Bispecific antibody and application thereof

Abstract

The invention relates to the technical field of biology, in particular to a bispecific antibody and application thereof. The invention provides a bispecific antibody which has neutralizing activity on an SARS-CoV-2 mutant strain (VOCs) and an SARS-related coronavirus (SARSr-CoVs), and the bispecific antibody is constructed by fusing single-chain variable fragments of the two antibodies through a GS linker. The bispecific antibody shows high binding capacity to S1, and has broad-spectrum and efficient activity to pseudovirus SARS-CoV-2 variants and SARS-related coronaviruses, so that the bispecific antibody provided by the invention has the potential of preventing and treating the SARS-CoV-2 and the SARS-related coronaviruses.

Description

Technical Field

[0001] The present invention relates to the field of biotechnology, and in particular to a bispecific antibody and application thereof. Background Art

[0002] The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused enormous global health and socioeconomic losses. SARS-CoV-2 has evolved rapidly, undergoing multiple lineages and variants. Vaccines and antibodies developed based on the original virus strain have become less effective against emerging variants. In addition, several recently discovered SARS-related coronaviruses (SARSr-CoVs) have significant genetic similarities with SARS-CoV and SARS-CoV-2, increasing their potential for transmission to humans. Therefore, it is critical to prepare strategies to respond to evolving SARS-CoV-2 variants in advance. Vaccination faces challenges related to accessibility and unresolved side effects, which may further affect its overall effectiveness. To date, the continuous emergence of variants of concern (VOCs) has challenged vaccines developed using sequences of the original SARS-CoV-2 virus strain (wild type). For example, the Delta variant (B.1.617.2) and its spread have been mainly attributed to its significant ability to evade neutralizing antibodies, which are present in recovered and vaccinated populations. The Omicron variant (BA.1) shows a higher propensity to mutate. Subsequently, a series of Omicron subvariants emerged, characterized by a set of mutations in the receptor binding domain (RBD) of the spike (S) protein. These mutations result in alterations in the RBD that may affect its binding affinity to human angiotensin-converting enzyme 2 (hACE2). Notably, the Omicron RBD was reported to have a lower affinity for ACE2 than the Alpha or Delta variants. The main transmission factor of Omicron is its enhanced ability to evade polyclonal antibodies, thereby reducing the potency of serum antibody binding and neutralization.

[0003] Although monoclonal antibodies (mAbs) have been considered as ideal tools to combat coronaviruses due to their proven broad-spectrum neutralizing activity and potency, the continuous emergence of mutations has posed a major challenge to existing mAbs. Bispecific antibodies (bsAbs) are artificially engineered antibodies that are able to bind to two epitopes of an antigen or two different antigens simultaneously. In order to construct bsAbs with broad-spectrum neutralizing activity, a scaffold with the insertion of a natural VH-CH1 switch region is a promising approach. Several bsAb formats targeting SARS-CoV-2 have been developed, including two immunoglobulin G (IgG) single-chain variable fragments and single-chain variable fragments (scFvs) fused to an Fc domain. Although these bsAbs exhibit enhanced neutralization against pseudoviruses and real VOCs, the scope of their potency remains insufficiently defined. However, the development of bsAbs with strong neutralizing activity and broad-spectrum coverage is considered an alternative solution to cope with emerging variants. Summary of the invention

[0004] The purpose of the present invention is to overcome the shortcomings of the prior art and provide a bispecific antibody and its application, which has broad-spectrum neutralizing activity against SARS-CoV-2 mutants and SARS-related coronaviruses.

[0005] To achieve the above object, the technical solution adopted by the present invention is:

[0006] In the first aspect, the present invention provides a bispecific antibody, comprising a first antigen-binding arm and a second antigen-binding arm; the first antigen-binding arm comprises a heavy chain variable region VH1 and a light chain variable region VL1; the second antigen-binding arm comprises a heavy chain variable region VH2 and a light chain variable region VL2; the amino acid sequence of the VH1 is shown in SEQ ID NO: 1; the amino acid sequence of the VL1 is shown in SEQ ID NO: 2; the amino acid sequence of the VH2 is shown in SEQ ID NO: 3; the amino acid sequence of the VL2 is shown in SEQ ID NO: 4.

[0007] The first antigen-binding arm of the bispecific antibody designed in the present invention is selected from the heavy chain variable region (H chain FV) sequence SEQ ID NO: 10 of one of the parent antibodies (the antibody can broadly neutralize SARS-related coronaviruses) and the light chain variable region (L chain FV) sequence SEQ ID NO: 11 of the parent antibody.

[0008] The second antigen binding arm is selected from the heavy chain variable region (H chain FV) sequence of another parent antibody (which can neutralize SARS-CoV-2 variants broadly): SEQ ID NO: 12; and the light chain variable region (L chain FV) sequence of the parent antibody: SEQ ID NO: 13. The first binding arm of the designed bispecific antibody targets the RBD binding domain of SARS-CoV-2, and the second binding arm targets the shared antigenic epitope of SARS-related coronaviruses. This specific antibody improves the neutralization ability by binding to the key regions of the two viruses at the same time, preventing the virus from entering the host cell.

[0009] Preferably, the first antigen binding arm of the bispecific antibody specifically binds to the RBD receptor binding domain of SARS-CoV-2, and the second antigen binding arm specifically binds to the shared antigenic epitope of SARS-related coronaviruses.

[0010] Preferably, the amino acid sequence of the heavy chain constant region of the bispecific antibody is as shown in SEQ ID NO:5, and the amino acid sequence of the light chain constant region is as shown in SEQ ID NO:6.

[0011] Preferably, the heavy chain amino acid sequence of the bispecific antibody is as shown in SEQ ID NO:7, and the light chain amino acid sequence is as shown in SEQ ID NO:8.

[0012] Preferably, the heavy chain variable regions VH1 and VH2, and the light chain variable regions VL1 and VL2 of the bispecific antibody are respectively connected by a GS linker.

[0013] Preferably, the amino acid sequence of the GS linker is shown in SEQ ID NO:9.

[0014] Preferably, the method for preparing the bispecific antibody comprises the following steps:

[0015] (a) designing the nucleic acid sequences of the light chain and heavy chain of a bispecific antibody comprising a first antigen binding arm and a second antigen binding arm, and connecting the Fv fragment through a GS linker;

[0016] (b) inserting the nucleic acid sequence into an expression vector;

[0017] (c) transferring the expression vector into 293F cells;

[0018] (d) culturing the host cell to express the bispecific antibody;

[0019] (e) purifying the bispecific antibody.

[0020] The construction strategy of the bispecific antibody is as follows: constructing the light chain and heavy chain of the bispecific antibody comprising the first antigen binding arm and the second antigen binding arm, inserting them into the expression vector pCDNA3.1, transfecting HEK293 cells, collecting the culture supernatant, and purifying using a Protein A column.

[0021] In a second aspect, the present invention provides a nucleic acid molecule encoding the above-mentioned bispecific antibody.

[0022] In a third aspect, the present invention provides a vector comprising the above-mentioned nucleic acid molecule.

[0023] In a fourth aspect, the present invention provides a host cell, wherein the host cell comprises the above-mentioned nucleic acid molecule or the above-mentioned vector.

[0024] Preferably, the host cell is a 293F cell.

[0025] In a fifth aspect, the present invention provides a pharmaceutical composition comprising the bispecific antibody described above and a pharmaceutically acceptable carrier.

[0026] In a sixth aspect, the present invention provides the use of the above-mentioned bispecific antibodies or a combination of the bispecific antibodies in the preparation of a medicament for preventing and / or treating SARS-CoV-2 mutant strains and / or SARS-related coronavirus infections.

[0027] The bispecific antibodies provided by the present invention exhibit high binding ability to S1, and have broad-spectrum and high-efficiency activity against pseudovirus SARS-CoV-2 variants (VOCs) and SARS-related coronaviruses (SARSr-CoVs), indicating that the bispecific antibodies provided by the present invention have the potential to prevent and treat SARS-CoV-2 and SARS-related coronaviruses, and can be used to prevent and / or treat SARS-CoV-2 mutants and / or SARS-related coronavirus infections.

[0028] The present invention is beneficial in that:

[0029] The present invention provides a bispecific antibody having neutralizing activity against SARS-CoV-2 mutants and SARS-related coronaviruses, and the bispecific antibody is constructed by fusing the single-chain variable fragments of the bispecific antibody through a GS linker. The bispecific antibody exhibits high binding ability to S1, and has broad-spectrum and high-efficiency activity against pseudovirus SARS-CoV-2 variants (VOCs) and SARS-related coronaviruses (SARSr-CoVs), indicating that the bispecific antibody provided by the present invention has the potential to prevent and treat SARS-CoV-2 and SARS-related coronaviruses. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 Schematic diagram of the structural composition of the bispecific antibody 5-HI protein.

[0031] Figure 2 This is a graph showing the binding activity results of the bispecific antibody 5-HI against SARS-related coronaviruses.

[0032] Figure 3 This is a graph showing the binding activity of the bispecific antibody 5-HI against the mutant strain (VOC).

[0033] Figure 4 This is a graph showing the neutralizing activity of the bispecific antibody 5-HI against SARS-related coronaviruses.

[0034] Figure 5 This is a graph showing the neutralizing activity of the bispecific antibody 5-HI against the mutant strain (VOC).

[0035] Figure 6 This is a graph showing the neutralization activity of the bispecific antibody 5-HI against live viruses. DETAILED DESCRIPTION

[0036] In order to better illustrate the purpose, technical solutions and advantages of the present invention, the present invention will be further described below in conjunction with specific embodiments.

[0037] In the following examples, the experimental methods used are conventional methods unless otherwise specified, and the materials, reagents, etc. used are all available from commercial sources unless otherwise specified.

[0038] The amino acid sequence involved in the present invention is shown in the following table:

[0039]

[0040]

[0041]

[0042] Example 1: Construction of bispecific antibodies

[0043] DVD-Ig is an engineered bispecific antibody consisting of two variable domains, each of which can recognize and bind to different antigens; this design enables DVD-Ig to target two different antigens or epitopes at the same time, enhancing its therapeutic effect, especially in complex diseases that require simultaneous regulation of multiple targets, such as cancer, autoimmune diseases, or inflammatory diseases. In this example, a modified DVD-Ig is combined with a fragment variable region (Fv) from two different antibodies mAb1 and mAb2 (monoclonal antibodies SCM 15-45 and SCM 13-65) to construct a bispecific antibody (bsAbs).

[0044] Construction strategy: The light chain and heavy chain of a bispecific antibody comprising a first antigen-binding arm and a second antigen-binding arm were constructed and inserted into the expression vector pCDNA3.1. HEK293 cells were transfected, and the culture supernatant was collected and purified using a Protein A column.

[0045] 1. Construction of expression plasmids for the heavy and light chains of bispecific antibodies (bsAbs) (hereinafter referred to as 5-HI)

[0046] The nucleic acid sequences of the light chain and the heavy chain of the bispecific antibody comprising the first antigen binding arm and the second antigen binding arm are designed respectively.

[0047] The heavy chain amino acid sequence of the bispecific antibody is shown in SEQ ID NO: 7, and the light chain amino acid sequence is shown in SEQ ID NO: 8.

[0048] Among them, the first antigen binding arm can neutralize SARS-related coronavirus, and the second antigen binding arm can neutralize the mutant strain of SARS-CoV-2. The first antigen binding arm includes a heavy chain variable region VH1 and a light chain variable region VL1; the second antigen binding arm includes a heavy chain variable region VH2 and a light chain variable region VL2; the amino acid sequence of the VH1 is shown in SEQ ID NO: 1; the amino acid sequence of the VL1 is shown in SEQ ID NO: 2; the amino acid sequence of the VH2 is shown in SEQ ID NO: 3; the amino acid sequence of the VL2 is shown in SEQ ID NO: 4. It was synthesized and constructed by Nanjing GenScript Co., Ltd. to obtain 5-HI heavy chain plasmid and light chain plasmid respectively. Among them, VH was cloned into the AbVec2.0-hIgG1 vector (addgene, #80795), and VL lambda (VL lambda chain was synthesized and constructed by Nanjing GenScript Company, lambda gene (AFR33667.1)) was cloned into the MapAbVec1.1-IGLC (addgene, #99575) vector.

[0049] 2. Expression of bispecific antibody 5-HI in 293F eukaryotic cell expression system

[0050] (1) Use FreeStyle one day before transfection TM 293 expression medium (Gibco, catalog number: 12338018) was used to adjust the 293F cell density to 1×10 6 / mL; count the cells on the day of transfection and use fresh FreeStyle TM 293 expression medium: adjust the 293F cell density to 2 × 10 6 Pieces / mL.

[0051] (2) Prepare PEI mixed solution: Add Polyethylenimine (PEI) (Polysciences, Catalog No.: 23996-2) to OptiPRO TM SMF medium (ThermoFisher, catalog number: 12309019) was used to make the concentration of transfection 4 μg / mL; the plasmid mixture was prepared: the extracted paired heavy chain plasmid and light chain plasmid were added to OptiPRO TM Mix well in SFM medium to make the total plasmid concentration at the time of transfection to be 1 μg / mL.

[0052] (3) Finally, add the PEI mixture to the plasmid mixture and mix gently to prepare a transfection mixture system, and let it stand at room temperature for 20 minutes; add the transfection mixture system to the 293F cells in step (1), shake it gently to mix; place it in a 8% CO 2 The suspension culture was performed in a 37°C suspension incubator at 125 rpm for 7 days to obtain a suspension culture solution.

[0053] 3. Bispecific Antibody Purification

[0054] The above suspension culture solution was centrifuged at 4000 rpm for 15 min to collect the expression supernatant; the supernatant was filtered using a 0.22 μm filter (JET, catalog number: FPE204030); The protein purifier uses PBS to balance the Protein A column at a flow rate of 3 mL / min; then the filtered expression supernatant is loaded onto the Protein A column at a flow rate of 3 mL / min; PBS is used to wash away the nonspecific binding proteins on the Protein A column at a flow rate of 3 mL / min; the antibody bound to the Protein A column is eluted using a glycine buffer at pH = 3.0 at a flow rate of 1 mL / min; the eluate is collected to obtain the purified bispecific antibody 5-HI, the structure of which is as follows: Figure 1 shown.

[0055] Example 2: Detection of the binding activity of bispecific antibody 5-HI to viral protein S1

[0056] (1) Expression and purification of viral protein S1

[0057] The S1 gene with a Flag tag (located at the C-terminus) was cloned into the pVAX1 vector to obtain a plasmid encoding S1, which was then transiently expressed in 293F cells.

[0058] S1 sequences were selected from the following coronaviruses:

[0059] (a) SARS-related coronaviruses: SARS-CoV-2WT (wild type, QHD43416.1), SARS-CoV (AAP13567.1), bat-RaTG13 (QHR63300.2), pangolin-GD18 (QLR06867.1), pangolin-GX-P5L (QIA48632.1), bat-WIV1 (AGZ48828.1), civet-007 (AAU04646.1), civet-SZ3 (AY304486.1);

[0060] (b) Mutant strains VOC: Omicron mutant strains: including BA.1 (B.1.1.529, UGY75354.1), BA.2 (UHU97100.1), BA.3 (UNH00890.1), BA.5 (UYO86297.1), BQ.1 (UUN09240.1), BF.7 (UST29212.1) and BA.2.75 (UYM51877.1).

[0061] During transfection, the plasmid encoding S1 (30 μg) was mixed with PEI reagent (the mass ratio of plasmid to PEI was 1:3), incubated at room temperature for 15 min, and then transfected into 293F cells (30 mL, 2 × 10 6 The transfected cells were maintained for 5 days, and the supernatant was collected and filtered (0.22 μm), and then purified using Anti-DYKDDDDK G1 affinity resin (GenScript, L00432) to obtain purified S1 protein.

[0062] Purification process: resin wash buffer (formula: 50mM Tris-HCl, 150mM NaCl, pH = 7.4) wash three times, and with elution buffer (formula 0.1M Tris, 0.5M NaCl, pH = 12) protein is eluted into the bottle containing 1 / 20 volume of 1M HCl of eluent. The eluted protein is concentrated with PBS and Amicon tubes and stored at -80°C.

[0063] (2) ELISA detection of the half-maximal effective concentration (EC50) of the bispecific antibody 5-HI and the viral protein S1

[0064] The purified SARS-like S1 protein or SARS-CoV-2 S1 protein or SARS-CoV S1 protein was coated on an ELISA plate at a concentration of 2 μg / mL (100 μL / well); after incubation at 4°C overnight, unbound proteins were washed away with 1×PBST; the plate was blocked at room temperature for 2 h with a blocking solution containing 2% FBS (Gibco, catalog number: 10270-106) and 2% BSA (Sigma, catalog number: V900933); after the blocking solution was washed away with 1×PBST, the bispecific antibody purified in Example 1 was diluted (using a solution containing 2% FBS (Gibco, catalog number: 10270-106) and 2% The antibody solutions were diluted with blocking buffer containing BSA (Sigma, catalog number: V900933) to obtain antibody solutions with concentrations of 10, 3.33, 1.11, 0.37, 0.123, 0.041, 0.0137, 0.0046, 0.0015, 0.0005, 0.0017 and 0 μg / mL, respectively.

[0065] The above antibody solutions of different concentrations were added to the ELISA plate and incubated at 37°C for 1h; unbound antibodies were washed away with 1×PBST, and then RHP-labeled anti-human IgG antibodies (Jackson ImmunoResearch, Catalog No.: 109-035-003) were added and incubated at 37°C for 1h; unbound anti-human IgG antibodies were washed away with 1×PBST, and 100μL TMB colorimetric solution (ThermoFisher, Catalog No.: 002023) was added and incubated at room temperature for 5min; finally, 50μL 1M sulfuric acid was added to terminate the reaction; the OD value was measured using a Varioskan Flash full-wavelength scanning multifunctional reader (ThermoFisher); the data were calculated using Prism8.0 software (GraphPad) to obtain the half-effective concentration (EC50) of the bispecific antibody against the new coronavirus SARS-CoV-2 Wuhan-Hu-1 and Omicron mutants.

[0066] The results are as follows Figure 2-3 As shown, the bispecific antibody 5-HI has good binding activity against mutant strains and SARS-Related coronaviruses.

[0067] Example 3: Detection of the neutralizing activity of bispecific antibody 5-HI pseudovirus

[0068] (1) Pseudovirus Encapsulation

[0069] The S protein coding sequences of coronaviruses (CoVs), including SARS-CoV2 WT, SARS-CoV, Pangolin-GD18, Pangolin-GX-P5L, Bat-RaTG13, Bat-WIV1, Civet-007, Civet-SZ3, and Omicron (BA.1, BA.2, BA.3, BA.5, BQ.1, BF.7, BA.2.75, XBB 1.5 (YP_009724390.1), EG.5.1 (YP_009724390.1), and JN.1 (YP_009724390.1)), were cloned into the pcDNA3.1 vector.

[0070] To generate pseudovirus, when 293T cells grow to 70% confluence, co-transfect the packaging plasmid pNL4-3.Luc.RE and the plasmid expressing the S protein (the mass ratio of the packaging plasmid to the plasmid expressing the S protein is 3:1). After 6 hours, the transfection supernatant was replaced with fresh DMEM containing 10% fetal bovine serum (FBS) and placed for 48 hours. The culture supernatant was collected, centrifuged at 1500×g for 15 minutes, and filtered (0.45μm) to obtain the pseudovirus supernatant, which was stored at -80°C and subsequently diluted for titer determination and neutralization experiments.

[0071] (2) Pseudovirus neutralization experiment

[0072] One day before infection, 0.8×10 5 293T / hACE2 cells were inoculated into 96-well plates (the culture medium used was DMEM containing 10% FBS) at a density of 100 μL per well; on the day of infection, the bispecific antibody purified in Example 1 was diluted with complete culture medium (DMEM culture medium containing 10% FBS) according to a certain concentration gradient, and then an equal amount of pseudovirus supernatant was added to obtain multiple groups of mixed solutions. The antibody concentrations contained in each group of mixed solutions were 100, 33.3, 11.1, 3.7, 1.23, 0.41, 0.137, 0.046, 0.015, 0.005, 0.017 and 0 μg / mL, respectively.

[0073] The mixture of the antibody obtained above and the pseudovirus was incubated at 37°C for 1 hour. The culture medium in the 96-well plate was discarded, and the treated mixed solution was added, and centrifuged at 800×g for 30 minutes; after incubation at 37°C in a cell culture incubator for 6-8 hours, the mixed solution was discarded, and freshly prepared DMEM culture medium (Gibco, catalog number: 11995065) containing 10% FBS (Gibco, catalog number: 10270-106) was added; after the cells were cultured for 48 hours, 50 μL of cell lysis solution (Promega, catalog number: E153A) was added to each well of the 96-well culture plate, and lysed at 37°C for 2 minutes; then the 96-well culture plate was placed in a -40°C freezer for 30 minutes; after freezing, the 96-well culture plate was taken out and placed at 37°C for 3 minutes, and centrifuged at 2000 rpm for 1 minute to obtain a cell lysate; 40 μL of the above cell lysate was aspirated and added to the 96-well black flat floor; 50 μL of luciferase detection reagent (Promega, catalog number: E1501) was added, and the luciferase detection reagent was detected by Varioskan Flash full wavelength scanning multifunctional reader (ThermoFisher) was used to measure the OD value. Neutralization inhibition rate was calculated as follows: Neutralization inhibition rate = [1 - (OD value of the well with antibody and virus mixture - OD value of the blank well) / (OD value of the well with virus only without antibody - OD value of the blank well)] × 100%. Based on the results of neutralization inhibition rate, IC50 of the antibody was calculated using Prism 8.0 software (GraphPad).

[0074] The results are as follows Figure 4-5 As shown, the bispecific antibody 5-HI has good neutralizing activity against mutant strains and SARS-Related coronaviruses.

[0075] Example 4: Detection of live virus neutralization activity of bispecific antibody 5-HI against SARS-CoV-2 WT and mutant strains

[0076] 50 μL of antibody dilutions of different concentrations (antibody concentrations were 100, 33.3, 11.1, 3.7, 1.23, 0.41, 0.137, 0.046, 0.015, 0.005, 0.017, 0 μg / mL; diluted with DMEM medium (Gibco, Catalog No.: 11995065) containing 10% FBS (Gibco, Catalog No.: 10270-106)) of bispecific antibody 5-HI were mixed with 50 μL of SARS-CoV-2, XBB.5.1, EG.5.1 and JN.1 live viruses (200 focus forming units), incubated at 37°C for 1 hour, transferred to a 96-well plate seeded with Vero E6 cells, and cultured at 37°C for 1 hour for virus infection. After infection, the virus solution was discarded and DMEM virus maintenance solution containing 1.2% (v / v) sodium carboxymethyl cellulose was added. After 24 h, the cells were fixed with 4% (v / v) paraformaldehyde and 0.2% (v / v) TritonX-100 to increase cell permeability. Immunostaining was performed with SARS-CoV-2S protein rabbit monoclonal antibody (China Biological, 40143-T62) and peroxidase-labeled goat anti-rabbit antibody (H+L) (Jackson, 111-035-144), respectively. Finally, KPL TrueBle peroxidase substrate (SeraCare Life Science, 5510-0030) was used to display virus-infected lesions, and CTL Immuno Spot S6 (Ccell Technology Limited) was used to count the virus lesion units.

[0077] The results are as follows Figure 6 As shown, the bispecific antibody 5-HI exhibited good neutralizing activity against live viruses of SARS-CoV-2 WT and mutant strains.

[0078] Finally, it should be noted that the above embodiments are only used to illustrate the technical solution of the present invention rather than to limit the scope of protection of the present invention. Although the present invention has been described in detail with reference to the preferred embodiments, those skilled in the art should understand that the technical solution of the present invention can be modified or replaced by equivalents without departing from the essence and scope of the technical solution of the present invention.

Claims

1. A bispecific antibody, characterized in that: The bispecific antibody comprises a first antigen-binding arm and a second antigen-binding arm; the first antigen-binding arm comprises a heavy chain variable region VH1 and a light chain variable region VL1; the second antigen-binding arm comprises a heavy chain variable region VH2 and a light chain variable region VL2; the amino acid sequence of the VH1 is shown in SEQ ID NO: 1; the amino acid sequence of the VL1 is shown in SEQ ID NO: 2; the amino acid sequence of the VH2 is shown in SEQ ID NO: 3; the amino acid sequence of the VL2 is shown in SEQ ID NO:

4.

2. The bispecific antibody according to claim 1, characterized in that The first antigen binding arm of the bispecific antibody specifically binds to the RBD receptor binding domain of SARS-CoV-2, and the second antigen binding arm specifically binds to the shared antigenic epitope of SARS-related coronaviruses.

3. The bispecific antibody according to claim 1, characterized in that The amino acid sequence of the heavy chain constant region of the bispecific antibody is shown in SEQ ID NO: 5, and the amino acid sequence of the light chain constant region is shown in SEQ ID NO:

6.

4. The bispecific antibody according to claim 1, characterized in that The heavy chain amino acid sequence of the bispecific antibody is shown in SEQ ID NO: 7, and the light chain amino acid sequence is shown in SEQ ID NO:

8.

5. The bispecific antibody according to claim 1, wherein The heavy chain variable regions VH1 and VH2, and the light chain variable regions VL1 and VL2 of the bispecific antibody are connected via GS linkers, respectively.

6. A nucleic acid molecule encoding the bispecific antibody according to any one of claims 1 to 5.

7. A carrier, characterized in that The vector comprises the nucleic acid molecule according to claim 6.

8. A host cell, characterized in that The host cell comprises the nucleic acid molecule of claim 6 or the vector of claim 7.

9. A pharmaceutical composition, characterized in that The pharmaceutical composition contains the bispecific antibody according to any one of claims 1 to 5 and a pharmaceutically acceptable carrier.

10. Use of the bispecific antibody or a combination of bispecific antibodies according to any one of claims 1 to 5 in the preparation of a medicament for preventing and / or treating SARS-CoV-2 mutant strains and / or SARS-related coronavirus infection.

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