Three human papilloma virus antibodies and application thereof in prevention and treatment of HPV (human papilloma virus) infection

Abstract

The invention provides three human papilloma virus antibodies and application thereof in prevention and treatment of HPV infection. The antibody is derived from memory B cells of vaccinated people, a humanized cross neutralizing antibody capable of neutralizing multiple HPV types is obtained through separation, and the continuous high-concentration neutralizing antibody can neutralize virus particles continuously generated at the vagina mucosa part, prevent viruses from continuously infecting epithelial cells and help an organism to accelerate removal of HPV viruses. And a new scheme is provided for developing antibody drugs for clinical treatment of HPV persistent infection.

Description

Technical Field

[0001] The present invention belongs to the field of biology and new medicine technology, and specifically relates to three strains of human papillomavirus antibodies and their application in preventing and treating HPV infection. Background Art

[0002] Among the cancers caused by HPV virus infection, cervical cancer has attracted the most attention from researchers. HPV16 and HPV18 viruses have been identified as the most common types associated with cervical cancer, causing more than 70% of cervical cancers and cervical precancerous lesions (CIN). Cervical cancer is one of the most common cancers that threaten women's health.

[0003] Human papillomavirus is considered to be a necessary cause for the development of cervical cancer. Research data show that the vast majority of cervical cancer patients are infected with high-risk HPV. Most of the new cases of cervical cancer worldwide come from developing countries. Cervical cancer seriously affects the reproductive and health of women in my country and around the world. It takes about 3-5 years from high-risk HPV infection to cervical intraepithelial neoplasia, and about 5-10 years from cervical intraepithelial neoplasia to cervical cancer. One of the necessary conditions is persistent infection with high-risk HPV.

[0004] The HPV preventive vaccines currently under development can provide certain immune protection to the vaccinated population. Vaccination with HPV virus-like particles (VLP) vaccines can prevent HPV infection and immunize against subsequent viral attacks by inducing high and persistent titers of neutralizing antibodies. These neutralizing antibodies bind to natural HPV virus particles and neutralize the virus. However, HPV preventive vaccines also have their limitations. They can only provide protection against the HPV types possessed by the vaccine, and the cross-protection effect is not good. They cannot treat people who have already been infected with HPV viruses, and are useless for people who are infected with the virus and are persistently infected with HPV viruses. At present, no drugs have been found that can directly and effectively eliminate high-risk HPV infections, which has caused great anxiety and panic to women with persistent HPV infections, seriously affecting the psychological and physical health of the majority of women, and is a problem that needs to be urgently solved clinically.

[0005] Treating these people with persistent HPV infection is a more difficult problem to solve in clinical work, because the number of people with persistent HPV infection is larger than the number of people with cervical lesions. HPV persistent infection is manifested by the continuous synthesis of new virus particles in the epithelial cells of the vaginal mucosa after viral infection and their release locally. When the body's ability to clear local virus particles is weak, the virus particles cannot be cleared in time, leading to persistent infection of the body's virus. Therefore, how to effectively eliminate these virus particles is still a bottleneck problem that has not yet been solved. Summary of the invention

[0006] In order to solve at least one of the above problems, the present disclosure provides anti-human papillomavirus antibodies and applications thereof. Using the human papillomavirus antibodies provided by the present disclosure, drugs can be developed to provide a new solution for clinical treatment to eliminate persistent HPV infection.

[0007] According to one aspect of the present disclosure, an anti-human papillomavirus antibody or an antigen-binding fragment thereof is provided, wherein the antibody or the antigen-binding fragment thereof comprises: (1) three complementary determining regions of the following light chain variable regions: LCDR1 having the amino acid sequence of LCDR1 contained in the light chain variable region as shown in any one of SEQ ID NOs: 19 to 21, or having one or more amino acid substitutions, deletions, additions, and / or an amino acid sequence having at least 80% sequence identity with the amino acid sequence of LCDR1 contained in the light chain variable region and retaining its human papillomavirus binding activity; LCDR2 having the amino acid sequence of LCDR2 contained in the light chain variable region as shown in any one of SEQ ID NOs: 19 to 21, or having one or more amino acid substitutions, deletions, additions, and / or an amino acid sequence having at least 80% sequence identity with the amino acid sequence of LCDR2 contained in the light chain variable region and retaining its human papillomavirus binding activity; LCDR3 having the amino acid sequence of LCDR4 contained in the light chain variable region as shown in SEQ ID NOs: 19 to 21, or having one or more amino acid substitutions, deletions, additions, and / or an amino acid sequence having at least 80% sequence identity with the amino acid sequence of LCDR4 contained in the light chain variable region and retaining its human papillomavirus binding activity. NO:19-21, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of LCDR3 contained in the light chain variable region and retaining its human papillomavirus binding activity; and / or, (2) the following three complementarity determining regions (HCDRs) of heavy chain variable regions: HCDR1, which has an amino acid sequence of HCDR1 contained in a heavy chain variable region as shown in any one of SEQ ID NOs:22-24, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of HCDR1 contained in the heavy chain variable region and retaining its human papillomavirus binding activity; HCDR2, which has an amino acid sequence as shown in SEQ ID NOs:24-25. The amino acid sequence of HCDR2 contained in the heavy chain variable region as shown in any one of SEQ ID NOs: 22 to 24, or an amino acid sequence having one or several amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of HCDR2 contained in the heavy chain variable region and retaining its human papillomavirus binding activity; HCDR3, which has the amino acid sequence of HCDR3 contained in the heavy chain variable region as shown in any one of SEQ ID NOs: 22 to 24, or an amino acid sequence having one or several amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of HCDR3 contained in the heavy chain variable region and retaining its human papillomavirus binding activity.

[0008] In some embodiments, each CDR is defined by any numbering system routinely used by those skilled in the art. Exemplary numbering systems include, but are not limited to, Kabat, AbM, Chothia, Contact, IMGT, or a combination thereof.

[0009] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0010] (A1) an amino acid sequence of 3 LCDRs contained in a light chain variable region as shown in SEQ ID NO: 19, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of any one of the 3 LCDRs contained in the light chain variable region and retaining its human papillomavirus binding activity; and / or, an amino acid sequence of 3 HCDRs contained in a heavy chain variable region as shown in SEQ ID NO: 22, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of any one of the 3 HCDRs contained in the heavy chain variable region and retaining its human papillomavirus binding activity; or, (A2) an amino acid sequence as shown in SEQ ID NO:20, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of any one of the three LCDRs contained in the light chain variable region and retaining its human papillomavirus binding activity; and / or, an amino acid sequence such as the three HCDRs contained in the heavy chain variable region as shown in SEQ ID NO:23, or an amino acid sequence such as the amino acid sequence of any one of the three HCDRs contained in the heavy chain variable region having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of any one of the three HCDRs contained in the heavy chain variable region and retaining its human papillomavirus binding activity; or, (A3) an amino acid sequence such as SEQ ID The three LCDRs contained in the light chain variable region as shown in NO:21, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of any one of the three LCDRs contained in the light chain variable region and retaining its human papillomavirus binding activity; and / or, the three HCDRs contained in the heavy chain variable region as shown in SEQ ID NO:24, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of any one of the three HCDRs contained in the heavy chain variable region and retaining its human papillomavirus binding activity.

[0011] In some embodiments, the antibody or antigen-binding fragment thereof comprises: LCDR1, which has an amino acid sequence as shown in any one of SEQ ID NOs: 1 to 3, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity therewith and retaining its human papillomavirus binding activity; and / or LCDR2, which has an amino acid sequence as shown in any one of SEQ ID NOs: 4 to 6, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity therewith and retaining its human papillomavirus binding activity; and / or LCDR3, which has an amino acid sequence as shown in any one of SEQ ID NOs: 7 to 9, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity therewith and retaining its human papillomavirus binding activity; and / or HCDR1, which has an amino acid sequence as shown in SEQ ID NOs: 8 to 10, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity therewith and retaining its human papillomavirus binding activity. NO: 10-12, or an amino acid sequence having one or several amino acid substitutions, deletions, additions thereto, and / or an amino acid sequence having at least 80% sequence identity thereto and retaining its human papillomavirus binding activity; and / or HCDR2, which has an amino acid sequence as shown in any one of SEQ ID NO: 13-15, or an amino acid sequence having one or several amino acid substitutions, deletions, additions thereto, and / or an amino acid sequence having at least 80% sequence identity thereto and retaining its human papillomavirus binding activity; and / or HCDR3, which has an amino acid sequence as shown in any one of SEQ ID NO: 16-18, or an amino acid sequence having one or several amino acid substitutions, deletions, additions thereto, and / or an amino acid sequence having at least 80% sequence identity thereto and retaining its human papillomavirus binding activity.

[0012] In some embodiments, the antibody or antigen-binding fragment thereof comprises: (B1) an amino acid sequence such as LCDR1, LCDR2 and LCDR3 as shown in SEQ ID NO:1, SEQ ID NO:4 and SEQ ID NO:7, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of any one of LCDR1, LCDR2 and LCDR3 and retaining its human papillomavirus binding activity; and / or an amino acid sequence such as HCDR1, HCDR2 and HCDR3 as shown in SEQ ID NO:10, SEQ ID NO:13 and SEQ ID NO:16, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of any one of HCDR1, HCDR2 and HCDR3 and retaining its human papillomavirus binding activity; or, (B2) an amino acid sequence such as SEQ ID NO:2, SEQ ID NO:5 and SEQ ID NO:6. NO:8, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of any one of said LCDR1, LCDR2 and LCDR3 and retaining its human papillomavirus binding activity; and / or, an amino acid sequence such as HCDR1, HCDR2 and HCDR3 as shown in SEQ ID NO:11, SEQ ID NO:14 and SEQ ID NO:17, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of any one of said HCDR1, HCDR2 and HCDR3 and retaining its human papillomavirus binding activity; or, (B3) an amino acid sequence such as SEQ ID NO:3, SEQ ID NO:6 and SEQ ID NO:7. The LCDR1, LCDR2 and LCDR3 shown in NO:9, or an amino acid sequence having one or several amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of any one of the LCDR1, LCDR2 and LCDR3 and retaining its human papillomavirus binding activity, and / or, an amino acid sequence such as HCDR1, HCDR2 and HCDR3 shown in SEQ ID NO:12, SEQ ID NO:15 and SEQ ID NO:18, or an amino acid sequence having one or several amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of any one of the HCDR1, HCDR2 and HCDR3 and retaining its human papillomavirus binding activity.

[0013] In some embodiments, the antibody or its antigen-binding fragment comprises: the antibody or its antigen-binding fragment comprises: a light chain variable region, comprising an amino acid sequence as shown in any one of SEQ ID NOs: 19 to 21, or an amino acid sequence having one or several amino acids substituted, deleted or added thereto, and / or an amino acid sequence having at least 80% sequence identity thereto and retaining its human papillomavirus binding activity; and / or a heavy chain variable region, comprising an amino acid sequence as shown in any one of SEQ ID NOs: 22 to 24, or an amino acid sequence having one or several amino acids substituted, deleted or added thereto, and / or an amino acid sequence having at least 80% sequence identity thereto and retaining its human papillomavirus binding activity.

[0014] In some embodiments, the antibody or antigen-binding fragment thereof comprises:

[0015] (C1) a light chain variable region comprising the amino acid sequence as shown in SEQ ID NO: 19, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity thereto and retaining its human papillomavirus binding activity; and / or a heavy chain variable region comprising the amino acid sequence as shown in SEQ ID NO: 22, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity thereto and retaining its human papillomavirus binding activity; or (C2) a light chain variable region comprising the amino acid sequence as shown in SEQ ID NO: 20, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity thereto and retaining its human papillomavirus binding activity; and / or a heavy chain variable region comprising the amino acid sequence as shown in SEQ ID NO: 21. NO:23, or an amino acid sequence having one or several amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity thereto and retaining its human papillomavirus binding activity; or (C3) a light chain variable region comprising the amino acid sequence shown in SEQ ID NO:21, or an amino acid sequence having one or several amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity thereto and retaining its human papillomavirus binding activity; and / or a heavy chain variable region comprising the amino acid sequence shown in SEQ ID NO:24, or an amino acid sequence having one or several amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity thereto and retaining its human papillomavirus binding activity.

[0016] In some embodiments, the antibody or antigen-binding fragment may be of, but is not limited to, IgA, IgD, IgE, IgG or IgM type.

[0017] In some embodiments, the antibody or antigen-binding fragment includes, but is not limited to, scFv, Fab, Fab', (Fab')2, Fv fragment, dsFv, bispecific antibody, and multispecific antibody;

[0018] In some embodiments, the antibody or antigen-binding fragment is used to neutralize human papillomavirus (HPV).

[0019] In some embodiments, the human papillomavirus includes HPV6, HPV11, HPV16, HPV18, HPV31, HPV33, HPV45, HPV52 or HPV58.

[0020] In some embodiments, the antibody or antigen-binding fragment includes, but is not limited to, a murine antibody, a chimeric antibody, a humanized antibody, or a fully human antibody.

[0021] In the present disclosure, the antigen-binding fragment generally includes at least a portion of the antigen-binding region or variable region of an antibody, such as one or more CDRs. Antibody fragments retain at least some of the binding specificity of the antibody.

[0022] According to another aspect of the present disclosure, a nucleic acid molecule is provided, which encodes the antibody or antigen-binding fragment thereof.

[0023] According to another aspect of the present disclosure, an expression vector is provided, which comprises the nucleic acid molecule and / or the expression vector.

[0024] In some embodiments, the expression vector includes a prokaryotic expression vector and a eukaryotic expression vector.

[0025] In some embodiments, the eukaryotic expression vector includes a yeast expression vector, a mammalian expression vector, an insect expression vector, and the like.

[0026] According to yet another aspect of the present disclosure, a host cell is provided, comprising the nucleic acid molecule and / or the expression vector.

[0027] In some embodiments, the host cell is selected from a prokaryotic cell and a eukaryotic cell.

[0028] In some embodiments, the prokaryotic cell comprises a bacterial cell.

[0029] In some embodiments, the bacterial cells include Escherichia coli and Streptomyces;

[0030] In some embodiments, the eukaryotic cells include yeast cells, mammalian cells, insect cells, etc.

[0031] In some embodiments, the mammal is selected from humans, monkeys, mice, rats, hamsters, goats, sheep, cows, pigs, dogs, or cats.

[0032] In some embodiments, the mammalian cells include CHO cells, 293 cells, Vero cells, BHK cells, NS0 cells, SP2 / 0 cells, YO myeloma cells, P3X63 mouse myeloma cells, PER cells, PER.C6 cells or hybridoma cells.

[0033] According to another aspect of the present disclosure, a chimeric antigen receptor is provided, which includes an extracellular antigen binding domain, a transmembrane domain and an intracellular signaling domain, wherein the extracellular antigen binding domain comprises the antibody or antigen-binding fragment thereof.

[0034] According to yet another aspect of the present disclosure, a modified immune cell is provided, comprising the chimeric antigen receptor.

[0035] According to another aspect of the present disclosure, an antibody-drug conjugate is provided, comprising: the antibody or antigen-binding fragment thereof; and a drug covalently linked to the antibody or antigen-binding fragment thereof.

[0036] According to yet another aspect of the present disclosure, a detection kit is provided, the kit comprising the antibody or the antigen-binding fragment thereof.

[0037] According to yet another aspect of the present disclosure, a composition is provided, comprising the antibody or antigen-binding fragment thereof, and a pharmaceutically acceptable carrier.

[0038] In some embodiments, the antibody or antigen-binding fragment thereof comprises one or more of: (A1), (A2), and (A3).

[0039] In some embodiments, the antibody or antigen-binding fragment thereof comprises: (A1), and (A2), and (A3).

[0040] According to another aspect of the present disclosure, provided is the use of the antibody or antigen-binding fragment thereof, the nucleic acid molecule, the expression vector, the host cell, the chimeric antigen receptor, the modified immune cell, the antibody-drug conjugate, the detection kit and / or the composition in drug research, drug preparation, non-clinical research, clinical research and clinical testing.

[0041] According to another aspect of the present disclosure, provided is the use of the antibody or antigen-binding fragment thereof, the nucleic acid molecule, the expression vector, the host cell, the chimeric antigen receptor, the modified immune cell, the antibody-drug conjugate, the detection kit and / or the composition in the preparation of a product for diagnosing, preventing and / or treating human papillomavirus infection.

[0042] According to another aspect of the present disclosure, a method for detecting human papillomavirus is provided, the method comprising the step of using the antibody or antigen-binding fragment thereof for detection.

[0043] According to another aspect of the present disclosure, a method for preventing or treating human papillomavirus infection is provided, comprising administering a therapeutically effective amount of the antibody or antigen-binding fragment thereof or the composition to a subject.

[0044] Beneficial effects:

[0045] The present invention discloses the isolation of high-neutralizing anti-human papillomavirus antibodies from the serum of volunteers vaccinated with HPV vaccines. These three antibodies are derived from the memory B cells of the vaccinated population, and humanized cross-neutralizing antibodies that can neutralize multiple HPV types are isolated, which can cover a variety of major HPV types. The sustained high concentration of neutralizing antibodies can neutralize the virus particles continuously produced in the vaginal mucosa, prevent the virus from continuously infecting epithelial cells, and help the body accelerate the clearance of HPV viruses. It provides a new solution for the clinical treatment of persistent HPV infection and the development of broad-spectrum or specific HPV-type antibody drugs. BRIEF DESCRIPTION OF THE DRAWINGS

[0046] Figure 1 The technical routes for antibody screening, preparation and detection described in Examples 1 to 4 are shown.

[0047] Figure 2 Orthogonal views of the H419 Fab fragment and L1 pentamer complex are shown, wherein blue and orange represent the light and heavy chains of the antibody, and the remaining five colors represent the pentamer of the HPV L1 protein.

[0048] Figure 3 Orthogonal views of the H421 Fab fragment and L1 pentamer complex are shown, wherein blue and orange represent the light and heavy chains of the antibody, and the remaining five colors represent the pentamer of the HPV L1 protein.

[0049] Figure 4 Orthogonal views of the H7 Fab fragment and L1 pentamer complex are shown, wherein blue and orange represent the light and heavy chains of the antibody, and the remaining five colors represent the pentamer of the HPV L1 protein. DETAILED DESCRIPTION

[0050] The cervix is ​​located between the vagina and the uterus and is characterized by a single layer of columnar secretory epithelium. HPV viruses infect basal epithelial cells through epithelial abrasion or wounds, and can also enter epithelial cells through cells at the junction of single-layer squamous cells in the cervix and outside the cervix. HPV viruses infect epithelial cells and induce cervical dysplasia and cervical precancerous lesions (CIN), which then develop into cervical cancer due to persistent infection with high-risk HPV. After viral infection, early HPV genes (E1, E2, E4, E5, E6 and E7) are expressed in the production cycle. In the upper layer of epithelial tissue, the viral genome is replicated, late HPV genes (L1 and L2) and E4 are expressed, and progeny HPV virus particles are assembled. The shed virus particles repeat the life cycle of the virus and continue to infect the remaining epithelial tissue cells. High-risk HPV infection that persists for more than 1 year increases the risk of developing high-grade squamous intraepithelial lesions or potentially invasive cancer. At present, no clear and effective drug has been found to eliminate high-risk HPV infection directly and effectively, and persistent infection with high-risk HPV causes great anxiety and panic to the vast majority of women.

[0051] According to the process of HPV virus invading epithelial cells, sustained high concentrations of neutralizing antibodies can neutralize the virus particles continuously produced in the vaginal mucosa, prevent the virus from continuously infecting epithelial cells, and help the body accelerate the elimination of HPV virus. Studies have shown that the initial binding of human papillomavirus capsid to host cells is mainly through the interaction between the viral capsid protein L1 and heparin sulfate proteoglycan (HSPG) on the cell surface.

[0052] Therefore, the present invention separates high-neutralizing anti-HPV L1 protein antibodies from the serum of volunteers vaccinated with HPV vaccines. These three antibodies (H419, H7, H421) are derived from memory B cells of the vaccinated population without any sequence optimization, and their sequences are shown in Table 1. And the binding ability and neutralization level of these antibodies to nine types of HPV were detected. The antibody drugs prepared from these neutralizing antibodies can act directly on the vaginal mucosa, which can neutralize the virus particles shed from the infected vaginal area, prevent the newly generated HPV virus from continuously infecting the vaginal epithelial cells, and assist the body in clearing the HPV virus, providing a new solution for clinical treatment to clear persistent HPV infection.

[0053] Table 1. Amino acid sequences of the CDR regions of the three antibodies

[0054]

[0055]

[0056] Note: The CDR regions described in Table 1 are defined by IMGT rules.

[0057] definition

[0058] Unless otherwise defined, all technical and scientific terms used in the present invention have the same meaning as commonly used in the field to which the present invention belongs. For the purpose of interpreting this specification, the following definitions will apply, and where appropriate, terms used in the singular will also include the plural form, and vice versa.

[0059] As used herein, the articles "a," "an," and "an" include plural referents unless the context clearly dictates otherwise.

[0060] The expression "about" as used herein is understood by one of ordinary skill in the art and varies within a certain range depending on the context in which it is used. If one of ordinary skill in the art is not aware of the use of the term based on the context in which it is used, "about" will mean up to plus or minus 10% of the specified value.

[0061] The term "specific binding" as used herein refers to having binding selectivity for an antigen and can be distinguished from unwanted or non-specific binding. The ability of an antigen binding molecule to bind to a specific antigen can be measured by techniques familiar to those skilled in the art. For example, surface plasmon resonance (SPR) technology, enzyme-linked immunosorbent assay (ELISA) or traditional binding assays are measured.

[0062] The term "antibody" as used herein encompasses various antibody structures, including but not limited to monoclonal antibodies, polyclonal antibodies, monospecific and multispecific antibodies (e.g., bispecific antibodies or trispecific antibodies), single-chain molecules and antibody fragments, as long as the desired antigen binding activity is exhibited. Each heavy chain may include a heavy chain variable region (VH) and a heavy chain constant region (CH). Each light chain may include a light chain variable region (VL) and a light chain constant region (CL).

[0063] The term "antigen-binding fragment" as used herein means an antibody fragment that retains the ability to specifically bind to an antigen bound by a full-length antibody, such as a fragment that retains one or more CDR regions. Examples of antigen-binding fragments include, but are not limited to, Fab, Fab', F(ab')2, and Fv fragments; diabodies; linear antibodies; single-chain antibody molecules, such as single-chain Fv (ScFv); nanobodies, etc.

[0064] The term "Fab fragment" used herein refers to the region in the antibody structure that can bind to the antigen. The Fab fragment consists of a complete light chain and a partial heavy chain structure, and the light chain and the heavy chain are connected by a disulfide bond and are relatively small in size. The Fab fragment can be obtained by protease digestion of the full-length antibody. For example, under the action of papain, human immunoglobulin G (IgG) can be degraded into two Fab fragments and an Fc fragment; or under the action of pepsin, IgG can be degraded into a F(ab')2 fragment and a pFc' fragment. The F(ab')2 fragment can be further reduced to form two Fab' fragments. Fab fragments can also be expressed and prepared by prokaryotic systems (such as E. coli systems) and mammalian cell systems. The E. coli expression system has the characteristics of low production cost and fast production speed, but it is easy to form inclusion bodies, and the later purification and renaturation are more troublesome, and the activity of the protein obtained by renaturation is very low or even inactive. The expression of Fab fragments in mammalian cells can smoothly form disulfide bonds, which is closer to the natural Fab fragment structure and has higher activity.

[0065] The term "complementarity determining region (CDR)" refers to the site in the variable region of an antibody that confers binding specificity to an antigen. It will be understood by those skilled in the art that, unless otherwise specified, "CDR" and "complementarity determining region" of a given antibody or its region (e.g., variable region) should be understood to encompass "CDR" and "complementarity determining region" as defined by any one known in the art. It is well known to those skilled in the art that the CDR of an antibody can be defined in the art by a variety of methods, such as the Kabat definition rules based on sequence variability, the Chothia definition rules based on the position of the structural loop region, the Martin definition rules based on the sequence and framework region, the IMGT definition rules based on the amino acid sequence alignment of the germline V gene, the reference tool for antibody humanization design based on CDR transplantation, etc. It will be understood by those skilled in the art that, unless otherwise specified, the term "CDR" and "complementarity determining region" of a given antibody or its region (e.g., variable region) should be understood to encompass CDR defined by any one of the above-mentioned known schemes described in the present invention, and the amino acid sequences corresponding to the defined CDR definition rules should also fall within the scope of protection of the present invention.

[0066] The term "chimeric antibody" as used herein refers to an antibody that combines antibody fragments from different species. Specifically, for example, a monoclonal antibody from one species (e.g., mouse) has its Fc constant region replaced with an Fc constant region from a species (e.g., human) via DNA recombination technology.

[0067] The term "humanized antibody" as used herein refers to an antibody comprising a human immunoglobulin framework region and one or more CDRs from a non-human (e.g., mouse, rat, rabbit or synthetic) immunoglobulin. Except for the CDRs, all other parts in the humanized antibody are substantially identical to the corresponding parts of natural human immunoglobulin sequences.

[0068] The term "fully human antibody" as used herein is intended to include antibodies with variable regions and constant regions derived from human germline immunoglobulin sequences. As used herein, the term "fully human antibody" refers to almost every part of its protein molecule (e.g., CDR, FR, CL, HC domains (e.g., CH1, CH2, CH3), hinges, VL, VH) that is almost non-immunogenic in the human body, and has only small sequence changes or variations relative to human natural immunoglobulins. Therefore, fully human antibodies are different from chimeric or humanized antibodies. It should be noted that fully human antibodies can be produced by human cells, non-human animals, or prokaryotic or eukaryotic cells that can express functionally rearranged human immunoglobulin (e.g., heavy chain and / or light chain) genes.

[0069] The term "composition" as used in the present disclosure refers to any composition comprising at least one biologically active agent, for example, including the fusion protein. When used herein, the term "composition" also refers to a composition comprising an active pharmaceutical ingredient to be delivered to a subject, for example, to achieve a therapeutic, preventive, diagnostic, preventive or prognostic effect. The composition of the present invention can be administered as a food or health product, such as a nutritional supplement. Generally, the composition of the present invention is used to treat humans, but it can be used to treat animals, including monogastric mammals, such as poultry, pigs, cats, dogs, horses or rabbits. The composition of the present invention can be used to enhance the growth and performance of animals. If administered to an animal, oral gavage can be used.

[0070] The term "pharmaceutically acceptable carrier" used in the present disclosure refers to a substance that is suitable for use in humans and / or animals without excessive adverse side effects (such as toxicity, irritation, and allergic reactions), that is, a substance with a reasonable benefit / risk ratio. Examples of pharmaceutically acceptable carriers include cellulose and its derivatives (such as methylcellulose, ethylcellulose, hydroxypropylmethylcellulose, sodium carboxymethylcellulose, etc.), gelatin, talc, solid lubricants (such as stearic acid, magnesium stearate), calcium sulfate, vegetable oils (such as soybean oil, sesame oil, peanut oil, olive oil, etc.), polyols (such as propylene glycol, glycerol, mannitol, sorbitol, etc.), emulsifiers (such as Tween), wetting agents (such as sodium lauryl sulfate), buffers, chelating agents, thickeners, pH regulators, transdermal enhancers, colorants, flavoring agents, stabilizers, antioxidants, preservatives, antibacterial agents, pyrogen-free water, etc.

[0071] As used herein, the term "nucleic acid molecule" refers to any one or more nucleic acid segments, such as DNA or RNA fragments, present in a polynucleotide.

[0072] As used herein, "percent homology", "% homology" or "% sequence identity" is used in the context of the present invention to describe the degree of similarity between two nucleotide sequences or two amino acid sequences, and has the same meaning as "percent identity". The percent homology of two sequences can be calculated as follows, after the two sequences are aligned, by dividing the number of positions where the residues are identical by the total length of the aligned sequences and multiplying by 100%. Methods and tools for aligning two amino acid sequences or nucleotide sequences are well known in the art, such as the BLAST suite provided on the NCBI website. As used herein, having "at least 85% sequence identity" to a sequence refers to having at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% sequence identity to the sequence.

[0073] The term "expression vector" as used herein refers to a DNA molecule that introduces a specific gene operably linked thereto into a target cell and directs expression. The expression vector includes vectors that are self-replicating nucleic acid structures and vectors that are incorporated into the genome of the host cell into which it has been introduced. The expression vector can perform transcription of a large amount of stable mRNA. Once the expression vector is in the target cell, the cellular transcription and / or translation machinery generates a ribonucleic acid molecule or protein encoded by the gene.

[0074] The term "host cell" as used herein is used interchangeably and refers to a cell into which an exogenous nucleic acid has been introduced, and also includes the offspring of such cells. Host cells include primary transformed cells and offspring derived therefrom. The nucleic acid of the offspring is completely identical to the parent cell, or not completely identical, and may contain mutations. Host cells include cultured cells, for example cultured mammalian cells, such as CHO cells, 293 cells, Vero cells, BHK cells, NS0 cells, SP2 / 0 cells, YO myeloma cells, P3X63 mouse myeloma cells, PER cells, PER.C6 cells or hybridoma cells, yeast cells, insect cells and plant cells. Host cells of the present disclosure also include cells contained in transgenic animals, transgenic plants or cultured plants or animal tissues.

[0075] The composition can be in any dosage form, including, but not limited to, tablets, pills, powders, granules, capsules, suspensions, solutions, emulsions, syrups, injections, sprays, aerosols, powder sprays, lotions, liniments, ointments, plasters, pastes, patches, eye drops, nasal drops, suppositories, etc.

[0076] The compositions of the present disclosure can be administered in a pharmaceutically effective amount. "Pharmaceutically effective amount" means that the amount used is sufficient to treat the disease, with a reasonable benefit / risk ratio applicable to any medical treatment. The effective dosage level of the composition can be determined based on the type of subject, the severity of the disease, the age and gender of the subject, the activity of the drug, the sensitivity to the drug, the time of administration, the route of administration, the excretion rate, the duration of treatment, the drug used in combination with the composition, and other known factors in the medical field. The compositions of the present disclosure can be used alone or in combination with other therapeutic agents, and can be administered sequentially or simultaneously with conventional therapeutic agents. The composition can be administered in one or more dosage forms.

[0077] As used herein, the term "treating" a disease in a subject or "treating" a subject having or suspected of having a disease refers to administering a drug therapy, such as administering one or more pharmaceutical agents, to a subject, thereby reducing or preventing worsening of at least one symptom of the disease. Thus, in some embodiments, "treating" refers, inter alia, to delaying progression, accelerating remission, inducing remission, increasing remission, accelerating recovery, increasing the efficacy of alternative therapies, or reducing resistance to alternative therapies, or a combination thereof.

[0078] The term "diagnosis" as used herein refers not only to "judgment", but also includes "determination", "differentiation", "identification", "inspection", "detection" and "prediction", etc. In some embodiments, the diagnosis includes distinguishing or assisting in distinguishing lesions, judging the degree of disease development, or predicting the risk of disease, treatment effect, prognosis and recurrence risk.

[0079] As used herein, the terms "preventing" and "preventing" are art-recognized and well-known in the art when used in connection with a condition such as a local recurrence, and include administration of a composition that reduces the occurrence or delays the onset of symptoms of a medical condition in a subject relative to a subject not receiving the composition.

[0080] The term "EC50" as used herein refers to the concentration for 50% of maximal effect, that is, the antibody concentration that can induce 50% of the maximal effect (in this article: the ability of the antibody to bind to the HPV VLP protein).

[0081] The term "IC50" as used herein refers to the half-maximal inhibitory concentration, i.e., the antibody concentration that can cause 50% of the maximum inhibitory effect (in this article: the ability to inhibit cell activity). The following examples and drawings are provided to help understand the present invention. However, it should be understood that these examples and drawings are only used to illustrate the present invention and do not constitute any limitation. The actual protection scope of the present invention is set forth in the claims. It should be pointed out that for those skilled in the art, several variations and improvements made on the basis of this invention concept all belong to the protection scope of the present invention. In the following description, the description of well-known structures and technologies is omitted to avoid unnecessary confusion of the concepts disclosed herein. Such structures and technologies are also described in many publications. In addition, the reagents used in the examples without indicating the manufacturer are all conventional products that can be obtained commercially.

[0082] Example

[0083] The technical routes for antibody screening, preparation and detection described in Examples 1 to 4 are shown in Figure 1 .

[0084] Embodiment 1:

[0085] 1) Blood sample collection from volunteers

[0086] 100 mL of blood was collected from volunteers one month after receiving HPV vaccine. The age range of volunteers was between 18 and 38 years old, and they had no history of vaccine allergy. The experimental plan and ethical review were approved by the Ethics Review Committee of Peking Union Medical College Hospital, and all participants signed written informed consent.

[0087] 2) PBMC cell isolation

[0088] The blood of volunteers was subjected to density gradient centrifugation in a 50 mL centrifuge tube with Ficoll-Hypaque (Cytiva) as a lymphocyte separation medium at the bottom. After multiple washes with Hanks buffer (Solarbio), the isolated PBMC cells were placed in 90% heat-inactivated FBS supplemented with 10% dimethyl sulfoxide (DMSO) and stored in liquid nitrogen until thawed for experiments.

[0089] 3) Flow cytometry to isolate mononuclear B cells

[0090] The frozen PBMCs were quickly thawed in a 37°C water bath, then added to FACS buffer (PBS with 2% fetal bovine serum (PAN Biotech)), centrifuged at 300g for 8 min, the supernatant was discarded, and the cells were resuspended in FACS buffer. The cell suspension was filtered through a 40μm cell sieve and counted. Fluorescence staining was performed, and about 1×106 cells into a 1.5mL Ep tube; the remaining cells were used as sample tubes for fluorescent staining according to the purpose of the experiment. The staining solution included Percp-cy5.5anti-human CD3 (BioLegend), APC anti-human CD27 (BioLegend), Pacific Blue anti-human CD19 (BioLegend), PE anti-human IgG (BioLegend), PerCP / Cy5.5 anti-human IgM (BioLegend) and AF488-labeled HPV VLP proteins of various types. Block at 4℃ in the dark for 30min. Then wash the cells twice with 500μL FACS buffer and place them on an ice box for sorting. Using a BD FACSAria III flow cytometer (BD Biosciences), single memory B cells bound by AF488-labeled proteins were sorted into a 96-well PCR plate, and then antibody sequencing and cloning expression of B cell monoclonal clones were performed.

[0091] 4) Antibody cloning, expression and purification

[0092] The nucleic acid sequences encoding the variable regions of the naturally paired heavy and light chains of antibodies were artificially synthesized, and then subcloned into the pcDNA3.1 vector and expressed in HEK293F cells. The recombinant plasmids encoding each antibody heavy chain and light chain were transiently co-transfected into HEK293F cells and cultured at 37°C for 7 days under 8% CO2 to achieve antibody expression. After 7 days of expression, the cell suspension was collected, and then centrifuged at 3400rpm and 4°C for 40min to collect the cell culture supernatant. The supernatant was filtered using a 0.45μM filter cup, and concentrated and filtered using a membrane package with a cutoff value of 100kDa, and the solution was replaced with PBS more than 100 times. The complete antibody containing the Fc fragment was hung on the medium using a protein A affinity chromatography gravity column (Tian Di Renhe Company), and then the antibody was eluted with IgG Elution Buffer (Pierce), and the complete antibody protein was obtained after concentration and replacement with PBS.

[0093] Example 2: Elisa binding experiment of humanized antibody and HPV type 9 VLP protein

[0094] HPV VLP protein (GenBank of HPV6 is AAC80442.1; GenBank of HPV11 is AAA46935.1; GenBank of HPV16 is ANA05496; GenBank of HPV18 is AAQ92369; GenBank of HPV31 is P17388; GenBank of HPV33 is AMY16565; GenBank of HPV45 is P36741; GenBank of HPV52 is AML80965; GenBank of HPV58 is AFS33402) was diluted to 2 μg / mL with PBS for antigen coating, and 100 μL of the coating solution was added to each well of the ELISA plate and incubated at 4°C overnight. The coated plate was washed three times with PBST (PBS + 0.05% Tween20), adding 300 μL each time. Then add 300 μL of 2% BSA to each well for blocking and place at room temperature for 2 hours. Wash the blocked plate 3 times with PBST, dilute the antibody with an original concentration of 1 mg / mL with PBS (dilute 1000 times in the first well) in 8 gradients in a 1:3 sequence, add 100 μL to the corresponding wells of the ELISA plate, and incubate at 37°C for 30 minutes. Then add 100 μL of secondary antibody (horseradish peroxidase HRP-labeled anti-human IgG (Sigma-Aldrich)) diluted 1:5000 to each well, incubate at 37°C for 30 minutes, and wash the plate 5 times with PBST. Add 100 μL of TMB colorimetric solution to each well and incubate at room temperature for 15 minutes in the dark. Then, 50 μL of 2M H2SO4 solution was added to terminate the reaction, and the plate was read by an ELISA instrument. The absorbance value was read at an OD value of 450 nm, and candidate antibodies H419, H7, and H421 with high binding ability were screened out (the amino acid sequences of their heavy chain variable regions, light chain variable regions, and CDRs are shown in Table 1, and the DNA sequences of their light chain variable regions and heavy chain variable regions are shown in Table 2). The antibody concentration when the absorbance value reaches 50% of the maximum value is recorded as the EC value of the antibody. 50 Table 3 shows the EC values ​​of these three antibodies. 50 It can be seen that the screened HPV antibodies have good cross-binding ability for HPV18 / 45, HPV16 / 31 / 52 and HPV6 / 11 / 33.

[0095] Table 2. DNA sequences of light chain variable regions and heavy chain variable regions of three antibodies

[0096]

[0097]

[0098] Table 3. Binding ability of HPV antibodies to various types of VLP proteins

[0099]

[0100]

[0101] Example 3: Pseudovirus neutralization assay to detect the neutralizing ability of antibodies

[0102] The antibody concentration was adjusted to 1 mg / mL, and 250 μL of sterile water was added to the wells around the 96-well plate to seal the edges to reduce experimental errors. The antibody was diluted fourfold in seven gradients starting from 30 times. An equal volume of HPV pseudovirus (provided by the China Food and Drug Inspection Institute) was added to the antibody serial dilution solution in the 96-well plate and incubated at 37°C for 1 hour. After the incubation, 100 μL of digested HEK 293T cells was added to each well. Place in a 37°C, 5% CO2 cell culture incubator and culture for 72 hours. The Cytation5 enzyme plate reader was used for detection, and the inhibition rate (%) was calculated as 1-(sample detection value-cell control value) / (virus control value-cell control value)×100(%). The antibody concentration corresponding to the inhibition rate of 50% is IC 50 The IC of candidate antibodies was tested by in vitro pseudovirus neutralization assay. 50 As shown in Table 4, it can be seen that these HPV antibodies have good cross-neutralization activity against HPV18 / 45, HPV16 / 31 / 52 / 58 and HPV6 / 11 / 33.

[0103] Table 4 Neutralizing activity of HPV antibodies against various pseudoviruses

[0104]

[0105] At the same time, in order to make the antibody drug have obvious neutralization against nine types of HPV, we mixed the three antibodies to test the neutralizing antibody titer against HPV6, 11, 16, 18, 31, 33, 45, 52, and 58 pseudoviruses. As shown in Table 5, the mixed antibody has a good neutralizing effect on nine types of HPV pseudoviruses.

[0106] Table 5 Neutralization levels of mixed HPV cross-neutralizing antibodies in a group of three

[0107]

[0108] Example 4: Cryo-EM sample preparation and data collection

[0109] Because the HPVVLP protein morphology under electron microscopy is not uniform and is easily deformed, which is not conducive to subsequent structural analysis, we used the reducing agent dithiothreitol (DTT) to disaggregate the VLP protein into a uniform L1 pentamer protein. The specific operation is to repeatedly concentrate the HPVVLP protein in a centrifugal ultrafiltration tube with a disaggregation buffer containing 1mM DTT and change the buffer until the dilution ratio reaches 1:1000.

[0110] 0.8 mg / mL HPV L1 pentamer protein was mixed with purified HPV antibody Fab fragments at a molar ratio of 1:1.2 and incubated on ice for 30 min for sample preparation. We selected a 300-mesh gold grid covered with a discontinuous carbon film (C-flat, CF-1.2 / 1.3-3Au-50, USA). First, the grid was glow-discharged and placed in a Solarus plasma cleaner (Gatan, USA). The mode was selected as a mixed gas of H2 and O2, and the discharge power was set to 25 W and the discharge time was set to 25 s to complete the hydrophilic treatment of the grid. The parameters of the frozen sample preparation instrument Vitrobot were adjusted, the temperature was set to 22 ° C, the ambient humidity in the chamber was 100%, the filter paper clamping time was set to 6 s, and the clamping force was set to 0. After that, the treated gold grid was clamped with special tweezers for sample preparation, and 3 μL of the premixed sample was adsorbed on the grid. Lightly step on the foot control panel to start the preset program. After the filter paper is clamped on the grid, it is quickly put into the prepared liquid ethane to prepare the sample.

[0111] Afterwards, FEI's Titan Krios 300kV cryogenic transmission electron microscope was used to collect cryo-EM sample data, and Relion 3.0 and cryoSPARC software were used for data analysis. Figure 2 to Figure 4 As shown. The structural study of the antibody Fab fragment and the capsid protein L1 that constitutes the surface of the HPV virus particle shows the structural basis of the binding ability and high neutralization ability between the two. Most of the cross-neutralizing antibodies we selected bind to the relatively conserved amino acid sites at the top of the corresponding HPV type L1 pentamer. The antibody complex structure further verified the previous experimental results and provided structural support for the use of HPV neutralizing antibody drugs to neutralize virus particles shed from infected vaginal areas.

[0112] The technical solution of the present invention is not limited to the above-mentioned specific embodiments. All technical variations made according to the technical solution of the present invention fall within the protection scope of the present invention.

Claims

1. An anti-human papillomavirus antibody or an antigen-binding fragment thereof, the antibody or the antigen-binding fragment thereof comprising: (1) The complementarity determining regions of the following three light chain variable regions: LCDR1, which has the amino acid sequence of LCDR1 contained in the light chain variable region as shown in any one of SEQ ID NOs: 19 to 21, or has one or more amino acid substitutions, deletions, additions, and / or an amino acid sequence having at least 80% sequence identity therewith and retaining its human papillomavirus binding activity compared to the amino acid sequence of LCDR1 contained in the light chain variable region; LCDR2, which has the amino acid sequence of LCDR2 contained in the light chain variable region as shown in any one of SEQ ID NOs: 19 to 21, or has one or more amino acid substitutions, deletions, additions, and / or an amino acid sequence having at least 80% sequence identity therewith and retaining its human papillomavirus binding activity compared to the amino acid sequence of LCDR2 contained in the light chain variable region; LCDR3, which has the amino acid sequence of LCDR3 contained in the light chain variable region as shown in any one of SEQ ID NOs: 19 to 21, or has one or more amino acid substitutions, deletions, additions, and / or an amino acid sequence having at least 80% sequence identity therewith and retaining its human papillomavirus binding activity compared to the amino acid sequence of LCDR3 contained in the light chain variable region; and / or, (2) the following three heavy chain variable region complementarity determining regions (HCDRs): HCDR1 having the amino acid sequence of HCDR1 contained in the heavy chain variable region as shown in any one of SEQ ID NOs: 22 to 24, or having one or more amino acid substitutions, deletions, additions, and / or an amino acid sequence having at least 80% sequence identity therewith and retaining its human papillomavirus binding activity compared to the amino acid sequence of HCDR1 contained in the heavy chain variable region; HCDR2 having the amino acid sequence of HCDR2 contained in the heavy chain variable region as shown in any one of SEQ ID NOs: 22 to 24, or having one or more amino acid substitutions, deletions, additions, and / or an amino acid sequence having at least 80% sequence identity thereto and retaining its human papillomavirus binding activity compared to the amino acid sequence of HCDR2 contained in the heavy chain variable region; HCDR3, which has the amino acid sequence of HCDR3 contained in the heavy chain variable region as shown in any one of SEQ ID NOs: 22 to 24, or has one or more amino acids substituted, deleted, added, and / or has at least 80% sequence identity with the amino acid sequence of HCDR3 contained in the heavy chain variable region and retains its human papillomavirus binding activity.

2. The antibody or antigen-binding fragment thereof according to claim 1, characterized in that: The antibody or antigen-binding fragment thereof comprises: (A1) an amino acid sequence of three LCDRs contained in a light chain variable region as shown in SEQ ID NO: 19, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of any one of the three LCDRs contained in the light chain variable region and retaining its human papillomavirus binding activity; and / or an amino acid sequence of three HCDRs contained in a heavy chain variable region as shown in SEQ ID NO: 22, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of any one of the three HCDRs contained in the heavy chain variable region and retaining its human papillomavirus binding activity; or (A2) an amino acid sequence of three LCDRs contained in a light chain variable region as shown in SEQ ID NO: 20, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of any one of the three LCDRs contained in the light chain variable region and retaining its human papillomavirus binding activity; and / or, an amino acid sequence of three HCDRs contained in a heavy chain variable region as shown in SEQ ID NO: 23, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of any one of the three HCDRs contained in the heavy chain variable region and retaining its human papillomavirus binding activity; or (A3) an amino acid sequence such as the three LCDRs contained in the light chain variable region as shown in SEQ ID NO:21, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of any one of the three LCDRs contained in the light chain variable region and retaining its human papillomavirus binding activity; and / or an amino acid sequence such as the three HCDRs contained in the heavy chain variable region as shown in SEQ ID NO:24, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of any one of the three HCDRs contained in the heavy chain variable region and retaining its human papillomavirus binding activity.

3. The antibody or antigen-binding fragment thereof according to claim 1, characterized in that: The antibody or antigen-binding fragment thereof comprises: LCDR1, which has an amino acid sequence as shown in any one of SEQ ID NOs: 1 to 3, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity thereto and retaining its human papillomavirus binding activity; and / or LCDR2, which has an amino acid sequence as shown in any one of SEQ ID NOs: 4 to 6, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity thereto and retaining its human papillomavirus binding activity; and / or LCDR3, which has an amino acid sequence as shown in any one of SEQ ID NOs: 7 to 9, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity thereto and retaining its human papillomavirus binding activity; and / or HCDR1 having an amino acid sequence as shown in any one of SEQ ID NOs: 10 to 12, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity thereto and retaining its human papillomavirus binding activity; and / or HCDR2 having an amino acid sequence as shown in any one of SEQ ID NOs: 13 to 15, or having one or more amino acid substitutions, deletions, additions, and / or an amino acid sequence having at least 80% sequence identity thereto and retaining its human papillomavirus binding activity; and / or HCDR3 having an amino acid sequence as shown in any one of SEQ ID NOs: 16 to 18, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity thereto and retaining its human papillomavirus binding activity; Preferably, the antibody or antigen-binding fragment thereof comprises: (B1) an amino acid sequence such as LCDR1, LCDR2 and LCDR3 as shown in SEQ ID NO:1, SEQ ID NO:4 and SEQ ID NO:7, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of any one of said LCDR1, LCDR2 and LCDR3 and retaining its human papillomavirus binding activity; and / or an amino acid sequence such as HCDR1, HCDR2 and HCDR3 as shown in SEQ ID NO:10, SEQ ID NO:13 and SEQ ID NO:16, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of any one of said HCDR1, HCDR2 and HCDR3 and retaining its human papillomavirus binding activity; or, (B2) an amino acid sequence such as LCDR1, LCDR2 and LCDR3 as shown in SEQ ID NO:2, SEQ ID NO:5 and SEQ ID NO:8, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of any one of said LCDR1, LCDR2 and LCDR3 and retaining its human papillomavirus binding activity; and / or an amino acid sequence such as HCDR1, HCDR2 and HCDR3 as shown in SEQ ID NO:11, SEQ ID NO:14 and SEQ ID NO:17, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of any one of said HCDR1, HCDR2 and HCDR3 and retaining its human papillomavirus binding activity; or, (B3) an amino acid sequence such as LCDR1, LCDR2 and LCDR3 as shown in SEQ ID NO:3, SEQ ID NO:6 and SEQ ID NO:9, or an amino acid sequence having one or several amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of any one of said LCDR1, LCDR2 and LCDR3 and retaining its human papillomavirus binding activity; and / or an amino acid sequence such as HCDR1, HCDR2 and HCDR3 as shown in SEQ ID NO:12, SEQ ID NO:15 and SEQ ID NO:18, or an amino acid sequence having one or several amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity with the amino acid sequence of any one of said HCDR1, HCDR2 and HCDR3 and retaining its human papillomavirus binding activity.

4. The antibody or antigen-binding fragment thereof according to claim 1, characterized in that: The antibody or antigen-binding fragment thereof comprises: a light chain variable region comprising an amino acid sequence as shown in any one of SEQ ID NOs: 19 to 21, or an amino acid sequence having one or more amino acid substitutions, deletions or additions thereto, and / or an amino acid sequence having at least 80% sequence identity thereto and retaining its human papillomavirus binding activity; and / or a heavy chain variable region comprising an amino acid sequence as shown in any one of SEQ ID NOs: 22 to 24, or an amino acid sequence having one or more amino acid substitutions, deletions or additions thereto, and / or an amino acid sequence having at least 80% sequence identity thereto and retaining its human papillomavirus binding activity; Preferably, the antibody or antigen-binding fragment thereof comprises: (C1) a light chain variable region comprising the amino acid sequence shown in SEQ ID NO: 19, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity thereto and retaining its human papillomavirus binding activity; and / or a heavy chain variable region comprising the amino acid sequence shown in SEQ ID NO: 22, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity thereto and retaining its human papillomavirus binding activity; or (C2) a light chain variable region comprising the amino acid sequence as shown in SEQ ID NO: 20, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity thereto and retaining its human papillomavirus binding activity; and / or a heavy chain variable region comprising the amino acid sequence as shown in SEQ ID NO: 23, or an amino acid sequence having one or more amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity thereto and retaining its human papillomavirus binding activity; or (C3) a light chain variable region comprising the amino acid sequence shown in SEQ ID NO: 21, or an amino acid sequence having one or several amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity thereto and retaining its human papillomavirus binding activity; and / or a heavy chain variable region comprising the amino acid sequence shown in SEQ ID NO: 24, or an amino acid sequence having one or several amino acid substitutions, deletions, additions, and / or having at least 80% sequence identity thereto and retaining its human papillomavirus binding activity.

5. The antibody or antigen-binding fragment thereof according to claim 1, characterized in that: The antibody or antigen-binding fragment comprises IgA, IgD, IgE, IgG or IgM type; Preferably, the antibody or antigen-binding fragment comprises scFv, Fab, Fab', (Fab')2, Fv fragment, dsFv, bispecific antibody and multispecific antibody; Preferably, the antibody or antigen-binding fragment is used to neutralize human papillomavirus (HPV); Preferably, the human papillomavirus includes HPV6, HPV11, HPV16, HPV18, HPV31, HPV33, HPV45, HPV52 or HPV58; Preferably, the antibody or antigen-binding fragment comprises a murine antibody, a chimeric antibody, a humanized antibody or a fully human antibody.

6. A nucleic acid molecule encoding the antibody or antigen-binding fragment thereof according to claims 1 to 5.

7. An expression vector comprising the nucleic acid molecule according to claim 6.

8. A host cell comprising the nucleic acid molecule according to claim 6 and / or the expression vector according to claim 7; Preferably, the host cell is selected from prokaryotic cells and eukaryotic cells; Preferably, the prokaryotic cells include bacterial cells, such as Escherichia coli and Streptomyces; Preferably, the eukaryotic cells include yeast cells, mammalian cells, and insect cells; Preferably, the mammal is selected from humans, monkeys, mice, rats, hamsters, goats, sheep, cattle, pigs, dogs or cats; Preferably, the mammalian cells include CHO cells, 293 cells, Vero cells, BHK cells, NS0 cells, SP2 / 0 cells, YO myeloma cells, P3X63 mouse myeloma cells, PER cells, PER.C6 cells or hybridoma cells.

9. A chimeric antigen receptor, comprising an extracellular antigen binding domain, a transmembrane domain and an intracellular signaling domain, wherein the extracellular antigen binding domain comprises the antibody or antigen-binding fragment thereof according to any one of claims 1 to 5.

10. A modified immune cell comprising the chimeric antigen receptor of claim 9. 11 . An antibody-drug conjugate, comprising the antibody or antigen-binding fragment thereof according to any one of claims 1 to 5; and a drug covalently linked to the antibody or antigen-binding fragment thereof.

12. A detection kit, comprising the antibody or antigen-binding fragment thereof according to claims 1 to 5.

13. A composition comprising the antibody or antigen-binding fragment thereof according to claims 1 to 5, and a pharmaceutically acceptable carrier; Preferably, the antibody or antigen-binding fragment thereof comprises: one or more of (A1), (A2) and (A3); Preferably, the antibody or antigen-binding fragment thereof comprises: (A1), and (A2), and (A3).

14. Use of the antibody or antigen-binding fragment thereof according to any one of claims 1 to 5, the nucleic acid molecule according to claim 6, the expression vector according to claim 7, the host cell according to claim 8, the chimeric antigen receptor according to claim 9, the modified immune cell according to claim 10, the antibody-drug conjugate according to claim 11, the detection kit according to claim 12 and / or the composition according to claim 13 in the preparation of a product for diagnosing, preventing and / or treating human papillomavirus infection.

15. A method for detecting human papillomavirus, comprising the step of using the antibody or antigen-binding fragment thereof according to any one of claims 1 to 5 for detection.

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