ICAM-1 antibody and application thereof

Abstract

The invention provides a monoclonal antibody or an antigen binding part thereof, the monoclonal antibody can bind to ICAM-1 and comprises a heavy chain variable region and a light chain variable region, the amino acid sequences of the CDR1-3 region of the heavy chain variable region are sequentially shown as SEQ ID NOs: 1-3, and the amino acid sequences of the CDR1-3 region of the light chain variable region are sequentially shown as SEQ ID NOs: 4-6. The invention further provides application of the method. According to the present invention, the LFA-1 / ICAM-1 interaction can be specifically inhibited, and especially the LFA-1 / ICAM-1 can be used as the diagnostic reagent to prevent or treat related tumors and inflammations.

Description

Technical Field

[0001] The present invention relates to an antibody and application thereof. Background Art

[0002] LFA-1 is a member of the integrin β2 family, expressed on the surface of leukocytes, and composed of αL (CD11a) subunit and β2 (CD18) subunit. ICAM-1 is mainly involved in leukocyte transport and immune cell recruitment, and its expression is upregulated when stimulated by inflammatory factors. The LFA-1 / ICAM-1 interaction plays a role in T cell activation, T cell migration and potential secondary activation of ocular surface T cells in lymph nodes. ICAM-1 plays an important role in promoting adhesion of inflammatory sites, controlling tumor deterioration and metastasis, and regulating the body's immune response. It enhances the adhesion between leukocytes, inflammatory cells, tumor cells, etc. and endothelial cells through specific binding to its receptors, promotes endothelial cell activation, and makes it easier for them to penetrate the endothelium. Intercellular adhesion molecules on endothelial cells mediate contact and binding between cells or between cells and matrix, thereby participating in physiological and pathological processes such as cell signal transduction and activation, cell tissue growth and differentiation, immune response, inflammatory response, angiogenesis and tumor metastasis. Studies have shown that effector T cells reach the ocular surface along blood vessels and secrete various cytokines to promote the production of pro-inflammatory mediators in ocular tissues, causing ocular surface damage. Inhibiting LFA-1 / ICAM-1 interaction and thus destroying various key steps of T cell-mediated inflammation can inhibit the inflammatory pathways of tumors such as solid tumors or dry eye disease (DED). Summary of the invention

[0003] The present invention provides an antibody or an antigen binding portion thereof that can bind to ICAM-1. The present invention can specifically inhibit the interaction between LFA-1 / ICAM-1 and treat malignant tumors or related inflammation such as DED.

[0004] Specifically, the present invention includes the following aspects: The first aspect of the present invention relates to an anti-ICAM-1 monoclonal antibody or an antigen-binding portion thereof, wherein the amino acid sequences of the CDR1-3 regions of the heavy chain variable region VH are shown in SEQ ID NOs: 1-3 (NOs in the art represent a parallel listing of sequence numbers, i.e., sequence numbers 1 / 2 / 3), and the amino acid sequences of the CDR1-3 regions of the light chain variable region VL are shown in SEQ ID NOs: 4-6. Optionally, the VH amino acid sequence is SEQ ID NO: 7, or the VL sequence is SEQ ID NO: 8. Optionally, the monoclonal antibody or its antigen-binding portion is a complete antibody, a bispecific antibody, scFv, Fab, Fab', F(ab')2 or Fv, such as scFv, in which case: its VH and VL can be coupled by a flexible linker, and the linker can be a short peptide such as (GGGS)nC, (GGSC)n, (GGGGS)n or (GGG)n, wherein n=1-10, preferably n=3; the connection position of VH and VL can be the N-terminus of the heavy chain connected to the C-terminus of the light chain, or the N-terminus of the light chain can be connected to the C-terminus of the heavy chain. The scheme containing the constant region of the present invention includes: In one embodiment, the heavy chain constant region of the anti-ICAM-1 monoclonal antibody or antigen-binding portion thereof is selected from the group consisting of IgG, IgM, IgE, IgD and IgA derived from human.

[0005] In one embodiment of the present invention, the heavy chain constant region of the anti-ICAM-1 monoclonal antibody or antigen-binding portion thereof is selected from the group consisting of IgG1, IgG2, IgG3 and IgG4 derived from human.

[0006] In a specific embodiment of the present invention, the heavy chain constant region of the anti-ICAM-1 monoclonal antibody or the antigen-binding portion thereof is derived from human IgG1.

[0007] In one embodiment of the present invention, the light chain constant region of the anti-ICAM-1 monoclonal antibody or antigen-binding portion thereof is derived from human κ or λ.

[0008] In a specific embodiment of the present invention, the light chain constant region of the anti-ICAM-1 monoclonal antibody or antigen-binding portion thereof is derived from human κ.

[0009] The present invention also relates to a nucleic acid molecule comprising a sequence encoding any one of the anti-ICAM-1 monoclonal antibodies or antigen binding portions thereof according to the first aspect of the present invention.

[0010] In one embodiment of the present invention, the nucleic acid molecule comprises the sequence h and / or l: Sequence h: CAGGTCAAACTGCAGCAGTCAGGGGCTGACCTGGTGAAGCCTGGGGCTTCAGTGATGTTGTCCTGCAAGGCTTCTGGCTACACTTTCACCAGCTACTATATGTACTGGGTGAAGCAGAGGCCTGGACAAGGCCTTGAGTGGATTGGAGAGATTAATCCTAGCAATGGTGGTACTAACTTCAATGAGAAGTTCAAGACCAAGGCCACACTGACTGTAGACAGATCCTCCAGCACAGCATACATGCAACTCAGCAATCTGACATCTGAGGACTCTGCGGTCTATTACTGTACAAGACAGGCTGGTTACTACGGATACTATGCTATGGACTACTGGGGCCAAGGGACCACGGTCACCGTCTCCTCA Sequence l: GACATTGAGCTCACCCAGTCTCCAGCAATCATGTCTGCATCTCCAGGGGAAAAGGTCACCATGACCTGCAGGGCCAGCTCAAGTGTAAGTTCCAGTTACTTGCACTGGTACCAGCAGAAGTCAGGTGCCTCCCCCAAACTCTTGATTTATAGGACATCCAATCTGGCTTCTGGAGTCCCAGCTCGCTTCAGTGGCAGTGGGTCTGGGACCTCTTATTCTCTCACAGTCAGCAGTGTAGAGGCTGAAGATGCTGCCACTTATTACTGCCAGCAGTACAGTGGTTACCCACTATTCACGTTCGGCTCGGGGACAAAGTTGGAAATAAAACGG, for example, including: .

[0011] The present invention also relates to a recombinant vector comprising any of the above-mentioned nucleic acid molecules of the present invention.

[0012] In the present invention, the vector may be a cloning vector or an expression vector. The vector contains the nucleic acid molecule described in any one of the present invention, and the vector may be obtained by, for example, inserting the nucleic acid molecule into a cloning vector or an expression vector, or may be obtained by artificial synthesis.

[0013] In the present invention, the expression vector is, for example, a prokaryotic expression vector, a eukaryotic expression vector, a phage vector or a viral vector. The prokaryotic expression vector is, for example, a PET vector or a PGEX vector, the eukaryotic expression vector is, for example, pcDNA3.1, pEGFP-C1, pPIC9K, the phage vector is, for example, a λ phage vector λgt, λgt-λB, and the viral vector is, for example, a retrovirus, a lentivirus, an adenovirus or an adeno-associated virus vector.

[0014] The present invention also relates to a recombinant cell comprising any recombinant vector of the present invention.

[0015] In one embodiment of the present invention, the cell is a mammalian cell, or a bacterium such as Escherichia coli, or a fungus such as yeast. The mammalian cell is a cell suitable for expressing antibodies, such as a mammalian cell derived from humans, mice or monkeys. In a more specific implementation scheme, the mammalian cell is a CHO cell, such as a CHO-K1 cell.

[0016] The present invention also relates to a composition comprising any anti-ICAM-1 monoclonal antibody or antigen binding portion thereof of the present invention, any nucleic acid molecule of the present invention, any recombinant vector of the present invention, or any recombinant cell of the present invention, and optionally a pharmaceutically acceptable carrier or excipient.

[0017] The present invention also relates to a method for preparing any anti-ICAM-1 monoclonal antibody or antigen-binding portion thereof of the present invention, which comprises the step of using any nucleic acid molecule of the present invention, any recombinant vector of the present invention, or any recombinant cell of the present invention.

[0018] In one embodiment of the present invention, it specifically comprises the following steps: 1) cloning the nucleotide sequence of any nucleic acid molecule of the present invention into an expression vector to obtain a recombinant expression vector; 2) Transfer the recombinant expression vector into host cells to obtain recombinant cells; 3) culturing the recombinant cells obtained in step 2) in a target culture medium to obtain a cell line capable of expressing the antibody; 4) Gradually scale up the cell line obtained in step 3) and harvest the culture supernatant; 5) Purifying the culture supernatant obtained in step 4) to obtain any anti-ICAM-1 monoclonal antibody or antigen-binding portion thereof of the present invention.

[0019] In one embodiment of the present invention, the cell is a mammalian cell, which is a cell suitable for expressing antibodies, such as a mammalian cell derived from human, mouse or monkey. In a specific embodiment of the present invention, the mammalian cell is a CHO cell, such as a CHO-K1 cell.

[0020] The present invention also relates to any anti-ICAM-1 monoclonal antibody or antigen binding portion thereof of the present invention for specifically inhibiting LFA-1 / ICAM-1 interaction, especially as a diagnostic reagent, prevention or treatment of related tumors and inflammation. Tumors include solid tumors such as breast cancer and thyroid cancer. An example of inflammation is DED.

[0021] The present invention is further described below: In the present invention, unless otherwise specified, the scientific and technical terms used herein have the meanings commonly understood by those skilled in the art. In addition, the protein and nucleic acid chemistry, molecular biology, cell and tissue culture, microbiology, immunology and laboratory operation procedures used herein are terms and routine procedures widely used in the corresponding fields. At the same time, in order to better understand the present invention, the definitions and explanations of the relevant terms are provided below.

[0022] In the present invention, the term "antibody" refers to an immunoglobulin molecule that is generally composed of two pairs of identical polypeptide chains, each pair having one "light" (L) chain and one "heavy" (H) chain. Antibody light chains can be classified as κ and λ light chains. Heavy chains can be classified as μ, δ, γ, α or ε, and define the isotype of the antibody as IgM, IgD, IgG, IgA and IgE, respectively. Within the light and heavy chains, the variable and constant regions are connected by a "J" region of about 12 or more amino acids, and the heavy chain also includes a "D" region of about 3 or more amino acids. Each heavy chain consists of a heavy chain variable region (V H ) and the heavy chain constant region (C H ). The heavy chain constant region consists of three domains (C H 1. C H 2 and C H 3). Each light chain consists of a light chain variable region (V L ) and the light chain constant region (C L ). The light chain constant region consists of a domain C L The constant region of the antibody can mediate the binding of the immunoglobulin to host tissues or factors, including various cells of the immune system (e.g., effector cells) and the first component (C1q) of the classical complement system. H and V L The V region can be further subdivided into regions of high variability, termed complementarity determining regions (CDRs), interspersed with regions that are more conserved, termed framework regions (FRs). H and VL It consists of three CDRs and four FRs arranged from the amino terminus to the carboxyl terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. The variable region (V H and V L ) form antibody binding sites respectively. The allocation of amino acids to each region or domain follows the Kabat Sequences of Proteins of Immunological Interest (National Institutes of Health, Bethesda, Md. (1987 and 1991)), or the definitions of Chothia & Lesk (1987) J. Mol. Biol. 196:901-917; Chothia et al. (1989) Nature 342:878-883. The term "antibody" is not limited by any particular method for producing antibodies. For example, it includes, in particular, recombinant antibodies, monoclonal antibodies and polyclonal antibodies. Antibodies can be antibodies of different isotypes, for example, IgG (e.g., IgG1, IgG2, IgG3 or IgG4 subtypes), IgA1, IgA2, IgD, IgE or IgM antibodies.

[0023] In the present invention, the term "antigen binding portion" of an antibody refers to one or more portions of a full-length antibody that retain the ability to bind to the same antigen (e.g., PCSK9) to which the antibody binds, competing with the intact antibody for specific binding to the antigen. See generally, Fundamental Immunology, Ch. 7 (Paul, W., ed., 2nd edition, Raven Press, NY (1989), which is incorporated herein by reference in its entirety for all purposes. Antigen binding portions can be produced by recombinant DNA technology or by enzymatic or chemical cleavage of intact antibodies. In some cases, antigen binding portions include Fab, Fab', F(ab') 2、 Fd, Fv, dAb and complementarity determining region (CDR) fragments, single chain antibodies (e.g., scFv), chimeric antibodies, diabodies and polypeptides that contain at least a portion of an antibody sufficient to confer specific antigen binding ability to the polypeptide.

[0024] In the present invention, the term "Fd fragment" means a fragment consisting of V H and C H The term "Fv fragment" means an antibody fragment consisting of a single arm of an antibody V L and V H The term "dAb fragment" means an antibody fragment consisting of a VH The term "Fab fragment" refers to an antibody fragment consisting of V and V domains (Ward et al., Nature 341:544-546 (1989)); L 、V H , C L and C H The term "F(ab')2 fragment" refers to an antibody fragment comprising two Fab fragments linked by a disulfide bridge at the hinge region.

[0025] In some cases, the antigen-binding portion of the antibody is a single-chain antibody (e.g., scFv) in which V L and V H The domains are paired to form monovalent molecules via linkers that enable them to be produced as a single polypeptide chain (see, e.g., Bird et al., Science 242:423-426 (1988) and Huston et al., Proc. Natl. Acad. Sci. USA 85:5879-5883 (1988)). Such scFv molecules may have the general structure: NH2-V L -Connector-V H -COOH or NH2-V H -Connector-V L -COOH. Suitable prior art linkers (connecting peptides) consist of repeated GGGGS amino acid sequences or variants thereof. For example, a linker having the amino acid sequence (GGGGS) 4 may be used, but variants thereof may also be used (Holliger et al. (1993), Proc. Natl. Acad. Sci. USA 90: 6444-6448). Other linkers useful in the present invention are described by Alfthan et al. (1995), Protein Eng. 8: 725-731, Choi et al. (2001), Eur. J. Immunol. 31: 94-106, Hu et al. (1996), Cancer Res. 56: 3055-3061, Kipriyanov et al. (1999), J. Mol. Biol. 293: 41-56 and Roovers et al. (2001), Cancer Immunol. In an embodiment of the present invention, the sequence of the connecting peptide is (GGGGS)3.

[0026] In some cases, the antibody is a bispecific antibody, which can bind to two antigens or antigenic epitopes respectively, and includes a light chain, a heavy chain, or an antigen-binding portion thereof of an antibody that specifically binds to a first antigen, and a light chain, a heavy chain, or an antigen-binding portion thereof of an antibody that specifically binds to a second antigen. In an embodiment of the present invention, the light chain, the heavy chain, or an antigen-binding portion thereof of the antibody that binds to the first antigen in the bispecific antibody may be an antibody or an antigen-binding portion thereof according to any one of the present invention, and the light chain, the heavy chain, or an antigen-binding portion thereof of the antibody that specifically binds to the second antigen may be other anti-ICAM-1 antibodies or antigen-binding portions thereof or antibodies or antigen-binding portions thereof against other antigens.

[0027] In some cases, the antibody is a diabody, i.e., a bivalent antibody in which V H and V L The domains are expressed on a single polypeptide chain, but with a linker that is too short to allow pairing between the two domains on the same chain, thereby forcing the domains to pair with the complementary domains of another chain and create two antigen-binding sites (see, e.g., Holliger P. et al., Proc. Natl. Acad. Sci. USA 90:6444-6448 (1993), and Poljak RJ et al., Structure 2:1121-1123 (1994)).

[0028] Antigen-binding portions of antibodies (e.g., antibody fragments described above) can be obtained from a given antibody (e.g., monoclonal antibody 2E12) using conventional techniques known to those skilled in the art (e.g., recombinant DNA techniques or enzymatic or chemical cleavage methods) and screened for specificity in the same manner as for intact antibodies.

[0029] In the present invention, the antigen binding portion includes single chain antibody (scFv), chimeric antibody, diabody, scFv-Fc bivalent molecule, dAb and complementarity determining region (CDR) fragment, Fab fragment, Fd fragment, Fab' fragment, Fv and F(ab')2 fragment.

[0030] In the present invention, the λ light chain constant region includes various allotypes, such as λ I, λ II, λ III, and λ VI. In an embodiment of the present invention, the λ light chain constant region is of λ II type.

[0031] The antibody nucleic acid molecules involved in the present invention can also be obtained by conventional genetic engineering recombination technology or chemical synthesis methods. On the one hand, the sequence of the antibody nucleic acid molecules involved in the present invention includes the heavy chain variable region of the anti-ICAM-1 antibody or a partial nucleic acid sequence of the antibody molecule. On the other hand, the sequence of the antibody nucleic acid molecules involved in the present invention also includes the light chain variable region of the anti-ICAM-1 antibody or a partial nucleic acid sequence of the antibody molecule. On the other hand, the sequence of the antibody nucleic acid molecules involved in the present invention also includes the CDR sequence of the heavy chain or light chain variable region. The complementary determining region (CDR) is the site that binds to the antigen epitope. The CDR sequence in the present invention is determined by IMGT / V-QUEST (http: / / imgt.cines.fr / textes / vquest / ). However, the CDR sequences obtained by different division methods are slightly different.

[0032] The present invention relates to a recombinant expression vector containing the nucleic acid molecule, and also to a host cell transformed with the molecule. Moreover, the present invention also relates to a method for obtaining the antibody of the invention by culturing and isolating the host cell containing the nucleic acid molecule under specific conditions.

[0033] In the present invention, the term "vector" refers to a nucleic acid carrier into which a polynucleotide encoding a protein can be inserted and the protein can be expressed. The vector can be transformed, transduced or transfected into the host cell so that the genetic material elements it carries are expressed in the host cell. For example, vectors include: plasmids; phagemids; cosmids; artificial chromosomes such as yeast artificial chromosomes (YAC), bacterial artificial chromosomes (BAC) or P1-derived artificial chromosomes (PAC); bacteriophages such as λ phage or M13 phage and animal viruses. The types of animal viruses used as vectors include retroviruses (including lentiviruses), adenoviruses, adeno-associated viruses, herpes viruses (such as herpes simplex virus), poxviruses, baculoviruses, papillomaviruses, and papillomaviruses (such as SV40). A vector may contain a variety of elements for controlling expression, including promoter sequences, transcription start sequences, enhancer sequences, selection elements and reporter genes. In addition, the vector may also contain a replication start site. A vector may also include components that assist it in entering cells, such as viral particles, liposomes, or protein coats, but is not limited to these substances.

[0034] In the present invention, the term "host cell" refers to a cell into which a vector is introduced, including many cell types, such as prokaryotic cells such as Escherichia coli or Bacillus subtilis, fungal cells such as yeast cells or Aspergillus, insect cells such as S2 Drosophila cells or Sf9, or animal cells such as fibroblasts, CHO cells, COS cells, NSO cells, HeLa cells, BHK cells, HEK 293 cells or human cells.

[0035] The antibody fragments of the present invention can be obtained by hydrolyzing intact antibody molecules (see Morimoto et al., J. Biochem. Biophys. Methods 24: 107-117 (1992) and Brennan et al., Science 229: 81 (1985)). In addition, these antibody fragments can also be directly produced by recombinant host cells (reviewed in Hudson, Curr. Opin. Immunol. 11: 548-557 (1999); Little et al., Immunol. Today, 21: 364-370 (2000)). For example, Fab' fragments can be directly obtained from E. coli cells or chemically coupled to form F(ab')2 fragments (Carter et al., Bio / Technology, 10: 163-167 (1992)). For another example, F(ab')2 fragments can be obtained by connecting with leucine zipper GCN4. In addition, Fv, Fab or F(ab')2 fragments can also be directly isolated from the culture medium of recombinant host cells. Other techniques for preparing antibody fragments are well known to those skilled in the art.

[0036] In the present invention, "specific binding" refers to a non-random binding reaction between two molecules, such as the reaction between an antibody and the antigen that produced the antibody. Here, the antibody that binds to the first antigen has an undetectable or weak binding affinity for the second antigen. In certain embodiments, an antibody specific for an antigen is one that binds to the second antigen with an affinity (KD) ≤ 10 -5 M (e.g. 10 -6 M, 10 -7 M, 10 -8 M, 10 -9 M, 10 -10 M, etc.) binds to the antigen, wherein KD refers to the ratio of the dissociation rate to the binding rate (koff / kon), which can be determined by methods familiar to those skilled in the art.

[0037] In the present invention, the 20 conventional amino acids and their abbreviations follow conventional usage. See Immunology-A Synthesis (2nd edition, ES Golub and DR Gren, Eds., Sinauer Associates, Sunderland, Mass. (1991)), which is incorporated herein by reference. BRIEF DESCRIPTION OF THE DRAWINGS

[0038] Figure 1 The activity test results of the antibody in Example 1 of the present invention are shown in FIG. DETAILED DESCRIPTION

[0039] Example 1: Construction, screening and identification of anti-ICAM-1 phage single-chain antibody library 1. The phagemid vector pCANTAB5E and helper phage M13K07 were provided by our laboratory. Based on the known ICAM-1 sequence, the extracellular sequence of its transmembrane protein was found to achieve the expression and purification of ICAM-1 extracellular protein. The expressed and purified ICAM-1 ECD protein was immunized into BALB / c mice. After confirming that the immunization was successful, the mouse spleen was separated, the total RNA was extracted, and reverse transcribed into cDNA. The antibody light and heavy chain genes were amplified by PCR using specific degenerate primers, and then a single-chain antibody phage library was constructed. After multiple rounds of screening, the anti-ICAM-1 phage scFv antibody was obtained. The antibody gene sequence was: , and the antibody chain composition was V H -linker-V L , VH The sequence is QVKLQQSGADLVKPGASVMLSCKAS GYTFTSYY MYWVKQRPGQGLEWIGE INPSNGGT NFNEKFKTKATLTVDRSSSTAYMQLSNLTSEDSAVYYC I AM TRQAGYY WGQGTTVTVSS (the underline indicates CDR1-3, the same below), V L The sequence is DIELTQSPAIMSASPGEKVTMTCRAS SSVSSSY LHWYQQKSGASPKLLIY RTS NLASGVPARFSGSGSGTSYSLTVSSVEAEDAATYYC QQYSGYPLFT FGSGTKLEIKR, the linker is GGGGSGGGGSGGGGS; The above scFv antibody gene was used to construct an expression vector, which was expressed and purified in E. coli. Western Blot showed that the recombinant protein was successfully expressed and was used for further activity confirmation below.

[0040] 2. Elisa detection of the affinity activity of anti-ICAM-1 single-chain antibody and antigen (1) Coating: Dilute the ICAM-1 ECD protein with coating solution (pH = 9.6) and add 100 μL of protein solution to each well of the ELISA plate and incubate at 4°C overnight.

[0041] (2) Washing: Discard the coating solution, add 200 μL PBST to each well, wash for 5 min, and wash 3 times in total.

[0042] (3) Blocking: Add 200 μL of blocking solution to each well and incubate at 37°C for 2 h.

[0043] (4) Repeat step (2).

[0044] (5) Incubation with primary antibody: Add 100 μL of diluted scFv antibody to each well and incubate at 37°C for 2 h.

[0045] (6) Repeat step (2).

[0046] (7) Incubation with secondary antibody: Add 100 μL of HRP-labeled mouse anti-His antibody to each well and incubate at 37°C for 1 h.

[0047] (8) Repeat step (2).

[0048] (9) Color development: Add 100 μL TMB color development solution to each well and incubate at 37°C for 15-30 min. Then add 100 μL Elisa stop solution to each well.

[0049] (10) Reading: Open the multifunctional microplate reader in advance, set the parameters, and record the OD450 value. Figure 1 .

[0050] The concentrations of ICAM-1 ECD antigen were 1 μg / mL and 5 μg / mL, respectively. Substituting IC50 into the affinity constant calculation formula, the affinity constant of the scFv was calculated to be 1.02×10 6 L / mol.

Claims

1. A monoclonal antibody or an antigen-binding portion thereof, which can bind to ICAM-1 and comprises a heavy chain variable region and a light chain variable region, wherein the amino acid sequences of the CDR1-3 regions of the heavy chain variable region are shown in SEQ ID NOs: 1-3, and the amino acid sequences of the CDR1-3 regions of the light chain variable region are shown in SEQ ID NOs: 4-6.

2. The monoclonal antibody or antigen-binding portion thereof of claim 1, wherein the amino acid sequence of the heavy chain variable region is as shown in SEQ ID NO:

7.

3. The monoclonal antibody or antigen-binding portion thereof according to claim 1, wherein the amino acid sequence of the light chain variable region is shown in SEQ ID NO:

8.

4. The monoclonal antibody or antigen binding portion thereof according to any one of claims 1 to 3, which is a complete antibody, a bispecific antibody, a scFv, a Fab, a Fab', a F(ab')2 or a Fv.

5. The monoclonal antibody or antigen-binding portion thereof according to the preceding claim, which is a scFv, and the connecting peptide between the heavy chain variable region and the light chain variable region is (GGGS) n C. (GGGGS) n 、(GGSC) n or (GGG) n , where n=1-10, such as 3.

6. The monoclonal antibody or antigen-binding portion thereof according to the preceding claim, wherein the C-terminus of the heavy chain variable region is connected to the N-terminus of the light chain variable region via the connecting peptide.

7. A nucleic acid molecule comprising a sequence encoding the monoclonal antibody or antigen binding portion thereof according to any one of claims 1 to 6.

8. A recombinant cell comprising the nucleic acid molecule according to the preceding claim, preferably, the cell is a mammalian cell or a bacterium, a fungus, such as an Ecoli, a yeast or a CHO cell.

9. A pharmaceutical composition comprising the monoclonal antibody or antigen-binding portion thereof according to any one of claims 1 to 6, the nucleic acid molecule according to claim 7, or the recombinant cell according to claim 8.

10. Use of a monoclonal antibody or antigen binding portion thereof as claimed in any preceding claim in the preparation of a diagnostic reagent or a medicament for treating solid tumours or dry eye.

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