Antibody specifically binding to TCTP and use thereof

Abstract

The present invention relates to: an antibody specifically binding to TCTP; or an antigen-binding fragment thereof. In addition, the present invention relates to: a method for preparing the antibody or antigen-binding fragment; a composition for preventing or treating cancer, the composition comprising the antibody or antigen-binding fragment; a method for preventing or treating cancer by administering the antibody or antigen-binding fragment to a subject; a composition for diagnosing cancer, the composition comprising the antibody or antigen-binding fragment; a kit for diagnosing cancer, the kit comprising the antibody or antigen-binding fragment; a method for providing information for cancer diagnosis by using the antibody or antigen-binding fragment; a method for screening anticancer drugs by using the antibody or antigen-binding fragment; and a method for providing information for cancer treatment by using the antibody or antigen-binding fragment. The antibody or antigen-binding fragment has excellent effects in inhibiting ERK phosphorylation of cancer cells and inhibiting the proliferation or growth of tumors. The antibody or antigen-binding fragment has excellent anticancer effects, particularly on cancers exhibiting tolerance, resistance, or non-responsiveness to conventional anticancer drugs.

Description

Antibody specifically binding to TCTP and uses thereof

[0001] This application claims the benefit of priority based on Korean Patent Application No. 10-2025-0014800 filed on February 5, 2025, and all contents disclosed in the document of said Korean patent application are incorporated herein as part of this specification.

[0002] The present invention relates to an antibody that specifically binds to TCTP or an antigen-binding fragment thereof. Furthermore, the present invention relates to a method for manufacturing said antibody or antigen-binding fragment, a composition for preventing or treating cancer comprising said antibody or antigen-binding fragment, a method for preventing or treating cancer by administering said antibody or antigen-binding fragment to a subject, a composition for diagnosing cancer comprising said antibody or antigen-binding fragment, a kit for diagnosing cancer comprising said antibody or antigen-binding fragment, a method for providing information for diagnosing cancer using said antibody or antigen-binding fragment, a method for screening anticancer agents using said antibody or antigen-binding fragment, and a method for providing information for treating cancer using said antibody or antigen-binding fragment.

[0003] Despite intensive research on cancer over the past few years, it remains a leading cause of death worldwide. While numerous cancer treatments have been developed, they are not effective for all types of cancer or all patients. Current methods used to treat cancer are relatively non-selective. Treatments involve removing diseased tissue through surgery, reducing the size of solid tumors through radiation therapy, or rapidly killing cancer cells through chemotherapy. In particular, chemotherapy can lead to the development of drug resistance and, in some cases, cause severe side effects that limit the dosage, ultimately excluding the use of potentially effective drugs. Therefore, there is an urgent need to develop targeted and more effective cancer treatments.

[0004] In this regard, targeted anticancer drugs that specifically target only cancer cells, or immunotherapies that utilize the patient's immune system, are being developed. For example, there are about 30 types of targeted anticancer drugs approved in Korea, and immunotherapies approved in Korea include Opdivo (active ingredient: nivolumab), Keytruda (active ingredient: pembrolizumab), Tecentriq (active ingredient: atezolizumab), Imfinzi (active ingredient: durvalumab), and Yervoy (active ingredient: ipilimumab), which are used for various types of cancer, including non-small cell lung cancer, melanoma, renal cell carcinoma, Hodgkin lymphoma, head and neck cancer, and bladder cancer.

[0005] However, there are patients who exhibit resistance, tolerance, or refractoryness to existing anticancer drugs such as targeted therapies and immunotherapies, and accordingly, the need for novel anticancer drugs still exists in the industry.

[0006] The inventors identified a new target for anticancer drugs and conducted research to develop a treatment for cancers that exhibit resistance, tolerance, or refractoryness to existing anticancer drugs by utilizing this. As a result, they succeeded in manufacturing an anti-TCTP antibody that has high binding ability to the TCTP protein and exhibits excellent anticancer effects, thereby completing the present invention.

[0007] Each description and embodiment disclosed in the present invention may be applied to each other description and embodiment. That is, all combinations of the various elements disclosed in the present invention fall within the scope of the present invention. Furthermore, the scope of the present invention should not be considered limited by the specific descriptions provided below.

[0008] Furthermore, terms not specifically defined in this specification should be understood to have the meanings commonly used in the technical field to which the present invention pertains. Additionally, unless specifically defined in the context, the singular includes the plural, and the plural includes the singular.

[0009]

[0010] One aspect of the present invention provides an antibody or an antigen-binding fragment thereof that specifically binds to a Translationally-controlled tumor protein (TCTP).

[0011] As used herein, the term "antibody" refers to an immunoglobulin molecule that is immunologically reactive with a specific antigen or a protein molecule that acts as a receptor specifically recognizing an antigen. Accordingly, in the present invention, "antibody" is interpreted to include all of the following: monoclonal antibodies, polyclonal antibodies, whole antibodies (antibodies consisting of at least two heavy chains and two light chains interconnected by disulfide bonds), antibody fragments, Fab, Fab', F(ab')2, Fv, short-chain Fv (scFv), diabodies, linear antibodies, bispecific antibodies, multispecific antibodies, chimeric antibodies, humanized antibodies, human antibodies, or antigen-binding portions of antibodies. The whole antibodies include IgA, IgD, IgE, IgM, and IgG, and the IgG may include IgG1, IgG2, IgG3, and IgG4 as subtypes. The Fc region of the above antibody may be used with a mutation introduced as needed.

[0012] As used herein, the term “antigen-binding fragment” means an antibody fragment or an antibody analog that retains at least some of the binding specificity of the parent antibody and includes a portion of the antigen-binding region of the parent antibody (e.g., one or more CDRs) or a variable region. The antigen-binding fragment may be, for example, a Fab, Fab', F(ab')2, Fv fragment, scFv, unibody, diabody, linear antibody, nanobody, domain antibody, or a multispecific antibody formed from an antibody fragment.

[0013] As used herein, the term “heavy chain” means a full-length heavy chain and a fragment thereof comprising a heavy chain variable region and a heavy chain constant region. The heavy chain may be any one of the gamma (γ), mu (μ), alpha (α), delta (δ), and epsilon (ε) types.

[0014] As used in this specification, the term “light chain” means a full-length light chain and a fragment thereof, comprising a light chain variable region and a light chain invariant region. The light chain may be of the kappa (κ) and lambda (λ) types.

[0015] As used herein, the term "CDR (complementarity determining region)" refers to the amino acid sequence of a hypervariable region that forms the antigen-binding site, as part of the variable region of antibodies produced by B cells and T cells. The amino acid sequences of the heavy chain and light chain each contain three discontinuously arranged CDRs, namely the heavy chain CDR H1 , CDR H2 , CDR H3 and light chain CDR L1 , CDR L2 , CDR L3 This is included. The CDR is a site involved in antigen recognition and plays a decisive role in the diversity of antigen specificity by providing key contact residues for the binding of antibodies to antigens or epitopes.

[0016] In the present invention, the antibody is a full-length antibody or an antibody fragment having antigen-binding ability, the heavy chain may be any one of gamma (γ), mu (μ), alpha (α), delta (δ), or epsilon (ε) types, and the light chain may be kappa (κ) or lambda (λ) type.

[0017] As used herein, the term "TCTP" is an abbreviation for translationally-controlled tumor protein, and the amino acid sequence constituting TCTP and the base sequence encoding it are known in existing databases (NCBI reference sequence: NP_003286.1, NM_003295.4, etc.). The full-length amino acid sequence of TCTP may be represented, for example, by SEQ ID NO. 1. The TCTP protein is known to be overexpressed in various types of cancer cells, including not only common cancers but also cancers that exhibit resistance, tolerance, or refractoryness to anticancer drugs, and is known to promote the proliferation, migration, invasion, and growth of cancer cells. Therefore, the antibody of the present invention or its antigen-binding fragment, which binds to the TCTP protein with high specificity and affinity, can be usefully employed for the treatment, prevention, improvement, or diagnosis of cancer.

[0018] As used herein, the term "antibody specifically binding to TCTP" means an antibody or an antigen-binding fragment thereof that specifically binds to TCTP to neutralize or inhibit the activity of TCTP. The antibody specifically binding to TCTP may bind to or be reactive to human TCTP proteins and / or animal TCTP proteins, such as those of mice or rats. The term "antibody specifically binding to TCTP" may be used interchangeably with the terms "anti-TCTP antibody" or "αTCTP."

[0019] The anti-TCTP antibody according to the present invention may comprise a heavy chain CDR1 comprising the amino acid sequence GFTFSNYGMH of SEQ ID NO. 2; a heavy chain CDR2 comprising the amino acid sequence YDX1SNKYYADSV of SEQ ID NO. 3; and a heavy chain CDR3 comprising the amino acid sequence GVLDV of SEQ ID NO. 4. The anti-TCTP antibody according to the present invention may comprise a light chain CDR1 comprising the amino acid sequence QGX2X3SX4LA of SEQ ID NO. 5; a light chain CDR2 comprising the amino acid sequence ATX5TLQS of SEQ ID NO. 6; and a light chain CDR3 comprising the amino acid sequence QLNSYPLX6 of SEQ ID NO. 7. Here, X1, X2, X3, X4, X5, and X6 are any amino acids independently of each other. The above amino acids may be arginine (R), histidine (H), lysine (K), aspartic acid (D), glutamic acid (E), serine (S), threonine (T), asparagine (N), glutamine (Q), cysteine ​​(C), glycine (G), proline (P), alanine (A), valine (V), isoleucine (I), leucine (L), methionine (M), phenylalanine (F), tyrosine (Y), or tryptophan (W).

[0020] The sequences disclosed in the present invention include sequences that exhibit substantial identity with the sequences listed in the sequence list. The substantial identity means that, as a result of aligning the two sequences to correspond as much as possible and analyzing them using an algorithm commonly used in the art, they exhibit homology between the sequences of 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more.

[0021] Additionally, the antibody or its antigen-binding fragment of the present invention may include the sequence of the antibody or its antigen-binding fragment described herein, a sequence exhibiting substantial identity with said sequence, as well as biological equivalents thereof, within a range capable of specifically recognizing and binding to the TCTP protein. For example, it may include additional variations in the sequence to improve antibody binding affinity and / or biological properties, and may include additional variations within a range that does not alter the overall activity of the molecule.

[0022] The antibody or its antigen-binding fragment according to the present invention may be a chimeric antibody, a humanized antibody, or a human antibody, but is not limited thereto.

[0023] The term "chimeric antibody" refers to an antibody whose amino acid sequence is derived from two or more species. For example, it may be an antibody produced by combining a variable region derived from a mouse antibody with a constant region of a human antibody. Through such a structural combination, the immunogenicity of the existing mouse antibody can be reduced and its biocompatibility improved.

[0024] The term "humanized antibody" refers to an antibody whose amino acid sequence, originating from a species other than humans (e.g., mouse, llama, camel, non-human primate, etc.), has been redesigned to be similar to the sequence of a human antibody. For example, a humanized antibody can be produced by combining the CDR of a non-human antibody with the framework region of a human antibody to form a humanized variable region, and then linking this to a suitable constant region of a human antibody. While the antibody-antigen binding characteristics of the humanized antibody are maintained, its antigenicity within the human body may be reduced.

[0025] As used herein, the term "human antibody" refers to an antibody having a framework and a CDR region having a variable region derived from a human immunoglobulin sequence. In the present invention, the human antibody may include amino acid residues not encoded by a human-derived immunoglobulin sequence (e.g., mutations introduced by in vitro random or site-specific mutations, or mutations introduced by in vivo somatic mutations).

[0026] The antibody or its antigen-binding fragment according to the present invention may form a bispecific antibody, a multispecific antibody, an antibody-drug conjugate (ADC), or a fusion protein, but is not limited thereto, and may form any form in which said antibody or its antigen-binding fragment may be used or included in the art.

[0027]

[0028] Another aspect of the present invention provides a nucleic acid encoding the antibody or its antigen-binding fragment.

[0029] In the nucleic acid according to the present invention, unless specifically stated otherwise, related terms are understood to have the same meaning as the terms described above.

[0030] As used herein, the term "nucleic acid" has a meaning that comprehensively includes DNA (gDNA and cDNA) and RNA. Nucleotides, which form the basic building blocks of nucleic acid molecules, include not only natural nucleotides but also analogue nucleotides in which sugar or base sites are modified (Scheit, Nucleotide Analogs, John Wiley, New York (1980); Uhlman and Peyman, Chemical Reviews, 90:543-584 (1990)).

[0031] The sequence of a nucleic acid molecule encoding the antibody of the present invention or its antigen-binding fragment may be modified, and said modification includes the addition, deletion, or non-conservative or conservative substitution of nucleotides.

[0032] A nucleic acid molecule encoding an antibody or an antigen-binding fragment thereof according to the present invention comprises a nucleotide sequence encoding an amino acid sequence constituting the antibody or the antigen-binding fragment thereof. The nucleotide sequence comprises, taking into account the degeneracy of codons, a nucleotide sequence comprising functionally equivalent codons or codons encoding the same amino acid (e.g., due to the degeneracy of codons, there are six codons for arginine or serine), or codons encoding biologically equivalent amino acids.

[0033] Considering the variant having the aforementioned biological equivalent activity, the nucleic acid molecule of the present invention encoding the antibody or its antigen-binding fragment comprises a sequence exhibiting substantial identity therewith. The substantial identity means that, as a result of aligning the two sequences to correspond as much as possible and analyzing them using an algorithm commonly used in the art, they exhibit homology between the sequences of 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more. Accordingly, sequences having high homology with the sequences represented by SEQ ID NOs 1 to 127 of the present invention, for example, sequences having high homology of 70% or more, specifically 80% or more, and more specifically 90% or more, should also be interpreted as being included within the scope of the present invention.

[0034]

[0035] Another aspect of the present invention provides a vector comprising the nucleic acid and a host cell transformed with the vector.

[0036] In the vector or cell according to the present invention, unless specifically stated otherwise, related terms are understood to have the same meaning as the terms described above.

[0037] As used herein, the term "vector" refers to any that can be inserted into a host cell to perform gene replication. Such vectors include plasmids, linear nucleic acids, cosmids, RNA vectors, viral vectors, etc. Viral vectors include, but are not limited to, retroviruses, adenoviruses, and adeno-associated viruses. The recombinant vector system of the present invention can be constructed through various methods known in the art. Furthermore, the vector of the present invention can be constructed as a vector for cloning or expression, and can be constructed using a prokaryotic or eukaryotic cell as a host.

[0038] As used herein, the term "host cell" refers to a cell or its progenitor capable of stably and continuously cloning and expressing the antibody of the present invention or its antigen-binding fragment, and said host cell comprises an expression vector of said antibody or its antigen-binding fragment. The said host cell may be a fungus, a eukaryotic cell, a plant cell, an animal cell, etc., but is not limited thereto. The said host cell may be, for example, a Chinese hamster ovary (CHO) cell, a COS7 cell (monkey kidney cell), an NSO cell, SP2 / 0, W138, a baby hamster kidney (BHK) cell, an MDCK, a myeloma cell, a HELA cell, a HuT 78 cell, or a 293 cell, etc. However, it is not limited thereto, and any known host cell may be used as the host cell of the present invention. The above host cells may include all cells isolated in vitro from the subject, such as cells derived from or obtained from the subject, or commercially available cell lines.

[0039] As used herein, the term "subject" includes a human or any non-human animal, and said non-human animal may be a vertebrate, e.g., a primate, a dog, a cow, a horse, a pig, a rodent, e.g., a mouse, a rat, a guinea pig, etc. In this specification, said "subject" may be used interchangeably with "individual" or "patient."

[0040] The vector of the present invention may be one in which a nucleic acid molecule encoding the antibody or its antigen-binding fragment is operably linked to a promoter.

[0041] As used herein, the term “operably linked” means the functional linkage of two molecules, such as domains, polypeptides, nucleic acid sequences, nucleic acid expression regulatory sequences (e.g., promoters, signal sequences, or arrays of transcription factor binding sites). For example, when a nucleic acid expression regulatory sequence is linked to another nucleic acid sequence, the expression regulatory sequence controls the transcription and / or translation of the other nucleic acid sequence.

[0042] When the vector of the present invention has a eukaryotic cell as a host, a promoter derived from the genome of a mammalian cell or a promoter derived from a mammalian virus may be used in the vector, and may include a polyadenylation sequence as a transcription termination sequence, but is not limited thereto, and the vector may be constructed using various methods and sequences known in the art.

[0043] When the vector of the present invention has a prokaryotic cell as a host, the vector may include a promoter capable of proceeding transcription in the prokaryotic cell, a ribosome binding site for initiating translation, and a transcription / translation termination sequence, but is not limited thereto, and the vector may be constructed using various methods and sequences known in the art.

[0044] As used herein, the term "transformation" means introducing DNA into a host so that the DNA becomes replicable as a chromosomal factor or through the completion of chromosomal integration, thereby artificially inducing a genetic change by introducing external DNA into a cell. Transforming a host cell can be performed using any transformation method, and various transformation or transfection methods known in the art may be used without limitation.

[0045]

[0046] Another aspect of the present invention provides a method for producing the antibody or its antigen-binding fragment, comprising the step of introducing the vector into a separated host cell.

[0047] In the method for preparing an antibody or an antigen-binding fragment thereof according to the present invention, unless specifically stated otherwise, related terms are understood to have the same meaning as the terms described above.

[0048] Specifically, the method for producing the antibody or its antigen-binding fragment of the present invention is,

[0049] (a) a step of preparing a vector comprising a nucleic acid encoding an antibody or an antigen-binding fragment thereof;

[0050] (b) a step of introducing the vector of the present invention into a separated host cell;

[0051] (c) a step of culturing the host cells; and

[0052] (d) may include the step of obtaining an antibody or an antigen-binding fragment thereof from the host cell.

[0053] The step (a) of preparing a vector comprising a nucleic acid encoding an antibody of the present invention or an antigen-binding fragment thereof is a step of preparing a vector according to the present invention.

[0054] The step (b) of introducing the vector of the present invention into isolated host cells may be a step of preparing a transformant comprising a vector expressing the antibody or an antigen-binding fragment thereof, and the step of preparing the transformant may be to transform the host cells.

[0055] As used in this specification, the term "transformed organism" refers to a cell in which an artificial genetic change has occurred by introducing external DNA into a host cell, such that the DNA becomes replicable as a chromosomal factor or through the completion of chromosomal integration, or to an organism into which said transformed cell has been introduced.

[0056] The step (c) of culturing the host cells of the present invention can be performed according to the media and culture conditions known in the art.

[0057] The step (d) of obtaining an antibody or an antigen-binding fragment thereof from a host cell of the present invention may be a step of recovering the antibody or the antigen-binding fragment thereof from a culture medium obtained through the step (c). The antibody or the antigen-binding fragment thereof may be used in an unpurified state, or may be used after further purification and concentration.

[0058]

[0059] Another aspect of the present invention provides a pharmaceutical composition for the prevention or treatment of cancer, comprising the antibody or an antigen-binding fragment thereof.

[0060] In the pharmaceutical composition according to the present invention, unless specifically stated otherwise, related terms are understood to have the same meaning as the terms described above.

[0061] As used in this invention, the term "treatment" refers to all instances where cancer improves, reverses, or is completely cured by the administration of the composition according to this invention. As used in this invention, the term "prevention" refers to all instances where the occurrence or recurrence of cancer is suppressed, delayed, or prevented by the administration of the composition according to this invention.

[0062] In the present invention, the cancer may be, but is not limited to, bone cancer, lung cancer, non-small cell lung cancer, head cancer, neck cancer, thyroid cancer, parathyroid cancer, stomach cancer, liver cancer, pancreatic cancer, skin cancer, melanoma, small intestine cancer, colorectal cancer, rectal cancer, pro-anal cancer, colon cancer, uterine cancer, breast cancer, ovarian cancer, fallopian tube cancer, endometrial cancer, cervical cancer, vaginal cancer, vulvar cancer, Hodgkin's disease, esophageal cancer, endocrine gland cancer, adrenal cancer, soft tissue sarcoma, urethral cancer, penile cancer, prostate cancer, bladder cancer, kidney cancer, ureteral cancer, renal cell carcinoma, renopelvic carcinoma, central nervous system tumor, central nervous system lymphoma, spinal cord tumor, glioblastoma, brainstem glioma, or pituitary adenoma.

[0063] The above-mentioned cancer may exhibit anticancer drug resistance, resistance, or refractoryness. The term "anticancer drug" used in the present invention refers to a substance that exhibits a preventive or therapeutic effect against cancer, specifically a substance capable of killing cancer cells or tumors, or inhibiting their growth. The terms "anticancer drug resistance" or "anticancer drug resistance" used in the present invention refer to a situation where, when treating a cancer patient with an anticancer drug, there is no effect from the beginning of treatment, or although there is a therapeutic effect initially, the cancer treatment effect decreases or is lost during the course of continuous treatment. The term "anticancer drug refractoryness" used in the present invention refers to a situation where, when treating a cancer patient with an anticancer drug, there is no effect from the beginning of treatment, or the response to treatment does not persist for a long period.

[0064] TCTP is known to be increased in cancer cells and to cause resistance, tolerance, or refractoryness to anticancer agents such as chemotherapy drugs or immunotherapy drugs. Therefore, since the antibody or its antigen-binding fragment according to the present invention can reduce or lower the activity or expression of TCTP protein, it can be usefully utilized for the prevention or treatment of cancers exhibiting resistance, tolerance, or refractoryness to existing anticancer agents. The aforementioned existing anticancer agents may be those reported to have caused resistance, tolerance, or refractoryness, and include, for example, chemotherapy drugs such as 5-FU, methotrexate, gemcitabine, cytarabine, paclitaxel, vinorelbine, cisplatin, oxaliplatin, and irinotecan; Targeted anticancer agents such as bortezomib, cetuximab, crizotinib, dasatinib, gefitinib, imatinib, vemurafenib; and / or immunotherapies such as CTLA-4 inhibitors, PD-1 inhibitors, PD-L1 inhibitors, and ACT (adoptive cell therapy), but not limited thereto.

[0065] The pharmaceutical composition of the present invention may be administered to a subject requiring prevention or treatment of cancer by containing an effective amount of the antibody or its antigen-binding fragment.

[0066] As used herein, the term "administration" means the physical introduction of a composition to a subject using any of the various methods and delivery systems known to those skilled in the art. Such administration may be performed, for example, by oral administration, or by intravenous, intramuscular, subcutaneous, intraperitoneal, spinal, or other parenteral administration, such as by injection or infusion, but is not limited thereto. Such administration may be performed, for example, as a single dose, multiple doses, or over one or more extended periods.

[0067] The above effective dose may be a "therapeutic effective dose" or a "preventive effective dose." As used herein, the term "therapeutic effective dose" refers to any amount that, when a drug or therapeutic agent is used alone or in combination with other therapeutic agents, can result in a reduction in the severity of disease symptoms, an increase in the frequency and duration of symptom-free periods, or the prevention of damage or disability caused by disease suffering. As used herein, the term "preventive effective dose" refers to any amount that inhibits the occurrence or recurrence of cancer in a subject. The effective dose, administration cycle, and frequency may be determined based on factors including the subject's severity, age, sex, body weight, drug activity, sensitivity to the drug, administration time, route of administration and elimination rate, duration of treatment, concurrently used drugs, and other factors well known in the medical field. For example, 0.1 to 100 mg / kg of the composition of the present invention may be administered once or several times daily, or at intervals of several days to several months.

[0068] Additionally, the pharmaceutical composition may further include suitable carriers, excipients, and diluents commonly used in the manufacture of pharmaceutical compositions. Carriers, excipients, and diluents that may be included in the composition may be, for example, lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, microcrystalline cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and mineral oil, but are not limited thereto.

[0069] In addition, the above pharmaceutical composition may be administered in combination with other therapeutic agents. In this case, the pharmaceutical composition of the present invention and the other therapeutic agents may be administered simultaneously, sequentially, or individually. The other therapeutic agents may be drugs, such as compounds or proteins, having effects of preventing, treating, and improving cancer, but are not limited thereto.

[0070] In addition, the above pharmaceutical composition may be formulated to be administered simultaneously, sequentially, or individually with other therapeutic agents. For example, the antibody or its antigen-binding fragment and other therapeutic agents may be administered simultaneously as a single formulation, or they may be administered simultaneously, sequentially, or individually as separate formulations. The antibody or its antigen-binding fragment and other therapeutic agents included in the pharmaceutical composition of the present invention may be formulated separately in individual containers or formulated together in the same container. Furthermore, the antibody or its antigen-binding fragment and other therapeutic agents included in the pharmaceutical composition of the present invention may have the same or different pharmaceutical effective dose, administration time, administration interval, administration route, treatment duration, etc. As used herein, the term "simultaneous" administration means administering the antibody or its antigen-binding fragment and other therapeutic agents at once as a single formulation, or administering the antibody or its antigen-binding fragment and other therapeutic agents at once as separate formulations. As used herein, the term "sequential" administration means administering an antibody or its antigen-binding fragment and other therapeutic agents relatively continuously, allowing for the minimum possible time consumed in the administration interval. As used herein, the term "individual" administration means administering an antibody or its antigen-binding fragment and other therapeutic agents at regular time intervals.

[0071]

[0072] Another aspect of the present invention provides a method for preventing or treating cancer, comprising the step of administering the antibody or an antigen-binding fragment thereof.

[0073] In the prevention or treatment method according to the present invention, unless specifically stated otherwise, related terms are understood to have the same meaning as the terms described above.

[0074]

[0075] Another aspect of the present invention provides a composition for diagnosing cancer, comprising the antibody or an antigen-binding fragment thereof.

[0076] Another aspect of the present invention provides a cancer diagnostic kit comprising the antibody or an antigen-binding fragment thereof.

[0077] In the cancer diagnostic composition and kit according to the present invention, unless specifically stated otherwise, related terms are understood to have the same meaning as the terms described above.

[0078] The TCTP protein disclosed in the present invention is known to be overexpressed in various types of cancer cells, including not only common cancers but also cancers that exhibit resistance, tolerance, or refractoryness to anticancer drugs. Therefore, the antibody of the present invention or its antigen-binding fragment, which binds to the TCTP protein with high specificity and affinity, can be usefully used for the diagnosis of cancer.

[0079] Cancer diagnosis according to the present invention can be performed by reacting the antibody or its antigen-binding fragment with a biological sample to confirm whether cancer has developed or the possibility of developing it.

[0080] Specifically, cancer diagnosis according to the present invention can be performed by reacting the antibody or its antigen-binding fragment with a biological sample to confirm whether cancer has developed or the possibility of developing cancer, and more specifically, by contacting the antibody or its antigen-binding fragment with a biological sample to confirm whether an antigen-antibody complex is formed.

[0081] The term “biological sample” as used in this specification includes, but is not limited to, tissues, cells, blood, serum, plasma, tissue autopsy samples (brain, skin, lymph nodes, spinal cord), body fluids, etc.

[0082] As used herein, the term "antigen-antibody complex" refers to a complex of a TCTP protein antigen in a sample and an antibody of the present invention that recognizes it, or an antigen-binding fragment thereof. The formation of such an antigen-antibody complex can be detected by any method, such as a colorimetric method, an electrochemical method, a fluorimetric method, a luminometric method, a particle counting method, a visual assessment method, or a scintillation counting method, but is not limited thereto, and various applications are possible according to methods known in the art. In addition, various labels may be used to detect the antigen-antibody complex. Specific examples include enzymes, fluorescent agents, ligands, luminescent agents, microparticles, or radioactive isotopes, but are not limited thereto. In this case, the enzymes used as detection labels include acetylcholinesterase, alkaline phosphatase, β-D-galactosidase, horseradish peroxidase, β-latamase, etc., and the fluorescent agents include fluorescein, Eu 3+ , Eu 3+ It includes chelates or cryptates, etc., as ligands include biotin derivatives, etc., as luminescent materials include acridinium esters, isoluminol derivatives, etc., as microparticles include colloidal gold, colored latex, etc., and as radioactive isotopes 57 Co, 3 H, 125 I, 125 It may include, but is not limited to, I-Bolton Hunter reagents.

[0083] The diagnostic composition or kit of the present invention may be manufactured to be suitable for various immunoassays or immunostainings. The immunoassay or immunostaining includes, but is not limited to, enzyme-linked immunosorbent assay (ELISA), immunofluorescence, western blotting, immunohistochemistry staining, flow cytometry, immunocytochemistry, immunoprecipitation assay, radioimmunoassay (RIA), and protein chips.

[0084]

[0085] Another aspect of the present invention provides a method for providing information for diagnosing cancer in a subject, comprising the step of detecting or quantifying the expression or activity level of a TCTP protein in a biological sample separated from a subject suspected of having cancer, using the antibody or antigen-binding fragment.

[0086] Another aspect of the present invention provides a method for providing information for selecting a cancer treatment target, comprising the step of detecting or quantifying the expression or activity level of a TCTP protein in a biological sample separated from a cancer treatment target using the antibody or antigen binding fragment.

[0087] Another aspect of the present invention provides a method for providing information for selecting a cancer treatment subject, comprising: a step of detecting or quantifying the expression or activity level of a TCTP protein in a biological sample separated from a subject to be treated for cancer using the antibody or antigen-binding fragment; and a step of selecting a subject to be administered the antibody or antigen-binding fragment as a subject in which the expression or activity level of the TCTP protein is detected or quantified at a higher level than a control group.

[0088] Another aspect of the present invention provides a method for providing information for predicting anticancer drug resistance, resistance, or refractoryness of a subject, comprising the step of detecting or quantifying the expression or activity level of a TCTP protein in a biological sample isolated from a subject suspected of exhibiting resistance, resistance, or refractoryness to an anticancer drug using the antibody or antigen binding fragment.

[0089] In the method for providing information according to the present invention, unless specifically stated otherwise, related terms are understood to have the same meaning as the terms described above.

[0090] In one embodiment according to the present invention, the method for providing information for cancer diagnosis or the method for providing information for selecting a cancer treatment subject may be a method for diagnosing cancer, and the method for providing information for predicting anticancer drug resistance, resistance, or refractoryness of the subject may be a method for diagnosing anticancer drug resistance, resistance, or refractoryness.

[0091] The method for providing information according to the present invention can be performed by contacting the antibody or its antigen-binding fragment with a biological sample to confirm whether an antigen-antibody complex is formed.

[0092] Specifically, the method for providing information according to the present invention may include: (a) a step of treating a biological sample separated from a subject with an antibody according to the present invention or an antigen-binding fragment thereof to form an antigen-antibody complex; (b) a step of detecting or quantifying the antigen-antibody complex formed in step (a) to analyze the expression or activity level of a TCTP protein; and (c) a step of comparing the expression or activity level of a TCTP protein analyzed in step (b) with a control group, and if the expression or activity level of the TCTP protein is higher than that of the control group, determining that the subject has developed cancer or is likely to develop cancer.

[0093] Additionally, the method for providing information according to the present invention may include: (a) a step of treating a biological sample isolated from a cancer patient with an antibody according to the present invention or an antigen-binding fragment thereof to form an antigen-antibody complex; (b) a step of detecting or quantifying the antigen-antibody complex formed in step (a) to analyze the expression or activity level of a TCTP protein; and (c) a step of comparing the expression or activity level of a TCTP protein analyzed in step (b) with a control group, and if the expression or activity level of the TCTP protein is higher than that of the control group, determining that the cancer patient exhibits or is likely to exhibit anticancer drug resistance, resistance, or refractoryness.

[0094] In this case, the control group may be a normal person who has not developed cancer, a person who has developed cancer and been cured, or a biological sample isolated from them.

[0095]

[0096] Another aspect of the present invention provides an anticancer drug screening method comprising: (a) treating cancer cells with an anticancer drug candidate substance; (b) measuring the expression or activity level of a TCTP protein in cancer cells treated with the anticancer drug candidate substance using the antibody or an antigen-binding fragment thereof; and (c) determining the anticancer drug candidate substance treated in step (a) as an anticancer drug if the expression or activity level of the TCTP protein in step (b) is reduced compared to a negative control group.

[0097] In the screening method according to the present invention, unless specifically stated otherwise, related terms are understood to have the same meaning as the terms described above.

[0098] Step (a) above is a step of treating cancer cells with a candidate anticancer drug, and can be performed using methods known in the art. For example, the candidate anticancer drug may be treated by treating cancer cells with the candidate anticancer drug and culturing them together, or by administering it into a living organism containing cancer cells, but is not limited thereto, and a person skilled in the art may use a method suitable for the purpose of the present invention.

[0099] As used in this specification, the term "anticancer agent" may be used interchangeably with the term "drug," and the term "treatment" may be used interchangeably with the terms "addition" or "administration." As used in this specification, the term "anticancer agent candidate" refers to a substance expected to exhibit a preventive or therapeutic effect against cancer, specifically a substance expected to kill cancer cells or tumors, or to inhibit their growth. The anticancer agent or anticancer agent candidate may include, without limitation, but is not limited to, compounds, proteins, fusion proteins, compound-protein complexes, drug-protein complexes, antibodies, compound-antibody complexes, antibody-drug conjugates (ADCs), amino acids, peptides, viruses, carbohydrates, lipids, nucleic acids, extracts, fractions, etc.

[0100] Step (b) above is a step of measuring the expression or activity level of TCTP protein, and any method known to those skilled in the art may be used. Specific examples may include Western blotting, co-immunoprecipitation assay, ELISA, tissue immunostaining, and flow cytometry analysis, but are not limited thereto, and those skilled in the art may use a method suitable for the purpose of the present invention.

[0101] Step (c) above is a step of determining whether the anticancer drug candidate substance can be used as an anticancer drug by comparing the expression or activity level of the TCTP protein measured in Step (b) above with a negative control group. The negative control group may be a group not treated with the anticancer drug candidate substance, a group treated with a non-specific antibody such as an IgG antibody, etc. Since TCTP is overexpressed in cancer cells and promotes the proliferation, migration, invasion, and growth of cancer cells, an anticancer drug candidate substance that reduces the expression or activity level of the TCTP protein can be used as an anticancer drug. Optionally, since TCTP is overexpressed in cancer cells that exhibit resistance, tolerance, or refractoryness to anticancer drugs and promotes the proliferation, migration, invasion, and growth of cancer cells that exhibit resistance, tolerance, or refractoryness to anticancer drugs, an anticancer drug candidate substance that reduces the expression or activity level of the TCTP protein can be used as an anticancer drug for the prevention or treatment of cancer that exhibits resistance, tolerance, or refractoryness to anticancer drugs.

[0102]

[0103] Another aspect of the present invention provides an antibody that specifically binds to TCTP according to the present invention or an antigen-binding fragment thereof, or a pharmaceutical composition comprising the same, for use in the treatment, improvement, or prevention of cancer of a subject.

[0104] Another aspect of the present invention provides the use of an antibody that specifically binds to TCTP or an antigen-binding fragment thereof, or a pharmaceutical composition comprising the same, for treating, improving, or preventing cancer of a subject.

[0105] In the use of the antibody or its antigen-binding fragment according to the present invention and the pharmaceutical composition containing the same, related terms are understood to have the same meaning as the terms described above, unless specifically stated otherwise.

[0106] According to one embodiment of the present invention, the antibody or its antigen-binding fragment, or the pharmaceutical composition may be used as a medicine for the treatment or prevention of cancer, or may be used in the preparation or manufacture thereof.

[0107] The antibody or its antigen-binding fragment that specifically binds to TCTP according to the present invention inhibits ERK phosphorylation in cancer cells and has excellent effects in inhibiting the proliferation or growth of tumors. In particular, it has excellent anticancer effects against cancers that exhibit resistance, tolerance, or refractoryness to existing anticancer drugs.

[0108] Figures 1a to 1d are graphs showing the relative expression levels of ERK phosphorylation (pERK) protein relative to ERK protein when each cancer cell is treated with an anti-TCTP antibody according to the present invention or a comparative anti-TCTP antibody (Sino, Abnova, Santacruz).

[0109] Figures 2a to 2c are graphs showing the degree of cancer cell death by treatment with an anti-TCTP antibody according to the present invention or a comparative anti-TCTP antibody (Sino, Abnova, Santacruz).

[0110] Figures 3a to 3e are graphs showing the degree of tumor growth inhibition in vivo by treatment with the anti-TCTP antibody according to the present invention or a comparative anti-TCTP antibody (Sino, Abnova, Santacruz).

[0111] FIGS. 4 to 10 are graphs showing the degree of in vivo tumor growth inhibition by treatment with the anti-TCTP antibody alone or in combination with the anti-PD-1 antibody or the anti-PD-L1 antibody according to the present invention. In the graphs, * indicates p < 0.05, ** indicates p < 0.01, *** indicates p < 0.001, and **** indicates p < 0.0001.

[0112] Figure 11 is a graph showing the degree of in vivo tumor growth inhibition by anti-TCTP antibody treatment according to the present invention. In the graph, **** means p < 0.0001.

[0113] FIGS. 12 to 14 are graphs showing the degree of in vivo tumor growth inhibition by treatment with an anti-TCTP antibody, an anti-PD-1 antibody, or an anti-VEGF antibody according to the present invention. In the graphs, * indicates p < 0.05, ** indicates p < 0.01, *** indicates p < 0.001, and **** indicates p < 0.0001.

[0114] Figures 15 and 16 respectively show a photograph of a tumor taken by luciferin luminescence in an orthotopic tumor mouse model and a graph showing the tumor growth inhibition rate after antibody administration.

[0115] Figure 17 is a graph showing the degree of binding between TCTP and TLR2 according to antibody concentration.

[0116] The present invention will be explained in more detail below by way of examples. However, these examples are intended to illustrate the invention and the scope of the invention is not limited by these examples.

[0117]

[0118] Example 1. Preparation of anti-TCTP antibody

[0119] The inventors selected Fab antibody phage clones exhibiting TCTP binding ability using a human synthetic Fab phage library and phage display technology. The CDR sequences for the selected Fab antibodies are shown in Table 1. The obtained CDR sequences are IgG4 S228PAnti-TCTP antibodies that bind with high affinity to TCTP proteins were selected by transplanting them into a backbone. Subsequently, a recombinant plasmid was constructed by cloning a gene encoding an amino acid sequence containing the sequences of heavy chains CDR1 to CDR3 and light chains CDR1 to CDR3 in Table 1 into an expression vector. The recombinant plasmid prepared above was transfected into CHO cells to establish a high-expression cell line capable of stably producing anti-TCTP antibodies. The established cell line was cultured in large quantities in a bioreactor through a stepwise scale-up process, and anti-TCTP antibodies were obtained after culture.

[0120] CDR Type Amino Acid Sequence Sequence Number Heavy Chain CDR1GFTFSNYGMH Sequence Number Double Chain CDR2YDX1SNKYYADSV Sequence Number Triple Chain CDR3GVLDV Sequence Number Quadruple Light Chain CDR1QGX2X3SX4LA Sequence Number Five Light Chain CDR2ATX5TLQS Sequence Number Six Light Chain CDR3QLNSYPLX6 Sequence Number 7

[0121] In Table 1 above, X1, X2, X3, X4, X5, and X6 are any amino acids independently of each other. Representative examples of anti-TCTP antibodies having the sequences listed in Table 1 above are AO2P, AO2M1, AO2M2, AO2M3, AO2M4, AO2M5, AO2M6, AO2M7, AO2M8, AO2M9, AO2M10, AO2M11, AO2M12, AO2M13, AO2M14, AO2M15, AO2M16, AO2M17, AO2M18, and AO2M19, and their sequences are as follows. The anti-TCTP antibody AO2P comprises a heavy chain CDR1 containing the amino acid sequence of SEQ ID NO. 2, a heavy chain CDR2 containing the amino acid sequence of SEQ ID NO. 8, a heavy chain CDR3 containing the amino acid sequence of SEQ ID NO. 4, a light chain CDR1 containing the amino acid sequence of SEQ ID NO. 9, a light chain CDR2 containing the amino acid sequence of SEQ ID NO. 10, and SEQ ID NO. 11. It includes a light chain CDR3 containing an amino acid sequence, or a heavy chain variable region containing the amino acid sequence of SEQ ID NO. 12 and a light chain variable region containing the amino acid sequence of SEQ ID NO. 13.

[0122] The anti-TCTP antibody AO2M1 comprises a heavy chain CDR1 having the amino acid sequence of SEQ ID NO. 2, a heavy chain CDR2 having the amino acid sequence of SEQ ID NO. 14, a heavy chain CDR3 having the amino acid sequence of SEQ ID NO. 4, a light chain CDR1 having the amino acid sequence of SEQ ID NO. 15, a light chain CDR2 having the amino acid sequence of SEQ ID NO. 16, and a light chain CDR3 having the amino acid sequence of SEQ ID NO. 17, or comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO. 18 and a light chain variable region having the amino acid sequence of SEQ ID NO. 19.

[0123] The anti-TCTP antibody AO2M2 comprises a heavy chain CDR1 having the amino acid sequence of SEQ ID NO. 2, a heavy chain CDR2 having the amino acid sequence of SEQ ID NO. 20, a heavy chain CDR3 having the amino acid sequence of SEQ ID NO. 4, a light chain CDR1 having the amino acid sequence of SEQ ID NO. 21, a light chain CDR2 having the amino acid sequence of SEQ ID NO. 22, and a light chain CDR3 having the amino acid sequence of SEQ ID NO. 23, or comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO. 24 and a light chain variable region having the amino acid sequence of SEQ ID NO. 25.

[0124] The anti-TCTP antibody AO2M3 comprises a heavy chain CDR1 having the amino acid sequence of SEQ ID NO. 2, a heavy chain CDR2 having the amino acid sequence of SEQ ID NO. 26, a heavy chain CDR3 having the amino acid sequence of SEQ ID NO. 4, a light chain CDR1 having the amino acid sequence of SEQ ID NO. 27, a light chain CDR2 having the amino acid sequence of SEQ ID NO. 28, and a light chain CDR3 having the amino acid sequence of SEQ ID NO. 29, or comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO. 30 and a light chain variable region having the amino acid sequence of SEQ ID NO. 31.

[0125] The anti-TCTP antibody AO2M4 comprises a heavy chain CDR1 having the amino acid sequence of SEQ ID NO. 2, a heavy chain CDR2 having the amino acid sequence of SEQ ID NO. 32, a heavy chain CDR3 having the amino acid sequence of SEQ ID NO. 4, a light chain CDR1 having the amino acid sequence of SEQ ID NO. 33, a light chain CDR2 having the amino acid sequence of SEQ ID NO. 34, and a light chain CDR3 having the amino acid sequence of SEQ ID NO. 35, or comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO. 36 and a light chain variable region having the amino acid sequence of SEQ ID NO. 37.

[0126] The anti-TCTP antibody AO2M5 comprises a heavy chain CDR1 having the amino acid sequence of SEQ ID NO. 2, a heavy chain CDR2 having the amino acid sequence of SEQ ID NO. 38, a heavy chain CDR3 having the amino acid sequence of SEQ ID NO. 4, a light chain CDR1 having the amino acid sequence of SEQ ID NO. 39, a light chain CDR2 having the amino acid sequence of SEQ ID NO. 40, and a light chain CDR3 having the amino acid sequence of SEQ ID NO. 41, or comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO. 42 and a light chain variable region having the amino acid sequence of SEQ ID NO. 43.

[0127] The anti-TCTP antibody AO2M6 comprises a heavy chain CDR1 having the amino acid sequence of SEQ ID NO. 2, a heavy chain CDR2 having the amino acid sequence of SEQ ID NO. 44, a heavy chain CDR3 having the amino acid sequence of SEQ ID NO. 4, a light chain CDR1 having the amino acid sequence of SEQ ID NO. 45, a light chain CDR2 having the amino acid sequence of SEQ ID NO. 46, and a light chain CDR3 having the amino acid sequence of SEQ ID NO. 47, or comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO. 48 and a light chain variable region having the amino acid sequence of SEQ ID NO. 49.

[0128] The anti-TCTP antibody AO2M7 comprises a heavy chain CDR1 having the amino acid sequence of SEQ ID NO. 2, a heavy chain CDR2 having the amino acid sequence of SEQ ID NO. 50, a heavy chain CDR3 having the amino acid sequence of SEQ ID NO. 4, a light chain CDR1 having the amino acid sequence of SEQ ID NO. 51, a light chain CDR2 having the amino acid sequence of SEQ ID NO. 52, and a light chain CDR3 having the amino acid sequence of SEQ ID NO. 53, or comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO. 54 and a light chain variable region having the amino acid sequence of SEQ ID NO. 55.

[0129] The anti-TCTP antibody AO2M8 comprises a heavy chain CDR1 having the amino acid sequence of SEQ ID NO. 2, a heavy chain CDR2 having the amino acid sequence of SEQ ID NO. 56, a heavy chain CDR3 having the amino acid sequence of SEQ ID NO. 4, a light chain CDR1 having the amino acid sequence of SEQ ID NO. 57, a light chain CDR2 having the amino acid sequence of SEQ ID NO. 58, and a light chain CDR3 having the amino acid sequence of SEQ ID NO. 59, or comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO. 60 and a light chain variable region having the amino acid sequence of SEQ ID NO. 61.

[0130] The anti-TCTP antibody AO2M9 comprises a heavy chain CDR1 having the amino acid sequence of SEQ ID NO. 2, a heavy chain CDR2 having the amino acid sequence of SEQ ID NO. 62, a heavy chain CDR3 having the amino acid sequence of SEQ ID NO. 4, a light chain CDR1 having the amino acid sequence of SEQ ID NO. 63, a light chain CDR2 having the amino acid sequence of SEQ ID NO. 64, and a light chain CDR3 having the amino acid sequence of SEQ ID NO. 65, or comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO. 66 and a light chain variable region having the amino acid sequence of SEQ ID NO. 67.

[0131] The anti-TCTP antibody AO2M10 comprises a heavy chain CDR1 having the amino acid sequence of SEQ ID NO. 2, a heavy chain CDR2 having the amino acid sequence of SEQ ID NO. 68, a heavy chain CDR3 having the amino acid sequence of SEQ ID NO. 4, a light chain CDR1 having the amino acid sequence of SEQ ID NO. 69, a light chain CDR2 having the amino acid sequence of SEQ ID NO. 70, and a light chain CDR3 having the amino acid sequence of SEQ ID NO. 71, or comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO. 72 and a light chain variable region having the amino acid sequence of SEQ ID NO. 73.

[0132] The anti-TCTP antibody AO2M11 comprises a heavy chain CDR1 having the amino acid sequence of SEQ ID NO. 2, a heavy chain CDR2 having the amino acid sequence of SEQ ID NO. 74, a heavy chain CDR3 having the amino acid sequence of SEQ ID NO. 4, a light chain CDR1 having the amino acid sequence of SEQ ID NO. 75, a light chain CDR2 having the amino acid sequence of SEQ ID NO. 76, and a light chain CDR3 having the amino acid sequence of SEQ ID NO. 77, or comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO. 78 and a light chain variable region having the amino acid sequence of SEQ ID NO. 79.

[0133] The anti-TCTP antibody AO2M12 comprises a heavy chain CDR1 having the amino acid sequence of SEQ ID NO. 2, a heavy chain CDR2 having the amino acid sequence of SEQ ID NO. 80, a heavy chain CDR3 having the amino acid sequence of SEQ ID NO. 4, a light chain CDR1 having the amino acid sequence of SEQ ID NO. 81, a light chain CDR2 having the amino acid sequence of SEQ ID NO. 82, and a light chain CDR3 having the amino acid sequence of SEQ ID NO. 83, or comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO. 84 and a light chain variable region having the amino acid sequence of SEQ ID NO. 85.

[0134] The anti-TCTP antibody AO2M13 comprises a heavy chain CDR1 having the amino acid sequence of SEQ ID NO. 2, a heavy chain CDR2 having the amino acid sequence of SEQ ID NO. 86, a heavy chain CDR3 having the amino acid sequence of SEQ ID NO. 4, a light chain CDR1 having the amino acid sequence of SEQ ID NO. 87, a light chain CDR2 having the amino acid sequence of SEQ ID NO. 88, and a light chain CDR3 having the amino acid sequence of SEQ ID NO. 89, or comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO. 90 and a light chain variable region having the amino acid sequence of SEQ ID NO. 91.

[0135] The anti-TCTP antibody AO2M14 comprises a heavy chain CDR1 having the amino acid sequence of SEQ ID NO. 2, a heavy chain CDR2 having the amino acid sequence of SEQ ID NO. 92, a heavy chain CDR3 having the amino acid sequence of SEQ ID NO. 4, a light chain CDR1 having the amino acid sequence of SEQ ID NO. 93, a light chain CDR2 having the amino acid sequence of SEQ ID NO. 94, and a light chain CDR3 having the amino acid sequence of SEQ ID NO. 95, or comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO. 96 and a light chain variable region having the amino acid sequence of SEQ ID NO. 97.

[0136] The anti-TCTP antibody AO2M15 comprises a heavy chain CDR1 having the amino acid sequence of SEQ ID NO. 2, a heavy chain CDR2 having the amino acid sequence of SEQ ID NO. 98, a heavy chain CDR3 having the amino acid sequence of SEQ ID NO. 4, a light chain CDR1 having the amino acid sequence of SEQ ID NO. 99, a light chain CDR2 having the amino acid sequence of SEQ ID NO. 100, and a light chain CDR3 having the amino acid sequence of SEQ ID NO. 101, or comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO. 102 and a light chain variable region having the amino acid sequence of SEQ ID NO. 103.

[0137] The anti-TCTP antibody AO2M16 comprises a heavy chain CDR1 having the amino acid sequence of SEQ ID NO. 2, a heavy chain CDR2 having the amino acid sequence of SEQ ID NO. 104, a heavy chain CDR3 having the amino acid sequence of SEQ ID NO. 4, a light chain CDR1 having the amino acid sequence of SEQ ID NO. 105, a light chain CDR2 having the amino acid sequence of SEQ ID NO. 106, and a light chain CDR3 having the amino acid sequence of SEQ ID NO. 107, or comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO. 108 and a light chain variable region having the amino acid sequence of SEQ ID NO. 109.

[0138] The anti-TCTP antibody AO2M17 comprises a heavy chain CDR1 having the amino acid sequence of SEQ ID NO. 2, a heavy chain CDR2 having the amino acid sequence of SEQ ID NO. 110, a heavy chain CDR3 having the amino acid sequence of SEQ ID NO. 4, a light chain CDR1 having the amino acid sequence of SEQ ID NO. 111, a light chain CDR2 having the amino acid sequence of SEQ ID NO. 112, and a light chain CDR3 having the amino acid sequence of SEQ ID NO. 113, or comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO. 114 and a light chain variable region having the amino acid sequence of SEQ ID NO. 115.

[0139] The anti-TCTP antibody AO2M18 comprises a heavy chain CDR1 having the amino acid sequence of SEQ ID NO. 2, a heavy chain CDR2 having the amino acid sequence of SEQ ID NO. 116, a heavy chain CDR3 having the amino acid sequence of SEQ ID NO. 4, a light chain CDR1 having the amino acid sequence of SEQ ID NO. 117, a light chain CDR2 having the amino acid sequence of SEQ ID NO. 118, and a light chain CDR3 having the amino acid sequence of SEQ ID NO. 119, or comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO. 120 and a light chain variable region having the amino acid sequence of SEQ ID NO. 121.

[0140] The anti-TCTP antibody AO2M19 comprises a heavy chain CDR1 having the amino acid sequence of SEQ ID NO. 2, a heavy chain CDR2 having the amino acid sequence of SEQ ID NO. 122, a heavy chain CDR3 having the amino acid sequence of SEQ ID NO. 4, a light chain CDR1 having the amino acid sequence of SEQ ID NO. 123, a light chain CDR2 having the amino acid sequence of SEQ ID NO. 124, and a light chain CDR3 having the amino acid sequence of SEQ ID NO. 125, or comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO. 126 and a light chain variable region having the amino acid sequence of SEQ ID NO. 127.

[0141]

[0142] Example 2. TCTP binding activity of anti-TCTP antibody

[0143] The affinity of the human Fab antibodies of Example 1 for TCTP was confirmed. Specifically, using Biacore 8K (Cytiva), a culture medium containing Fab antibodies that bind to TCTP protein was immobilized on a sensor chip, and the binding rate constant (ka) and dissociation rate constant (kd) values ​​were measured using a single concentration of antigen TCTP as the analyte. Data for the dissociation (kd) and binding (ka) rate constants were obtained using Biacore 8K evaluation software, and the equilibrium dissociation constant (KD) was calculated from the ratio of kd to ka. The affinity confirmed by the single concentration SPR measurement method is shown in Table 2.

[0144] Antibody Name ka (1 / Ms)kd (1 / s)KD (M)AO2P 1.74 x 10 4 1.51 x 10 -3 8.67 x 10 -8 AO2M11.47 x 10 4 8.07 x 10 -4 5.49 x 10 -8 AO2M21.74 x 10 4 6.88 x 10 -4 3.96 x 10 -8 AO2M3 1.95 x 10 4 5.75 x 10 -4 2.94 x 10 -8 AO2M41.34 x 10 4 8.91 x 10 -4 6.66 x 10 -8 AO2M51.56 x 10 4 7.30 x 10 -4 4.68 x 10 -8 AO2M69.55 x 10 3 8.74 x 10 -4 9.15 x 10 -8 AO2M7 1.50 x 10 4 1.17 x 10 -3 7.77 x 10 -8AO2M81.45 x 10 4 1.13 x 10 -3 7.78 x 10 -8 AO2M91.13 x 10 4 1.41 x 10 -3 1.25 x 10 -7 AO2M101.57 x 10 4 9.42 x 10 -4 6.00 x 10 -8 AO2M111.56 x 10 4 1.26 x 10 -3 8.06 x 10 -8 AO2M121.51 x 10 4 1.44 x 10 -3 9.53 x 10 -8 AO2M131.68 x 10 4 1.03 x 10 -3 6.15 x 10 -8 AO2M141.46 x 10 4 9.00 x 10 -4 6.15 x 10 -8 AO2M151.58 x 10 4 9.97 x 10 -4 6.31 x 10 -8 AO2M161.55 x 10 4 1.43 x 10 -3 9.21 x 10 -8 AO2M171.33 x 10 4 1.22 x 10 -3 9.11 x 10 -8 AO2M181.45 x 10 4 1.16 x 10 -3 8.02 x 10 -8 AO2M191.64 x 10 4 1.12 x 10 -3 6.86 x 10 -8

[0145]

[0146] In addition, the inventors analyzed the TCTP binding affinity of the anti-TCTP antibody prepared according to Example 1 using a Biacore T200. TCTP was immobilized on the sensor chip, and the binding rate constant (ka) and dissociation rate constant (kd) values ​​were measured using the anti-TCTP antibody as the analyte. Data for the dissociation (kd) and binding (ka) rate constants were obtained using Biacore evaluation software, and the equilibrium dissociation constant (KD) was calculated from the ratio of kd to ka. The binding rate constant (ka) represents the complex formation rate, that is, the number of antigen-antibody complexes formed per second in a 1 molar solution of TCTP and the anti-TCTP antibody, and the unit of ka is 1 / Ms. The dissociation rate constant (kd) represents the stability of the complex, that is, the rate of complexes dissociating per second, and the unit of kd is 1 / s. For example, kd = 0.01 1 / s means that 1% of the complex dissociates per second. The equilibrium dissociation constant (KD) value calculated from the ratio of kd to ka represents the binding affinity of the antibody to the TCTP protein.

[0147] Antibody name ka (1 / Ms)kd (1 / s)KD (M)AO2P 1.54 x 10 6 6.93 x 10 -3 4.50 x 10 -9 AO2M12.20 x 10 6 1.09 x 10 -3 4.97 x 10 -10 AO2M21.53 x 10 6 6.30 x 10 -4 4.11 x 10 -10 AO2M31.38 x 10 6 8.57 x 10 -4 6.19 x 10 -10 AO2M42.06 x 10 6 1.51 x 10 -3 7.35 x 10 -10 AO2M51.78 x 10 6 7.45 x 10 -4 4.19 x 10 -10AO2M13 1.35 x 10 6 5.15 x 10 -3 3.81 x 10 -9

[0148] As shown in Tables 2 and 3 above, it was confirmed that the Fab antibody and anti-TCTP antibody according to the present invention have excellent binding affinity to TCTP protein.

[0149]

[0150] Example 3. Reduction of ERK phosphorylation (pERK) in cancer cells

[0151] To confirm the anticancer effect of the anti-TCTP antibody according to the present invention, the effect of reducing ERK activation (i.e., ERK phosphorylation) in cancer cells was analyzed. ERK is primarily activated by growth factors involved in cell proliferation, differentiation, and survival. ERK activation occurs through the phosphorylation of ERK proteins, and such sustained activation of ERK plays a crucial role in tumor formation. It is known that ERK activation is increased in various types of cancer, including pancreatic cancer, colorectal cancer, lung cancer, melanoma, ovarian cancer, kidney cancer, and breast cancer.

[0152] In this embodiment, colorectal cancer MC38 αPD-1 Cell line, colorectal cancer CT26 αPD-1 Cell line, melanoma B16F0 αPD-1 Cell line and melanoma B16F0 αPD-L1 A cell line was used. Here, colorectal cancer MC38 αPD-1 The cell line refers to a cell line refractory to PD-1 antibody therapy, constructed from the colorectal cancer MC38 cell line. Colorectal cancer CT26 αPD-1 The cell line refers to a cell line refractory to PD-1 antibody therapy, constructed from the colorectal cancer CT26 cell line. Melanoma B16F0 αPD-1 The cell line refers to a cell line refractory to PD-1 antibody treatment, constructed from the melanoma B16F0 cell line. Melanoma B16F0 αPD-L1The cell line refers to a cell line refractory to PD-L1 antibody treatment, constructed from the melanoma B16F0 cell line.

[0153] MC38 αPD-1 Cells and CT26 αPD-1 Cells 1.5 x 10⁶ per well in a 12-well plate 5 Seed with a cell count of 10, and B16F0 αPD-1 Cells and B16F0 αPD-L1 Cells 1.0 x 10⁶ per well in a 12-well plate 5 Cells were seeded at a rate of [number] cells. Medium containing 10% FBS was used for culture. After 16 hours, the medium was replaced with medium containing 0.1% FBS, and after an additional 6 hours of incubation, each anti-TCTP antibody was treated at a concentration of 10 or 50 ng / mL. After 24 hours, cells were washed with 1X PBS buffer and lysed by incubating in 150 µL of RIPA buffer at 4°C for 10 minutes. Subsequently, the cells were centrifuged at 13,000 rpm at 4°C for 15 minutes to obtain the supernatant cell extract. Proteins were quantified using the Bradford assay, and ERK protein levels and ERK protein phosphorylation levels were measured for equal amounts of protein using the SDS-PAGE Western blotting method. The measured ERK protein levels and ERK protein phosphorylation levels were quantified using Image J (densitometer), and the phosphorylated ERK protein levels were calculated as a ratio to total ERK protein.

[0154] The group without antibody treatment (indicated by "-") or the group treated with a non-specific IgG4 antibody (indicated by "Iso") was designated as the negative control group. For the comparison group, commercially available anti-TCTP antibodies were used, specifically the "Sino" antibody from Sino Biological (Cat. no. 14662-MM08), the "Abnova" antibody from Abnova (Cat. no. H00007178-M03), and the "Santacruz" antibody from Santa Cruz (Cat. no. sc-133131).

[0155] As a result, as can be seen in Figures 1a to 1d, ERK phosphorylation was significantly reduced in all cancer cell lines by the anti-TCTP antibody according to the present invention, and the ratio of ERK phosphorylation to total ERK was reduced by up to about 80% compared to the control group. On the other hand, when treated with the comparative anti-TCTP antibody (Sino, Abnova, Santacruz), there was no significant change in ERK phosphorylation in all cancer cell lines, or it even increased.

[0156] The above results suggest that the anti-TCTP antibody according to the present invention can reduce ERK activation in cancer cells, thereby exhibiting an excellent anticancer effect.

[0157]

[0158] Example 4. Increase in Apoptosis

[0159] To confirm the anticancer effect of the anti-TCTP antibody according to the present invention, the effect of increasing apoptosis in cancer cells was analyzed.

[0160] In this embodiment, lung cancer TC-1 αPD-1 Cell line, colorectal cancer MC38 αPD-1 Cell line, colorectal cancer MC38 αPD-L1 A cell line was used. Here, lung cancer TC-1 αPD-1The cell line refers to a cell line refractory to PD-1 antibody therapy, constructed from the lung cancer TC-1 cell line. Colorectal cancer MC38 αPD-1 The cell line refers to a cell line constructed from the colorectal cancer MC38 cell line that is refractory to PD-1 antibody therapy. Colorectal cancer MC38 αPD-L1 The cell line refers to a cell line refractory to PD-L1 antibody treatment, constructed from the colorectal cancer MC38 cell line.

[0161] Lung cancer TC-1 αPD-1 1.5 x 10 cells per well in a 12-well plate 5 Cells were seeded at a cell count, and a medium containing 10% FBS was used for culture. After 16 hours, the medium was replaced with one containing 0.1% FBS, and after an additional 6 hours of culture, each anti-TCTP antibody was treated at a concentration of 50 ng / mL. After 24 hours, the proportion of cells killed by apoptosis was determined using FACS.

[0162] Colorectal cancer MC38 αPD-1 Cell and Colorectal Cancer MC38 αPD-L1 5 x 10 cells per well in a 96-well plate 3 Cells were seeded at a cell count, and a medium containing 10% FBS was used for culture. After 16 hours, each anti-TCTP antibody was treated at a concentration of 50 ng / mL. After 8 hours, 50 μM cisplatin was added to induce apoptosis. After two days, the percentage of dead cells was determined using WST-8.

[0163] A sample treated with the non-specific antibody IgG4 (indicated as "IgG") was set as the negative control. For the comparison group, commercially available anti-TCTP antibodies were used, specifically the "Sino" antibody from Sino Biological (Cat. no. 14662-MM08), the "Abnova" antibody from Abnova (Cat. no. H00007178-M03), and the "Santacruz" antibody from Santa Cruz (Cat. no. sc-133131).

[0164] As a result, as can be seen in Figures 2a to 2c, the proportion of cells killed by the anti-TCTP antibody according to the present invention increased significantly in all cancer cell lines compared to the negative control group. On the other hand, when treated with the comparative anti-TCTP antibody (Sino, Abnova, Santacruz), there was no significant change in the proportion of cells killed in all cancer cell lines, or it decreased or increased to a low level.

[0165] The above results suggest that the anti-TCTP antibody according to the present invention can increase the sensitivity to apoptosis in cancer cells, thereby exhibiting an excellent anticancer effect.

[0166]

[0167] Example 5. In vivo inhibition of tumor growth by anti-TCTP antibody

[0168] To confirm the anticancer effect of the anti-TCTP antibody according to the present invention, the degree of tumor growth was analyzed after administering the antibody of the present invention to mice.

[0169] In this embodiment, colorectal cancer CT26 cell line, melanoma B16F0 cell line, colorectal cancer MC38 cell line, lung cancer TC-1 αPD-1 Cell lines and colorectal cancer MC38 αPD-1 A cell line was used. Here, lung cancer TC-1 αPD-1The cell line refers to a cell line refractory to PD-1 antibody therapy, constructed from the lung cancer TC-1 cell line. Colorectal cancer MC38 αPD-1 The cell line refers to a cell line refractory to PD-1 antibody treatment, constructed from the colorectal cancer MC38 cell line.

[0170] 1–3 x 10 cells per balb / c mouse or C57BL / 6 mouse 5 Tumors were formed by transplantation with a cell count. The average tumor size was approximately 10 to 40 mm. 3 When [the time] was reached, mice were distributed among the groups so that the tumor size was distributed as uniformly as possible (3 to 6 mice per experimental group), and antibodies were administered. The antibody dosage was 10 mg / kg, and it was administered via intraperitoneal injection three times a week for a total of five times, starting from the antibody administration start date (Day 0).

[0171] Subsequently, the length of the tumor was measured using calipers at intervals of 2 or 3 days starting from the date of antibody administration, and the tumor size was calculated using the following formula: Tumor size = {Length of major axis x (Length of minor axis) 2 2. The above results were analyzed using parametric multiple comparison procedures, assuming normality of the data. If the results of the two-way ANOVA were significant, post-hoc testing was performed using Sidak's multiple comparison test. Statistical analysis was performed using Prism 8.0.1 (GraphPad Software Inc., San Diego, CA, USA), and a P-value of less than 0.05 was considered statistically significant.

[0172] The sample treated with a non-specific antibody (designated as "Control IgG") was set as the negative control. The comparison group used commercially available anti-TCTP antibodies, specifically the "Sino" antibody from Sino Biological (Cat. no. 14662-MM08), the "Abnova" antibody from Abnova (Cat. no. H00007178-M03), and the "Santacruz" antibody from Santa Cruz (Cat. no. sc-133131).

[0173] As a result, as shown in FIGS. 3a to 3e, the anti-TCTP antibody according to the present invention can significantly inhibit tumor growth in vivo. Furthermore, this effect was observed not only in cancer cells responsive to existing immunotherapies but also in cancer cells that are refractory. On the other hand, when the control group anti-TCTP antibody (Sino, Abnova, Santacruz) was administered, no significant change in the inhibition of tumor growth in vivo was observed.

[0174] The results of Example 5 above indicate that the anti-TCTP antibody according to the present invention can inhibit tumor growth in vivo and exhibit an excellent anticancer effect, and in particular, exhibits an excellent anticancer effect even against cancers that are resistant or refractory to existing anticancer drugs. Furthermore, it indicates that the anti-TCTP antibody according to the present invention not only exhibits an anticancer effect alone but can also exhibit an excellent synergistic effect together with existing immunotherapies.

[0175]

[0176] Example 6. In vivo inhibition of tumor growth with anti-TCTP antibodies alone and in combination

[0177] To confirm the anticancer effects of the anti-TCTP antibody according to the present invention when used alone and in combination, the degree of tumor growth was analyzed after administering the antibody of the present invention to mice.

[0178] In this embodiment, colorectal cancer CT26αPD-1 Cell line, colorectal cancer SL4 αPD-1 Cell line, colorectal cancer MC38 αPD-1 Cell line, Colon cancer Colon 26 Cell line, melanoma B16BL6 Cell line, breast cancer 4T1 Cell lines, hepatocellular carcinoma RIL-175 cell line, colorectal cancer SW480 cell line, triple-negative breast cancer MDA-MB-231 cell line, lung adenocarcinoma A549 A cell line was used. Here, colorectal cancer CT26 αPD-1 The cell line refers to a cell line refractory to PD-1 antibody treatment constructed from the colorectal cancer CT26 cell line, and colorectal cancer SL4 αPD-1 The cell line refers to a cell line refractory to PD-1 antibody treatment constructed from colorectal cancer SL4 cell lines, and colorectal cancer MC38 αPD-1 The cell line refers to a cell line refractory to PD-1 antibody treatment constructed from the colorectal cancer MC38 cell line, and colorectal cancer Colon 26 Cell lines refer to cell lines known to be resistant to immunotherapies such as PD-1 antibodies and PD-L1 antibodies, and melanoma B16BL6 A cell line refers to a cell line known to be resistant to immunotherapies such as PD-1 antibodies and PD-L1 antibodies. In addition, breast cancer 4T1 Cell lines refer to cell lines known to be resistant to immunotherapies such as PD-1 antibodies and PD-L1 antibodies; the colorectal cancer SW480 cell line refers to a cell line that exhibits refractoryness to anti-PD-1 antibody treatment; the triple-negative breast cancer MDA-MB-231 cell line refers to a cell line that exhibits refractoryness to anti-PD-1 antibody treatment; and lung adenocarcinoma A549 A cell line refers to an epithelial cell line that is refractory to anti-PD-1 antibody treatment.

[0179]

[0180] Example 6-1.

[0181] In this Example 6-1, colorectal cancer CT26 αPD-1 A cell line was used.

[0182] Colorectal cancer CT26 P3 in BALB / c mice αPD-1 2 x 10 cells 5 Tumors were formed by transplanting with a cell count. When the tumor size reached an average of approximately 20 to 30 mm³, mice were distributed among the groups to ensure the tumor size was as uniform as possible (13 mice per experimental group), and antibodies were administered. The dose of the anti-TCTP antibody was 10 mg / kg, and it was administered via intraperitoneal injection three times a week for a total of six times, starting from the antibody administration start date (Day 0). The anti-PD-1 antibody was administered at 5 mg / kg, and it was administered via intraperitoneal injection twice a week for a total of four times, starting from the antibody administration start date (Day 0).

[0183] Subsequently, the length of the tumor was measured using calipers at intervals of 2 or 3 days starting from the date of antibody administration, and the tumor size was calculated using the following formula: Tumor size = {Length of major axis x (Length of minor axis) 2} / 2. In addition, the tumor growth inhibition rate (TGI, %) was calculated using the above tumor size according to the following formula: 100 - (100 x experimental group tumor size / IgG antibody-treated control group tumor size).

[0184] The above results were analyzed using parametric multiple comparison procedures, assuming the normality of the data. If the results of the two-way ANOVA were significant, post-hoc testing was performed using Sidak's multiple comparison test. Statistical analysis was conducted using Prism 8.0.1 (GraphPad Software Inc., San Diego, CA, USA), and a P-value of less than 0.05 was determined to be statistically significant. In the graphs of the figure, *: p < 0.05, **: p < 0.01, ***: p < 0.001, and ****: p < 0.0001.

[0185] As a result, as shown in Figure 4, on the 14th day from the antibody administration start date (Day 0), the negative control group treated with IgG antibodies (indicated as "Control IgG") had an average tumor size of 987 mm³, while the experimental group treated with the anti-TCTP antibody according to the present invention had an average tumor size of 533 mm³. The experimental group treated with the anti-TCTP antibody according to the present invention showed a tumor growth inhibition rate (TGI) of 46% compared to the negative control group treated with IgG antibodies, indicating that the anti-TCTP antibody according to the present invention significantly inhibits tumor growth even when administered alone. Furthermore, the group administered the anti-TCTP antibody according to the present invention in combination with an anti-PD-1 antibody showed an average tumor size of 312 mm³ and a tumor growth inhibition rate of 68%.

[0186]

[0187] Example 6-2.

[0188] In this Example 6-2, colorectal cancer SL4 αPD-1 A cell line was used.

[0189] Colorectal cancer SL4 in C57BL / 6N mice αPD-15 x 10 cells 5 Tumors were formed by transplanting with a cell count. When the tumor size reached an average of approximately 10 to 30 mm³, mice were distributed among the groups to ensure the tumor sizes were as uniform as possible (11 mice per experimental group).

[0190] Subsequently, as described in Example 6-1, the length and size of the tumor and the tumor growth inhibition rate were calculated after administering the antibody.

[0191] As a result, as shown in Fig. 5, on the 14th day from the antibody administration start date (Day 0), the negative control group treated with IgG antibodies (indicated as "Control IgG") had an average tumor size of 526 mm³, while the experimental group treated with the anti-TCTP antibody according to the present invention had an average tumor size of 355 mm³. The experimental group treated with the anti-TCTP antibody according to the present invention showed a tumor growth inhibition rate (TGI) of 33% compared to the negative control group treated with IgG antibodies, indicating that the anti-TCTP antibody according to the present invention significantly inhibits tumor growth even when administered alone. Furthermore, the group administered the anti-TCTP antibody according to the present invention in combination with an anti-PD-1 antibody showed an average tumor size of 223 mm³ and a tumor growth inhibition rate of 58%.

[0192]

[0193] Example 6-3.

[0194] In this Example 6-3, colorectal cancer MC38 αPD-1 A cell line was used.

[0195] Colorectal cancer MC38 in C57BL / 6N mice αPD-1 5 x 10 cells 5Tumors were formed by transplanting with a cell count. When the tumor size reached an average of approximately 10 to 30 mm³, mice were distributed among the groups to ensure the tumor size was as uniform as possible (11 mice per experimental group), and antibodies were administered. The dose of the anti-TCTP antibody was 10 mg / kg, and it was administered via intraperitoneal injection three times a week for a total of five times, starting from the antibody administration start date (Day 0). The anti-PD-1 antibody was administered at a dose of 5 mg / kg, and it was administered via intraperitoneal injection twice a week for a total of four times, starting from the antibody administration start date (Day 0).

[0196] Subsequently, as described in Example 6-1, the length and size of the tumor and the tumor growth inhibition rate were calculated.

[0197] As a result, as shown in Fig. 6, on the 12th day from the antibody administration start date (Day 0), the negative control group treated with IgG antibodies (indicated as "Control IgG") had an average tumor size of 1,477 mm³, while the experimental group treated with the anti-TCTP antibody according to the present invention had an average tumor size of 803 mm³. The experimental group treated with the anti-TCTP antibody according to the present invention showed a tumor growth inhibition rate of 45% compared to the negative control group treated with IgG antibodies, indicating that the anti-TCTP antibody according to the present invention significantly inhibits tumor growth even when administered alone. Furthermore, the group administered the anti-TCTP antibody according to the present invention in combination with an anti-PD-1 antibody showed an average tumor size of 557 mm³ and a tumor growth inhibition rate of 62%.

[0198]

[0199] Example 6-4.

[0200] In this Example 6-4, colon cancer Colon 26 A cell line was used.

[0201] Colon cancer in BALB / c mice Colon 26 3 x 10 cells 5Tumors were formed by transplanting the cells. When the tumor size reached an average of about 10 to 30 mm³, mice were distributed among the groups so that the tumor sizes were distributed as uniformly as possible (10 mice per experimental group). Subsequently, as described in Example 6-1, the length and size of the tumors and the tumor growth inhibition rate were calculated after administering the antibody.

[0202] As a result, as shown in Fig. 7, on the 14th day from the antibody administration start date (Day 0), the negative control group treated with IgG antibodies (indicated as "Control IgG") had an average tumor size of 735 mm³, while the experimental group treated with the anti-TCTP antibody according to the present invention had an average tumor size of 452 mm³. The experimental group treated with the anti-TCTP antibody according to the present invention showed a tumor growth inhibition rate (TGI) of 39% compared to the negative control group treated with IgG antibodies, indicating that the anti-TCTP antibody according to the present invention significantly inhibits tumor growth even when administered alone. Furthermore, the group administered the anti-TCTP antibody according to the present invention in combination with an anti-PD-1 antibody showed an average tumor size of 296 mm³ and a tumor growth inhibition rate of 60%.

[0203]

[0204] Example 6-5.

[0205] Examples 6-5 of this document are colon cancer Colon 26 The procedure was carried out in the same manner as in Example 6-4 using cell lines, but with the difference that 9 mice were used per experimental group, anti-TCTP antibodies were injected intraperitoneally 8 times in total, 3 times a week starting from the antibody administration start date (Day 0), and anti-PD-L1 antibodies were injected intraperitoneally 6 times in total, 2 times a week starting from the antibody administration start date (Day 0).

[0206] Subsequently, as described in Example 6-1, the length and size of the tumor and the tumor growth inhibition rate (TGI) were calculated.

[0207] As a result, as shown in Fig. 8, on the 18th day from the antibody administration start date (Day 0), the negative control group treated with IgG antibodies (indicated as "Control IgG") had an average tumor size of 1,289 mm³, while the experimental group treated with the anti-TCTP antibody according to the present invention had an average tumor size of 722 mm³. The experimental group treated with the anti-TCTP antibody according to the present invention showed a tumor growth inhibition rate (TGI) of 43% compared to the negative control group treated with IgG antibodies, indicating that the anti-TCTP antibody according to the present invention significantly inhibits tumor growth even when administered alone. Furthermore, the group administered the anti-TCTP antibody according to the present invention in combination with the anti-PD-L1 antibody showed an average tumor size of 557 mm³ and a tumor growth inhibition rate of 57%.

[0208]

[0209] Example 6-6.

[0210] In this Example 6-6, melanoma B16BL6 A cell line was used.

[0211] B16BL6 Melanoma in C57BL / 6N Mice 3 x 10 cells 5 Tumors were formed by transplanting with a cell count. When the tumor size reached an average of approximately 20 to 40 mm³, mice were distributed among the groups to ensure the tumor size was as uniform as possible (12 mice per experimental group), and antibodies were administered. The dose of the anti-TCTP antibody was 10 mg / kg, and it was administered via intraperitoneal injection three times a week for a total of five times, starting from the antibody administration start date (Day 0). The anti-PD-1 antibody was administered at a dose of 5 mg / kg, and it was administered via intraperitoneal injection twice a week for a total of four times, starting from the antibody administration start date (Day 0).

[0212] Subsequently, as described in Example 6-1, the length and size of the tumor and the tumor growth inhibition rate (TGI) were calculated.

[0213] As a result, as shown in Fig. 9, on the 12th day from the antibody administration start date (Day 0), the negative control group treated with IgG antibodies (indicated as "Control IgG") had an average tumor size of 906 mm³, while the experimental group treated with the anti-TCTP antibody according to the present invention had an average tumor size of 503 mm³. The experimental group treated with the anti-TCTP antibody according to the present invention showed a tumor growth inhibition rate (TGI) of 45% compared to the negative control group treated with IgG antibodies, indicating that the anti-TCTP antibody according to the present invention significantly inhibits tumor growth even when administered alone. Furthermore, the group administered the anti-TCTP antibody according to the present invention in combination with an anti-PD-1 antibody showed an average tumor size of 293 mm³ and a tumor growth inhibition rate of 68%.

[0214]

[0215] Examples 6-7.

[0216] In Examples 6-7, breast cancer 4T1 A cell line was used.

[0217] C57BL / 6N mice 4T1 breast cancer 3 x 10 cells 5 Tumors were formed by transplanting with a cell count. When the tumor size reached an average of approximately 20 to 30 mm³, mice were distributed among the groups to ensure the tumor size was as uniform as possible (9 mice per experimental group), and antibodies were administered. The dose of the anti-TCTP antibody was 10 mg / kg, and it was administered via intraperitoneal injection three times a week for a total of eight times, starting from the antibody administration start date (Day 0). The anti-PD-1 antibody was administered at a dose of 5 mg / kg, and it was administered via intraperitoneal injection twice a week for a total of six times, starting from the antibody administration start date (Day 0).

[0218] Subsequently, as described in Example 6-1, the length and size of the tumor and the tumor growth inhibition rate (TGI) were calculated.

[0219] As a result, as shown in Fig. 10, on the 18th day from the antibody administration start date (Day 0), the negative control group treated with IgG antibodies (indicated as "Control IgG") had an average tumor size of 1,174 mm³, while the experimental group treated with the anti-TCTP antibody according to the present invention had an average tumor size of 681 mm³. The experimental group treated with the anti-TCTP antibody according to the present invention showed a tumor growth inhibition rate (TGI) of 42% compared to the negative control group treated with IgG antibodies, indicating that the anti-TCTP antibody according to the present invention significantly inhibits tumor growth even when administered alone. Furthermore, the group administered the anti-TCTP antibody according to the present invention in combination with an anti-PD-1 antibody showed an average tumor size of 406 mm³ and a tumor growth inhibition rate of 65%.

[0220]

[0221] Examples 6-8.

[0222] In Examples 6-8, hepatocellular carcinoma RIL-175 A cell line was used.

[0223] RIL-175 hepatocellular carcinoma in C57BL / 6N mice 1 x 10 cells 5 Tumors were formed by transplanting with a cell count. When the tumor size reached an average of approximately 20 to 40 mm³, mice were distributed among the groups to ensure the tumor size was as uniform as possible (8 mice per experimental group), and then antibodies were administered. The dosage of the anti-TCTP antibody was 10 mg / kg, and it was administered via intraperitoneal injection three times a week for a total of 10 times, starting from the antibody administration start date (Day 0).

[0224] Subsequently, as described in Example 6-1, the length and size of the tumor and the tumor growth inhibition rate (TGI) were calculated.

[0225] As a result, as shown in Fig. 11, based on the antibody administration start date (Day 0), the negative control group treated with IgG antibodies (indicated as "Control IgG") had an average tumor size of 885.27 mm³, while the experimental group treated with the anti-TCTP antibody according to the present invention (indicated as "AO2M5") had an average tumor size of 330.29 mm³. The tumor growth inhibition rate of the experimental group treated with the anti-TCTP antibody according to the present invention was 62.69% compared to the negative control group treated with IgG antibodies, indicating that the anti-TCTP antibody according to the present invention significantly inhibits tumor growth.

[0226]

[0227] Examples 6-9.

[0228] In Examples 6-9, colorectal cancer SW480 NCG mice transplanted with cell lines and human PBMCs (peripheral blood mononuclear cells) were used.

[0229] NCG mice are triple immunodeficiency mice in which major immune cells such as T cells, B cells, and NK cells are almost completely eliminated, and are known as an immunodeficient mouse model in which immune rejection is minimized when human cells are transplanted.

[0230] Specifically, colorectal cancer SW480 in NCG mice 5 x 10 cells 5 Tumors were formed by transplanting with a cell count of [number]. When the tumor size reached an average of approximately 100 mm³, mice were distributed among the groups to ensure the tumor size was as uniform as possible (7 mice per experimental group), and then 5 x 10 human PBMCs (CD45+ cells) were introduced. 6The antibodies were administered intravenously with the number of cells. Three hours after administering human PBMC, each antibody was administered. The dosage of the anti-TCTP antibody, anti-PD-1 antibody, and anti-VEGF antibody was 200 µg / mouse, and they were intravenously administered once a week for a total of three times, starting from the antibody administration start date (Day 0). The anti-PD-1 antibody was pembrolizumab, and the anti-VEGF antibody was bevacizumab, both of which are anticancer drugs approved by the US FDA.

[0231] Subsequently, the length and size of the tumor and the tumor growth inhibition rate (TGI) were calculated twice a week at intervals of 3 or 4 days using the same method as described in Example 6-1.

[0232] As a result, as shown in Figure 12, based on the antibody administration start date (Day 0), the average tumor size of Control 1 (indicated as "Control IgG") treated with IgG antibodies was 603.5 mm³. Control 2 (indicated as "αPD-1") treated with anti-PD-1 antibodies had an average tumor size of 581.4 mm³ and showed a tumor growth rate similar to that of Control 1 treated with IgG antibodies. Control 3 (indicated as "αVEGF") treated with anti-VEGF antibodies had an average tumor size of 491.1 mm³ and showed a tumor growth inhibition rate of 18.6% compared to Control 1 treated with IgG antibodies. In the experimental group treated with the anti-TCTP antibody according to the present invention (indicated as "AO2M5"), the average tumor size was 406.0 mm³, and the tumor growth inhibition rate was 33% compared to control group 1 treated with the IgG antibody, indicating that the anti-TCTP antibody according to the present invention significantly inhibits tumor growth. In addition, it can be seen that the anti-TCTP antibody according to the present invention exhibits a higher in vivo tumor inhibition effect than the anti-PD-1 antibody and the anti-VEGF antibody.

[0233]

[0234] Examples 6-10.

[0235] Examples 6-10 of this document describe triple-negative breast cancer MDA-MB-231 The procedure was carried out in the same manner as in Examples 6-9 using cell lines, except that anti-TCTP antibody, anti-PD-1 antibody, and anti-VEGF antibody were administered at 100 μg / mouse each.

[0236] Subsequently, as described in Example 6-1, the length and size of the tumor and the tumor growth inhibition rate were calculated.

[0237] As a result, as shown in Figure 13, based on the antibody administration start date (Day 0), the average tumor size of Control 1 (indicated as "Control IgG") treated with IgG antibodies was 536.3 mm³. Control 2 (indicated as "αPD-1") treated with anti-PD-1 antibodies had an average tumor size of 509.9 mm³ and showed a tumor growth rate similar to that of Control 1 treated with IgG antibodies. Control 3 (indicated as "αVEGF") treated with anti-VEGF antibodies had an average tumor size of 345.1 mm³ and showed a tumor growth inhibition rate of 36% compared to Control 1 treated with IgG antibodies. In the experimental group treated with the anti-TCTP antibody according to the present invention (indicated as "AO2M5"), the average tumor size was 265.2 mm³, and the tumor growth inhibition rate was 51% compared to control group 1 treated with the IgG antibody, indicating that the anti-TCTP antibody according to the present invention significantly inhibits tumor growth. In addition, it can be seen that the anti-TCTP antibody according to the present invention exhibits a higher in vivo tumor inhibition effect than the anti-PD-1 antibody and the anti-VEGF antibody.

[0238]

[0239] Examples 6-11.

[0240] Examples 6-11 of this document are lung adenocarcinoma A549 The procedure was carried out in the same manner as in Examples 6-9 using a cell line.

[0241] Subsequently, as described in Example 6-1, the length and size of the tumor and the tumor growth inhibition rate were calculated.

[0242] As a result, as shown in Figure 14, based on the antibody administration start date (Day 0), the average tumor size of Control 1 (indicated as "Control IgG") treated with IgG antibodies was 522.8 mm³. Control 2 (indicated as "αPD-1") treated with anti-PD-1 antibodies had an average tumor size of 511.7 mm³ and showed a tumor growth rate similar to that of Control 1 treated with IgG antibodies. Control 3 (indicated as "αVEGF") treated with anti-VEGF antibodies had an average tumor size of 345.3 mm³ and showed a tumor growth inhibition rate of 34% compared to Control 1 treated with IgG antibodies. In the experimental group treated with the anti-TCTP antibody according to the present invention (indicated as "AO2M5"), the average tumor size was 362.2 mm³, and the tumor growth inhibition rate was 30% compared to control group 1 treated with the IgG antibody, indicating that the anti-TCTP antibody according to the present invention significantly inhibits tumor growth. In addition, it can be seen that the anti-TCTP antibody according to the present invention exhibits an in vivo tumor inhibitory effect similar to that of the anti-VEGF antibody and a higher in vivo tumor inhibitory effect than that of the anti-PD-1 antibody.

[0243]

[0244] The results of Examples 6-1 to 6-11 above indicate that the anti-TCTP antibody according to the present invention can inhibit tumor growth in vivo and exhibit an excellent anticancer effect, and in particular, exhibits an excellent anticancer effect even against cancers that are resistant or refractory to existing anticancer drugs. Furthermore, it indicates that the anti-TCTP antibody according to the present invention not only exhibits an anticancer effect alone but can also exhibit an excellent synergistic effect together with existing immunotherapies.

[0245]

[0246] Example 7. Inhibition of tumor growth in orthotopic transplantation

[0247] In this embodiment, an orthotopic tumor mouse model was constructed. In orthotopic tumor models, the colon is a tissue with inherent immunosuppressive properties, and it is known that signals inhibiting the activation of immune cells such as T cells act strongly to limit the effects of immune checkpoint inhibitors (William W. Ho, et al., Dendritic cell paucity in mismatch repair-proficient colorectal cancer liver metastases limits immune checkpoint blockade efficacy", PNAS 2021 Vol. 118 No. 45 e2105323118).

[0248] Specifically, the colorectal cancer cell line SL4-Luc, transformed to express luciferase, was prepared. C57BL / 6N mice were anesthetized, the abdominal cavity was opened, and cancer cells were directly implanted into the cecum at a rate of 5 x 10⁶ 4 Tumors were formed by transplanting with a cell count. Three days after cancer cell transplantation, 750 µg of luciferin was administered, and five minutes later, the average luminescence value measured using an In Vitro Luminescence Imaging Analyzer (IVIS) was approximately 1,000,000 (p / s / cm²). 2 When the sr level was reached, antibodies were administered (12 mice per experimental group). The anti-TCTP antibody according to the present invention was AO2M5, and the negative control group used an IgG antibody. The antibody dosage was 200 μg, and it was administered via intraperitoneal injection three times a week for a total of six times, starting from the antibody administration start date (Day 0).

[0249] Subsequently, luminescence was captured at intervals of 3 or 4 days to calculate the size of the tumor in vivo (ROI). As a result, as shown in Figures 15 and 16, the average region of interest (ROI) measured in the group administered the IgG antibody on day 22, relative to the antibody administration start date (Day 0), was 2.11 x 10⁻⁶. 8 (p / s / cm 2 It was calculated as / sr), and the mean ROI value in the group administered anti-TCTP antibodies was 0.959 x 10⁻⁶ 8 (p / s / cm 2 The tumor growth inhibition rate was calculated as / sr) and was found to be 55%. The above results suggest that the anti-TCTP antibody according to the present invention can exhibit an excellent anticancer effect in inhibiting the growth of tumors present in vivo in an orthotopic transplantation model that reproduces the major site of tumor formation, thereby indicating a therapeutic effect in patients.

[0250]

[0251] Example 8. Inhibition of TCTP and TLR2 binding by anti-TCTP antibody

[0252] In this embodiment, the inhibitory effect of the anti-TCTP antibody according to the present invention on the interaction between TCTP and TLR2 (toll-like receptor 2) was confirmed. It is already known in the art that the interaction between TCTP and TLR2 plays an important role in tumor growth in the tumor microenvironment.

[0253] Specifically, recombinant TCTP was immobilized on an immunoplate and reacted with various concentrations of biotinized TLR2 (biotin-TLR2). The interaction between TCTP and biotin-TLR2 was quantitatively evaluated using Avidin-HRP to detect biotin-TLR2 bound to TCTP.

[0254] As a result, as shown in Fig. 17, a concentration-dependent decrease in the OD450 value was observed in the anti-TCTP antibody treatment group according to the present invention, which indicates that the anti-TCTP antibody effectively inhibits the binding between TCTP and TLR2.

[0255] In addition, quantitative analysis was performed to evaluate the extent to which the anti-TCTP antibody inhibits the interaction between TCTP and TLR2, and the half-maximal inhibitory concentration IC₀ 50 The value was measured at 4.35 nM. Such a low IC 50 The value indicates that the anti-TCTP antibody according to the present invention has a high binding affinity for TCTP, and thus has high efficacy in inhibiting TCTP-TLR2 interaction.

Claims

An antibody that specifically binds to TCTP (translationally-controlled tumor protein) or an antigen-binding fragment thereof, comprising the following: Heavy chain CDR1 containing the amino acid sequence of SEQ ID NO. 2; Heavy chain CDR2 containing the amino acid sequence of SEQ ID NO. 3; Heavy chain CDR3 containing the amino acid sequence of SEQ ID NO. 4; Light chain CDR1 containing the amino acid sequence of SEQ ID NO. 5; Light chain CDR2 comprising the amino acid sequence of SEQ ID NO. 6; and Light chain CDR3 containing the amino acid sequence of SEQ ID NO.

7. A nucleic acid encoding the antibody or antigen-binding fragment of claim 1. A vector containing the nucleic acid of paragraph 2. Cells transformed with the vector of paragraph 3. A pharmaceutical composition for the prevention or treatment of cancer, comprising the antibody of claim 1 or an antigen-binding fragment thereof. In paragraph 5, A pharmaceutical composition wherein the above cancer is selected from the group consisting of bone cancer, lung cancer, non-small cell lung cancer, head cancer, neck cancer, thyroid cancer, parathyroid cancer, stomach cancer, liver cancer, pancreatic cancer, skin cancer, melanoma, small intestine cancer, colorectal cancer, rectal cancer, pro-anal cancer, colon cancer, uterine cancer, breast cancer, ovarian cancer, fallopian tube cancer, endometrial cancer, cervical cancer, vaginal cancer, vulvar cancer, Hodgkin's disease, esophageal cancer, endocrine gland cancer, adrenal cancer, soft tissue sarcoma, urethral cancer, penile cancer, prostate cancer, bladder cancer, kidney cancer, ureteral cancer, renal cell carcinoma, renal pelvic carcinoma, central nervous system tumor, central nervous system lymphoma, spinal cord tumor, glioblastoma, brainstem glioma and pituitary adenoma. In paragraph 5, A pharmaceutical composition in which the above cancer is resistant, resistant, or refractory to anticancer drugs. In Paragraph 7, The above anticancer agent is a pharmaceutical composition comprising one or more selected from the group consisting of chemotherapy agents, targeted anticancer agents, and immunotherapy agents. A cancer diagnostic composition comprising the antibody of claim 1 or an antigen-binding fragment thereof. A cancer diagnostic kit comprising the antibody of claim 1 or an antigen-binding fragment thereof. A method for providing information for diagnosing cancer in a subject, comprising the step of detecting or quantifying the expression or activity level of a TCTP protein in a biological sample isolated from a subject suspected of having cancer, using the antibody of claim 1 or an antigen-binding fragment thereof. A method for providing information for selecting a cancer treatment subject, comprising the step of detecting or quantifying the expression or activity level of a TCTP protein in a biological sample separated from a cancer treatment subject using the antibody of claim 1 or an antigen-binding fragment thereof. A method for providing information for predicting anticancer drug resistance, resistance, or refractoryness of a subject, comprising the step of detecting or quantifying the expression or activity level of a TCTP protein in a biological sample isolated from a subject suspected of exhibiting resistance, resistance, or refractoryness to an anticancer drug, using the antibody of claim 1 or an antigen-binding fragment thereof. A screening method for anticancer drugs comprising the following steps: (a) A step of treating cancer cells with a candidate anticancer drug; (b) a step of measuring the expression or activity level of TCTP protein using the antibody of claim 1 or its antigen-binding fragment in cancer cells treated with the above anticancer drug candidate; and (c) A step of determining the anticancer drug candidate treated in step (a) as an anticancer drug if the expression or activity level of the TCTP protein in step (b) above is lower than that of the negative control group. As an antibody that specifically binds to TCTP or an antigen-binding fragment thereof for use in the treatment or prevention of cancer, Heavy chain CDR1 containing the amino acid sequence of SEQ ID NO. 2; Heavy chain CDR2 containing the amino acid sequence of SEQ ID NO. 3; Heavy chain CDR3 containing the amino acid sequence of SEQ ID NO. 4; Light chain CDR1 containing the amino acid sequence of SEQ ID NO. 5; Light chain CDR2 comprising the amino acid sequence of SEQ ID NO. 6; and A light chain CDR3 comprising the amino acid sequence of SEQ ID NO. 7; Antibody or its antigen-binding fragment.

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