Bispecific fully-humanized single-domain antibody targeting new coronavirus and CD16a and application of bispecific fully-humanized single-domain antibody

Abstract

The invention discloses a bispecific fully-humanized single-domain antibody targeting a new coronavirus and CD16a and an application of the bispecific fully-humanized single-domain antibody. The antibody comprises a single-domain antibody n3130v targeting a new coronavirus and a single-domain antibody n118 targeting a human CD16a, the amino acid sequence of the n3130v is as shown in SEQ ID NO: 1, and the amino acid sequence of the n118 is as shown in SEQ ID NO: 2. The antibody disclosed by the invention has the capabilities of broad-spectrum recognition and neutralization of novel coronavirus, can be efficiently combined with a CD16a target spot, and is used for mediating antibody-dependent cell-mediated cytotoxic action (ADCC). The bispecific antibody can be used for novel coronavirus related research and treatment of diseases caused by novel coronavirus infection.

Description

Technical Field

[0001] The present invention relates to a bispecific fully human single-domain antibody targeting the new coronavirus and CD16a and its application, belonging to the field of biomedicine technology. Background Art

[0002] The binding of the SARS-CoV-2 receptor binding domain (RBD) to human angiotensin converting enzyme 2 (ACE2) on the cell surface is the first step in viral infection of the host and is therefore a key target for antibody development. Five variants of concern (VOC) have emerged in the SARS-CoV-2, including Alpha, Beta, Gamma, Delta, and Omicron. Since the emergence of the Omicron variant, numerous mutant substrains have emerged and have continuously iterated within the circulating strains, resulting in severe immune escape and recurrent infection. The Omicron spike protein has accumulated dozens of mutation sites, posing a significant challenge to the development of broad-spectrum antibodies.

[0003] Fully human single-domain antibodies (SDOs) are antibody fragments consisting solely of the variable region of the heavy chain (VH). Their molecular weight is only one-tenth that of traditional monoclonal antibodies. They are highly efficient in binding to antibodies, exhibit stable properties, exhibit high solubility, and can be expressed in prokaryotes.

[0004] CD16a, or FcγRIIIA, is primarily expressed on macrophages, monocytes, and natural killer (NK) cells and is a major cytotoxic receptor. Binding of the Fc region of antigen-bound IgG to CD16a on the surface of immune cells can trigger cytokine release or antibody-dependent cytotoxicity, leading to antigen recognition and killing by immune cells.

[0005] Based on this, the bispecific antibody developed by the present invention will activate immune cells and recognize and eliminate viruses by directly binding to CD16a antibodies. Summary of the Invention

[0006] The present invention aims to solve the above-mentioned technical problems and provides two bispecific fully human single-domain antibodies that can effectively and broadly neutralize the new coronavirus while binding to CD16a on the surface of immune cells.

[0007] To achieve the above objectives, the present invention provides a bispecific antibody targeting the new coronavirus and CD16a, wherein the antibody comprises a single-domain antibody n3130v targeting the new coronavirus and a single-domain antibody n118 targeting human CD16a, the amino acid sequence of n3130v is shown in SEQ ID NO: 1, and the amino acid sequence of n118 is shown in SEQ ID NO: 2.

[0008] Preferably, the bispecific antibody comprises the amino acid sequence shown in SEQ ID NO: 1 or a sequence that has at least 90% identity and functionally the same as the amino acid sequence shown in SEQ ID NO: 2, and the amino acid sequence shown in SEQ ID NO: 2 or a sequence that has at least 90% identity and functionally the same as the amino acid sequence shown in SEQ ID NO: 2.

[0009] Preferably, the bispecific antibody is connected by the N-terminus of the n118 antibody to the C-terminus of the n3130v antibody via a peptide linker.

[0010] Preferably, the linker peptide sequence is (Gly4Ser)3 and (Gly4Ser)4.

[0011] Preferably, the single-domain antibody consists of a heavy chain variable region (VH).

[0012] Preferably, the heavy chain variable region (VH) is fully human.

[0013] Preferably, the sequence of the antibody is as shown in SEQ ID NO: 3 or SEQ ID NO: 4, or a sequence of a protein having the same function obtained by replacing, deleting or inserting one or more amino acids in SEQ ID NO: 3 or SEQ ID NO: 4.

[0014] The present invention also provides a fusion protein comprising the bispecific antibody described above and a heterologous protein.

[0015] Preferably, the heterologous protein is human immunoglobulin Fc, more preferably human IgG1 Fc.

[0016] The present invention provides a conjugate comprising the above-mentioned bispecific antibody or the above-mentioned fusion protein coupled with an effector molecule.

[0017] Preferably, the effector molecule is a detectable label.

[0018] Preferably, the detectable label is a fluorescent label, a radioactive label, avidin, biotin, or an enzyme.

[0019] The present invention provides nucleic acids encoding the bispecific antibodies that bind to the novel coronavirus and CD16a as described above. The present invention provides vectors comprising the nucleic acid, optionally operably linked to regulatory sequences. The present invention provides host cells comprising the vector, and methods for producing and optionally recovering the bispecific antibodies or antigen-binding fragments. The host cells of the present invention can be any prokaryotic or eukaryotic cells, including but not limited to bacterial cells (e.g., Escherichia coli), insect cells (e.g., using a baculovirus expression system), yeast or mammalian cells (e.g., CHO or BHK cell lines). Other suitable host cells are known to those skilled in the art. Preferably, the host cell is an Escherichia coli cell, which has a short culture cycle and low production cost.

[0020] The present invention provides a pharmaceutical composition comprising a prophylactic or therapeutic dose of the anti-COVID-19 and human CD16a bispecific antibody, or the aforementioned fusion protein, or the aforementioned conjugate, or the aforementioned nucleic acid molecule, or the aforementioned vector, mixed with a physiologically or pharmaceutically acceptable carrier, excipient, or stabilizer, and other pharmaceutically acceptable excipients, the composition including but not limited to a lyophilized dosage form, an aqueous solution dosage form, a liposome, or a capsule dosage form. The concentration of the human CD16a bispecific antibody, fusion protein, conjugate, bispecific antibody nucleic acid molecule, or vector comprising the bispecific antibody nucleic acid molecule of the present invention can vary from about 0.1% to 100% (by weight).

[0021] The present invention provides an immunotherapy method for treating or improving viral infection, which uses bispecific antibodies to mediate immune cell recognition of viruses and exert antibody-dependent cell-mediated cytotoxicity (ADCC).

[0022] The present invention provides the use of the aforementioned bispecific antibody, or the aforementioned fusion protein, or the aforementioned conjugate, or the aforementioned nucleic acid molecule, or the aforementioned vector, or the aforementioned pharmaceutical composition in the preparation of a medicament for treating and / or preventing the novel coronavirus or a disease caused by infection with the novel coronavirus.

[0023] The present invention provides the use of the aforementioned bispecific antibody, or the aforementioned fusion protein, or the aforementioned conjugate, or the aforementioned nucleic acid molecule, or the aforementioned vector, or the aforementioned pharmaceutical composition in the preparation of a diagnostic product or a detection product for detecting and / or diagnosing coronavirus infection. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 : Schematic diagram of the structure of bispecific antibodies against the new coronavirus.

[0025] Figure 2 : SDS-PAGE verification of the purity of the bispecific antibody.

[0026] Figure 3 : ELISA detects the affinity of bispecific antibodies to the RBD protein of the new coronavirus variant and to human CD16a.

[0027] Figure 4 : Bispecific antibody-dependent cellular cytotoxicity (ADCC) activity.

[0028] Figure 5 : Neutralizing activity of bispecific antibodies against SARS-CoV-2 pseudovirus. DETAILED DESCRIPTION

[0029] To make the present invention more clearly understood, preferred embodiments are described in detail below with reference to the accompanying drawings.

[0030] In order to better understand the present invention, some definitions are listed below. The above definitions are intended to include grammatical equivalents.

[0031] "CD16" herein is also referred to as Fcγ receptor III (FcγRIII), which is expressed from different genes in two forms: FcγRIIIa / CD16a and FcγRIIIb / CD16b. CD16a is an integral membrane protein expressed primarily on NK cells and monocytes. ADCC via CD16a is one of the primary mechanisms by which NK cells exert their effector functions.

[0032] The "spike protein" in this article refers to the key protein on the surface of the new coronavirus used to identify and invade target cells, also known as the Spike protein.

[0033] As used herein, "immune cells" refer to cells that have hematopoietic origin and play a role in immune responses. Immune cells include myeloid cells, such as monocytes, macrophages, and lymphocytes, such as B cells, T cells, and NK cells.

[0034] "Immunoglobulin Fc" refers to an antibody heavy chain fragment composed of CH2 and CH3 of an antibody. Fc can be selected from human IgG1, IgG2, IgG3, IgG4, or the corresponding domains after mutation of one or several sites therein.

[0035] As used herein, "antibody" refers to a protein composed of one or more domains encoded by all or part of substantially all recognized immunoglobulin genes. The recognized immunoglobulin genes, for example, in humans, include kappa (κ), lambda (λ), and heavy chain loci, which contain numerous variable region genes, as well as constant region genes mu (μ), delta (δ), gamma (γ), epsilon (ε), and alpha (α), encoding the IgM, IgD, IgG, IgE, and IgA isotypes, respectively. Antibodies herein are intended to include full-length antibodies, individual chains thereof, and all parts, domains, or fragments thereof, as well as natural antibodies from any organism, engineered antibodies, or antibodies recombinantly produced for experimental, therapeutic, or other purposes as further specified below. The term "antibody" includes antibody fragments, as known in the art, such as Fab, Fab', F(ab')2, Fv, scFv, or the antigen-binding domains of antibodies (e.g., VHH domains or VH / VL domains), or antibody fragments produced by modifying intact antibodies or those synthesized de novo using recombinant DNA technology. The term "antibody" includes monoclonal and polyclonal antibodies. Antibodies can be antagonists, agonists, neutralizing antibodies, inhibitory antibodies, or stimulatory antibodies. Antibodies of the present invention can be non-human antibodies, chimeric antibodies, humanized antibodies, or fully human antibodies.

[0036] As used herein, the term "heavy chain variable region (VH)" refers to the region of the immunoglobulin heavy chain near the N-terminus where the amino acid sequence varies significantly, and is the heavy chain variable region domain in conventional four-chain antibodies. It is particularly used to distinguish VHH antibodies from Camelidae.

[0037] As used herein, "bispecific antibody" refers to an artificial antibody that contains two specific antigen-binding sites.

[0038] As used herein, a "single domain antibody" refers to an immunoglobulin variable domain that is capable of specifically binding to an antigenic epitope without pairing with other immunoglobulin variable domains.

[0039] As used herein, "fully human" means that the antibodies have high homology with human antibodies and are entirely encoded by human antibody genes.

[0040] As used herein, "amino acid" means one of the 20 naturally occurring amino acids or any non-natural analogs, which may be located at a specifically defined position. As used herein, "protein" means at least two covalently linked amino acids, which include proteins, polypeptides, oligopeptides, and peptides. Proteins can be composed of naturally occurring amino acids and peptide bonds, or of synthetic peptide mimetic structures, which are "analogs." Therefore, as used herein, "amino acid" or "peptide residue" means both naturally occurring and synthetic amino acids. For example, for the purposes of the present invention, homophenylalanine, citrulline, and norleucine are considered amino acids for the purposes of the present invention. "Amino acid" also includes imino acid residues such as proline and hydroxyproline. The side chains may be in the (R) or (S) configuration. In preferred embodiments, amino acids are present in the (S) or L- configuration. If non-naturally occurring side chains are used, non-amino acid substitutions may be used, for example to prevent or delay degradation in vivo.

[0041] The "peptide linker" used herein refers to the (Gly4Ser)3 and (Gly4Ser)4 polypeptide segments that connect n3130v and n118 to form the bispecific antibody.

[0042] " nucleic acid " used herein means the polymer consisting of nucleotide units (ribonucleotides, deoxyribonucleotides, related naturally occurring structural variants and the analogs of the non-naturally occurring syntheses thereof) by phosphodiester bonds. Therefore, the term includes nucleotide polymers, wherein nucleotide and the bond therebetween include non-naturally occurring synthetic analogs, such as but not limited to phosphorothioate, phosphoramidate, methyl phosphonate, chiral methyl phosphonate, 2'-O-methyl ribonucleotides, peptide nucleic acid (PNA) etc. For example, automatic DNA synthesizer can be used to synthesize these polynucleotides. The term " oligonucleotide " generally refers to short polynucleotides, generally not more than about 50 nucleotides. It should be understood that when nucleotide sequence is represented by DNA sequence (i.e. A, T, G, C), this also includes the RNA sequence (i.e. A, U, G, C) in which " U " replaces " T ".

[0043] As used herein, "vector" refers to a vector artificially constructed from a natural plasmid to adapt to laboratory manipulation. A nucleic acid molecule can be introduced into a host cell, thereby producing a transformed host cell. A vector may include a nucleic acid sequence that permits replication in the host cell, such as an origin of replication, and may also include one or more selectable marker genes and other genetic elements known in the art.

[0044] As used herein, "host cell" is also called a recipient cell and can refer to a cell that is recognized and infected through a specific target, or a cell that is transferred with a specific plasmid or vector.

[0045] As used herein, " encoding " means the inherent properties of a specific nucleotide sequence in a polynucleotide, such as a gene, cDNA or mRNA, used as a template for synthesizing other polymers and macromolecules in a biological process with a determined nucleotide sequence, or a determined amino acid sequence and the resulting biological properties. Therefore, if the transcription and translation of the mRNA produced by the gene produce a protein in a cell or other biological system, the gene encodes a protein. Coding strands (its nucleotide sequence is identical to the mRNA sequence and is generally provided in a sequence table) and non-coding strands (used as transcription templates, genes or cDNAs) can be referred to as encoded proteins. Or other products of the gene or cDNA. Unless otherwise indicated, " nucleotide sequences encoding amino acid sequences " include all nucleotide sequences that are degenerate forms of each other and encode the same amino acid sequence. The nucleotide sequences of encoded proteins and RNA may include introns.

[0046] As used herein, "pharmaceutically acceptable carrier" means any and all solvents that are physiologically compatible. Remington's Pharmaceutical Sciences, E.W. Martin, Mack Publishing Co., Easton, Pa., 15th ed. (1975), describes compositions and formulations suitable for the pharmaceutical delivery of one or more therapeutic compounds or molecules (e.g., one or more antibodies), as well as additional pharmaceutical agents.

[0047] As used herein, an "effective preventive or therapeutic dose" means a specific agent sufficient to achieve the desired effect in a subject treated with the bispecific antibody. The exact dosage will depend on the purpose of the treatment and can be determined by those skilled in the art using known techniques. The dosage range may be 0.01-100 mg / kg body weight or greater, for example 0.1, 1, 10 or 50 mg / kg body weight, preferably 1-10 mg / kg. As is known in the art, adjustments may be necessary for the distribution, metabolism, excretion processes of the antibody drug throughout the body, as well as age, weight, general health status, sex, diet, administration time, drug interactions, and severity of the disease, and can be determined by those skilled in the art through conventional experimental methods. Ideally, a therapeutically effective amount of an antibody is an amount sufficient to prevent, treat, or ameliorate an infection or disease. The therapeutically effective amount of an agent used to prevent, ameliorate, and / or treat a subject will depend on the subject being treated, the type and severity of the pain, and the mode of administration of the therapeutic composition.

[0048] Unless otherwise indicated, all technical and scientific terms used herein have the same meaning as those generally understood by those of ordinary skill in the art to which this disclosure belongs. Unless the context clearly indicates otherwise, the singular terms "a", "an" and "the" include plural indicators. It should also be understood that all base sizes or amino acid sizes given for nucleic acids or polypeptides, and all molecular weights or molecular weight values are approximate and are provided for description. Although methods and materials similar or equivalent to those described herein can be used for the practice or testing of the present disclosure, suitable methods and materials are described below. The term "comprising" means "including". All publications, patent applications, patents and other references mentioned herein are incorporated by reference in their entirety. In the event of a conflict, this specification (including term explanations) will prevail. In addition, materials, methods and examples are illustrative only and not restrictive.

[0049] The experimental methods in the examples of the present invention, for which specific conditions are not specified, are generally carried out under conventional conditions or according to the conditions recommended by the manufacturer. The standard recombinant DNA technology and molecular cloning techniques used in the examples of the present invention are referenced to those well known in the art (Ausubel, FM et al., Current Protocols in Molecular Biology, published by Greene Publishing Assoc. and Wiley-Interscience). Materials and methods suitable for microbial growth are well known in the art; the materials, reagents, etc. used in the examples, unless otherwise specified, are reagents and materials obtained from commercial sources. The main chemical and biological reagents were purchased from KAPABiosystems, New England Biolabs, TransGen Biotech, Thermo Fisher Scientific, OMEGA bio-tek, etc.

[0050] Example 1 Bispecific Antibody Gene Synthesis

[0051] The single-domain antibody n3130v against the new coronavirus (SEQ ID NO: 1) and the single-domain antibody n118 against human CD16a (SEQ ID NO: 2) were fused using peptide linkers (Gly4Ser)3 and (Gly4Ser)4, i.e., n118-(GGGGS)3-n3130v and n118-(GGGGS)4-n3130v. The genes were synthesized by GenScript. The structural diagram is shown in the figure. Figure 1 shown.

[0052] Example 2 Construction of bispecific antibody expression vector

[0053] The synthesized bispecific antibody genes (n118-(GGGGS)3-n3130v and n118-(GGGGS)4-n3130v) were cloned into the prokaryotic expression vector pComb3x (Addgene, product number: 63891) using homologous recombination enzymes. The specific process was as follows: the gene fragment and the pComb3x vector linearized with the sfiI restriction endonuclease were mixed at a molar ratio of 3:1. 1 μL of homologous recombination enzyme and 4 μL of 5x buffer were added. The reaction system was made up to 20 μL with ultrapure water and incubated at 50°C in a metal bath for 20 minutes. The homologous recombination products were transformed into Escherichia coli TOP10 competent cells using the heat shock method and plated on ampicillin-resistant plates for overnight screening. The next day, single colonies were picked and sequenced to obtain the correct bispecific antibody expression vector.

[0054] Example 3 Expression and purification of bispecific antibodies

[0055] The bispecific antibody expression vector was transformed into competent E. coli HB2151 cells using the heat shock method. The next day, a single colony was picked and inoculated into SB medium for amplification at 37°C and 250 rpm. When the OD600 absorbance of the culture reached 0.8-1.0, 1 mM IPTG was added for induction at 30°C for 16-20 hours. The culture was then centrifuged at 8000 rpm to collect the solution, resuspend it in PBS buffer containing 0.5 M sodium chloride, and disrupt the cells using a high-pressure homogenizer. The supernatant was collected by centrifugation at 1000 rpm and the bispecific antibody was purified using Protein A resin. Purification procedures were described in the Protein A resin manual (GenScript).

[0056] Example 4: Purity of bispecific antibodies detected by polyacrylamide gel electrophoresis (SDS-PAGE)

[0057] The purified bispecific antibody protein was mixed with SDS-PAGE loading buffer and boiled at 95°C for 5-10 minutes. Electrophoresis was performed at a constant voltage of 80 V for 15 minutes, followed by 60 minutes at 120 V. After electrophoresis, the gel was stained with 0.25% Coomassie Brilliant Blue for 30 minutes and then destained in a destaining buffer. Figure 2 The band of the bispecific antibody was around 30 kD, which was in line with expectations. The band was single and had good purity.

[0058] Example 5 Bispecific antibodies bind to the RBD protein of the new coronavirus variant and the human CD16a protein respectively

[0059] The SARS-CoV-2 variant RBD protein (SinoBiological) and human CD16a protein (purified) were coated on an ELISA plate at a concentration of 2 μg / mL and incubated overnight at 4°C. The next day, 5% BSA-PBS solution was added for blocking for 1 hour. The bispecific antibody with a starting concentration of 2 μM (binding to RBD protein) and 3.3 μM (binding to human CD16a protein) was added and incubated at 37°C for 1.5 hours. The plate was washed three times with 0.05% PBST and the affinity was detected with a 1:5000 dilution of anti-Flag-HRP antibody (Sigma). Figure 3 As shown, the bispecific antibody has strong binding ability to the wild type and human CD16a, with no difference from the positive control group.

[0060] Example 6 Bispecific Antibody-Dependent Cellular Cytotoxicity (ADCC) Activity Assay

[0061] The recombinant plasmid of SARS-CoV-2 variant RBD protein fused to the transmembrane domain was transfected into HEK293T cells. After 48 hours, the cells were counted and collected. The target cells were plated at 7.5×10 per well. 4 Dilute 100 cells into 1640 medium containing 10% FBS, 30 μL per well, and dilute the antibody into 1640 medium at a starting concentration of 1.67 μM, 2-fold concentration gradient dilution, 30 μL per well, and mix the antibody and target cells. Human CD16a (158V) (luciferase) Jurkat reporter gene cells were cultured at 5×10 4 Dilute 1640 cells per well into 1640 culture medium, and add 30 μL of the mixture of antibodies and target cells to each well and incubate for 5 hours. Take 30 μL of cell mixture and 30 μL of ONE-Glo TM The luciferase reagent was mixed and shaken at room temperature for 4 minutes to achieve complete lysis. The fluorescence (RLU) value was read using a microplate reader, and the curve fitting analysis was performed using GraphPad Prism software by nonlinear regression (logarithmic dose-response curve, four-parameter variable slope). The results are shown in Figure 4 As shown, bispecific antibodies mediate immune cell recognition of viruses and exert antibody-dependent cell-mediated cytotoxicity (ADCC).

[0062] Example 7 Determination of Bispecific Antibody Pseudovirus Neutralization Activity

[0063] HEK293T cells were co-transfected with a recombinant plasmid encoding the SARS-CoV-2 variant S protein and a luciferase reporter gene-deleted HIV-1 backbone plasmid. After 48 hours, the supernatant was harvested, filtered, and stored at -80°C until further use. Huh-7 cells (10,000 cells / well) were plated in 96-well plates and incubated at 37°C, 5% CO₂, for 12 hours before use. n3130v and the bispecific antibody were serially diluted threefold in serum-free cell culture medium. Then, 50 μL of antibody was mixed with 50 μL of virus. After incubation at 37°C for 1 hour, the cells were added to Huh-7 cells, including control wells containing virus alone and no antibody, as well as wells containing background cells without virus or antibody. After infection for 8 hours at 37°C, 5% CO₂, the medium was replaced with fresh medium and cultured for an additional 48 hours. The cell culture medium in the 96-well plates was discarded, and the cells were washed twice with PBS. Relative fluorescence in the cell lysates was measured using a Luciferase Assay Kit (Promega). Inhibition rate = 100*(1-(antibody sample fluorescence value-background fluorescence mean) / (virus infection fluorescence value-background fluorescence mean)), the results are as follows Figure 5 As shown, the neutralizing activity of the bispecific antibody against the mutant strain had no significant difference from that of the positive control n3130v, indicating that its neutralizing activity was strong.

[0064] The sequences involved in the present invention are as follows:

[0065] n3130v variable heavy chain (SEQ ID NO: 1)

[0066]

[0067] n118 variable heavy chain (SEQ ID NO: 2):

[0068]

[0069]

[0070] Bispecific antibody n118-(GGGGS)3-n3130v (SEQ ID NO: 3):

[0071]

[0072] Bispecific antibody n118-(GGGGS)4-n3130v (SEQ ID NO: 4):

[0073]

[0074]

[0075] The above description is only a preferred embodiment of the present invention and does not constitute any formal or substantial limitation to the present invention. It should be noted that ordinary technicians in this technical field can make several improvements and supplements without departing from the present invention, and these improvements and supplements should also be regarded as the scope of protection of the present invention.

Claims

1. A bispecific antibody targeting novel coronavirus and CD16a, characterized in that: The antibody comprises a single-domain antibody n3130v targeting the new coronavirus and a single-domain antibody n118 targeting human CD16a. The amino acid sequence of n3130v is shown in SEQ ID NO: 1, and the amino acid sequence of n118 is shown in SEQ ID NO:

2.

2. The bispecific antibody according to claim 1, wherein The bispecific antibody comprises an amino acid sequence shown in SEQ ID NO: 1 or a sequence having at least 90% identity and functionally identical thereto, and an amino acid sequence shown in SEQ ID NO: 2 or a sequence having at least 90% identity and functionally identical thereto.

3. The bispecific antibody according to claim 1, wherein The bispecific antibody is connected by the N-terminus of the n118 antibody and the C-terminus of the n3130v antibody via a peptide linker.

4. The bispecific antibody according to claim 3, wherein The peptide linker is (Gly4Ser)3 or (Gly4Ser)4.

5. The bispecific antibody according to any one of claims 1 to 4, wherein The single-domain antibody is composed of a heavy chain variable region (VH), which is fully human; the sequence of the antibody is as shown in SEQ ID NO: 3 or SEQ ID NO: 4, or a sequence of a protein with the same function obtained by replacing, deleting or inserting one or more amino acids in SEQ ID NO: 3 or SEQ ID NO:

4.

6. A fusion protein, characterized in that The invention comprises the bispecific antibody according to any one of claims 1 to 4, fused with a heterologous protein.

7. The fusion protein according to claim 6, wherein The heterologous protein is immunoglobulin Fc.

8. The fusion protein according to claim 7, wherein The heterologous protein is Fc of IgG1.

9. A conjugate, characterized in that The bispecific antibody according to any one of claims 1 to 4, or the fusion protein according to any one of claims 6 to 8, is coupled to an effector molecule.

10. The conjugate according to claim 9, wherein The effector molecule is a detectable label.

11. The conjugate according to claim 10, wherein The detectable label is a fluorescent label, a radioactive label, avidin, biotin, or an enzyme.

12. A nucleic acid molecule, characterized in that It encodes the bispecific antibody according to any one of claims 1 to 4, or the fusion protein according to any one of claims 7 to 9.

13. A carrier, characterized in that Comprising the nucleic acid molecule of claim 12.

14. A host cell, characterized in that Comprising the nucleic acid molecule according to claim 12 or the vector according to claim 13.

15. A pharmaceutical composition, characterized in that Containing an effective preventive or therapeutic dose of the bispecific antibody according to any one of claims 1 to 4, or the fusion protein according to claim 6, or the conjugate according to claim 9, or the nucleic acid molecule according to claim 12, or the vector according to claim 13, and a pharmaceutically acceptable carrier.

16. Use of the bispecific antibody according to any one of claims 1 to 4, or the fusion protein according to claim 6, or the conjugate according to claim 9, or the nucleic acid molecule according to claim 12, or the vector according to claim 13, or the pharmaceutical composition according to claim 15 in the preparation of a medicament for treating and / or preventing a novel coronavirus or a disease caused by infection with the novel coronavirus.

17. Use of the bispecific antibody according to any one of claims 1 to 4, or the fusion protein according to claim 6, or the conjugate according to claim 9, or the nucleic acid molecule according to claim 12, or the vector according to claim 13, or the pharmaceutical composition according to claim 15 in the preparation of a diagnostic product or a test product for detecting and / or diagnosing coronavirus infection.

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