the amino acid sequence shown in SEQ ID NO:28; or e) The VH contains the amino acid sequence shown in SEQ ID NO:26, and the VL contains the amino acid sequence shown in SEQ ID NO:

29.

19. The antigen-binding protein according to any one of claims 1-18, comprising an antibody or an antigen-binding fragment thereof.

20. The antigen-binding protein of claim 19, wherein the antigen-binding fragment is Fab, Fab', F(ab)2, Fv fragment, F(ab')2, scFv, di-scFv, VHH and / or dAb.

21. The antigen-binding protein according to any one of claims 19-20, wherein the antibody is selected from the group consisting of monoclonal antibodies, chimeric antibodies, humanized antibodies, and fully human antibodies.

22. A fusion protein comprising the antigen-binding protein according to any one of claims 1-21.

23. A bispecific antibody comprising a first portion and a second portion, wherein the first portion comprises the antigen-binding protein of any one of claims 1-21.

24. An immunoconjugate comprising the antigen-binding protein of any one of claims 1-21, the fusion protein of claim 22, and / or the bispecific antibody of claim 23.

25. One or more isolated nucleic acid molecules encoding an antigen-binding protein according to any one of claims 1-21, a fusion protein according to claim 22, a bispecific antibody according to claim 23, and / or an immunoconjugate according to claim 24.

26. A vector comprising the nucleic acid molecule of claim 25.

27. A cell comprising and / or expressing the antigen-binding protein of any one of claims 1-21, the fusion protein of claim 22, the bispecific antibody of claim 23, the immunoconjugate of claim 24, the nucleic acid molecule of claim 25, and / or the vector of claim 26.

28. A pharmaceutical composition comprising the antigen-binding protein of any one of claims 1-21, the fusion protein of claim 22, the bispecific antibody of claim 23, the immunoconjugate of claim 24, the nucleic acid molecule of claim 25, the carrier of claim 26, and / or the cell of claim 27, and optionally a pharmaceutically acceptable carrier.

29. Use of the antigen-binding protein of any one of claims 1-21, the fusion protein of claim 22, the bispecific antibody of claim 23, the immunoconjugate of claim 24, the nucleic acid molecule of claim 25, the carrier of claim 26, the cell of claim 27, and / or the pharmaceutical composition of claim 28 in the preparation of a medicament for the prevention and / or treatment of diseases and / or conditions.

30. The use according to claim 29, wherein the disease and / or condition includes diseases and / or conditions associated with IL-11 expression.

31. The use according to any one of claims 29-30, wherein the disease and / or condition includes inflammatory diseases or tumors.

32. The use according to any one of claims 29-31, wherein the disease and / or condition includes pulmonary fibrosis, renal fibrosis, hepatic fibrosis, myocardial fibrosis, splenic fibrosis, arthritis, ankylosing spondylitis, gastritis, pneumonia, hepatitis, colitis, lung cancer, gastric cancer, bone cancer, colon cancer, and / or liver cancer.

33. A method for preparing the antigen-binding protein of any one of claims 1-21, the fusion protein of claim 22, and / or the bispecific antibody of claim 23, the method comprising culturing the cells of claim 27 under conditions in which the antigen-binding protein, the fusion protein, and the bispecific antibody are expressed.

34. A kit comprising the antigen-binding protein of any one of claims 1-21, the fusion protein of claim 22, the bispecific antibody of claim 23, the immunoconjugate of claim 24, the nucleic acid molecule of claim 25, the vector of claim 26, the cell of claim 27, and / or the pharmaceutical composition of claim 28.

35. A method for preventing and / or treating a disease and / or condition in a subject who requires such treatment, the method comprising administering to the subject an effective amount of any of the antigen-binding protein of claims 1-21, the fusion protein of claim 22, the bispecific antibody of claim 23, the immunoconjugate of claim 24, the nucleic acid molecule of claim 25, the carrier of claim 26, the cell of claim 27, and / or the pharmaceutical composition of claim 28.

36. The method of claim 35, wherein the disease and / or condition includes diseases and / or conditions associated with IL-11 expression.

37. The method according to any one of claims 35-36, wherein the disease and / or condition includes inflammatory diseases or tumors.

38. The use according to any one of claims 35-37, wherein the disease and / or condition includes pulmonary fibrosis, renal fibrosis, hepatic fibrosis, myocardial fibrosis, splenic fibrosis, arthritis, ankylosing spondylitis, gastritis, pneumonia, hepatitis, colitis, lung cancer, gastric cancer, bone cancer, colon cancer, and / or liver cancer.

39. The antigen-binding protein of any one of claims 1-21, the fusion protein of claim 22, the bispecific antibody of claim 23, the immunoconjugate of claim 24, the nucleic acid molecule of claim 25, the vector of claim 26, the cell of claim 27, and / or the pharmaceutical composition of claim 28, for the prevention and / or treatment of diseases and / or conditions.

40. The antigen-binding protein, fusion protein, bispecific antibody, immunoconjugate, nucleic acid molecule, vector, cell and / or pharmaceutical composition according to claim 39, wherein the disease and / or condition includes diseases and / or conditions associated with IL-11 expression.

41. The antigen-binding protein, fusion protein, bispecific antibody, immunoconjugate, nucleic acid molecule, carrier, cell and / or pharmaceutical composition according to any one of claims 39-40, wherein the disease and / or condition includes inflammatory diseases or tumors.

42. The antigen-binding protein, fusion protein, bispecific antibody, immunoconjugate, nucleic acid molecule, carrier, cell and / or pharmaceutical composition according to any one of claims 39-41, wherein the disease and / or condition includes pulmonary fibrosis, renal fibrosis, hepatic fibrosis, myocardial fibrosis, splenic fibrosis, arthritis, ankylosing spondylitis, gastritis, pneumonia, hepatitis, colitis, lung cancer, gastric cancer, bone cancer, colon cancer, and / or liver cancer.

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