Antibodies against mucin 17 and uses thereof

Abstract

The present invention relates to binding antibodies targeting Mucin 17 (MUC17). The present invention further relates to detection systems comprising such antibodies for the diagnosis of diseases and / or disorders associated with aberrant expression of MUC17.

Description

Technical Field

[0001] This invention relates to binding antibodies targeting mucin 17 (Mucin-17, MUC17), and detection systems comprising such antibodies for the diagnosis of diseases and / or conditions associated with abnormal expression of Mucin 17. Background Technology

[0002] Mucins (MUCs) are high-molecular-weight glycoproteins produced and secreted by epithelial cells, forming the main component of mucus. To date, more than twenty types of mucin proteins have been identified, which are classified into two categories based on their function: membrane-bound mucins and secretory mucins. Secretory mucins can form a physical barrier, acting as a mucus gel to protect respiratory and gastrointestinal epithelial cells. Transmembrane mucins form rod-like structures through O-glycosylated tandem repeats in their extracellular domains, constructing a protective mucus gel layer.

[0003] MUC17 (Mucin-17) is a transmembrane mucin encoded by the MUC17 gene located at locus q22 on chromosome 7. Discovered and identified in 2002, MUC17 consists of three parts: a transmembrane segment, an intracellular segment, and an extracellular segment. From the N-terminus, the sequence is as follows: a mucin-like structure (tandem repeat sequence), an EGF (epidermal growth factor)-like region, an N-glycosylated region, a second EGF-like region, a hydrophobic transmembrane segment, and a carboxyl terminus in the cytoplasm. MUC17 binds to the cell membrane via the transmembrane region near its C-terminus and participates in intracellular signal transduction. The extracellular segment determines the MUC-specific spatial structure and immunogenicity. Expression of MUC17 can enhance the intestinal mucus barrier and promote inflammatory healing. In epithelial cells, MUC17 participates in signal transduction, maintains luminal structure, provides cell protection, and imparts anti-adhesion properties to cancer cells that have lost polarity, thereby causing tumor cells to lose their polarity.

[0004] MUC17 is primarily expressed in the gut. Studies have shown that MUC17 expression gradually increases significantly during the development of gastric mucosa, intestinal metaplasia mucosa, and advanced gastric cancer, and low expression is associated with poor prognosis in gastric cancer. Further research indicates that MUC17 expression is negatively correlated with tumor differentiation, suggesting that MUC17 could serve as a diagnostic and therapeutic target for gastric cancer.

[0005] Immunohistochemistry (IHC) is a routine technique used in medical diagnostics to determine the presence and expression levels of proteins in tissues. Tumor tissues are typically fixed in formalin and embedded in paraffin before IHC analysis. MUC17 diagnostic antibodies should be able to detect MUC17 expressed in tissue samples. However, therapeutic antibodies that typically target MUC17 are not effective at detecting endogenous MUC17 in tumor tissues. Therefore, there remains a need to develop antibodies that can detect and quantify endogenously expressed MUC17. Summary of the Invention

[0006] This invention provides a series of antibodies that specifically bind to MUC17, which can be used in diagnostic methods on tissue samples obtained from individuals, particularly from patients suspected of having neoplastic diseases associated with and / or identifiable by aberrant MUC17 expression. The anti-MUC17 antibodies of this invention can specifically recognize cells, tissues (e.g., cancerous tissue) or PDX that aberrantly express MUC17, without recognizing normal cells or tissues. Therefore, they can be used as highly sensitive detection and diagnostic antibodies. These antibodies can also serve as a tool for detecting MUC17 in laboratory animals.

[0007] The present invention further provides a polynucleotide encoding the antibody, a vector containing the polynucleotide, a host cell expressing the polynucleotide, a diagnostic composition containing the polynucleotide, and a kit containing the antibody of the present invention as a diagnostic tool, wherein the other components include, but are not limited to, at least one of a second antibody, an enzyme, a buffer, an instruction manual, a calibration tool, a control, etc.

[0008] The present invention provides an antibody or antigen-binding fragment thereof that specifically binds to MUC17, wherein the antibody or antigen-binding fragment thereof binds to the MUC17 protein on the cell surface.

[0009] The present invention provides a MUC17-specific binding antibody or an antigen-binding fragment thereof, wherein the antibody or antigen-binding fragment comprises a heavy chain variable region, wherein the heavy chain variable region contains HCDR1 of the sequence SEQ ID NO: 3 or SEQ ID NO: 11; and / or, the heavy chain variable region contains HCDR2 of the sequence SEQ ID NO: 4 or SEQ ID NO: 12; and / or, the heavy chain variable region contains HCDR3 of the sequence SEQ ID NO: 5 or SEQ ID NO: 13.

[0010] In some embodiments, the heavy chain variable region contains HCDR1 of the SEQ ID NO: 3 sequence; and / or, the heavy chain variable region contains HCDR2 of the SEQ ID NO: 4 sequence; and / or, the heavy chain variable region contains HCDR3 of the SEQ ID NO: 5 sequence.

[0011] In some embodiments, the heavy chain variable region contains HCDR1 of the sequence SEQ ID NO: 11; and / or, the heavy chain variable region contains HCDR2 of the sequence SEQ ID NO: 12; and / or, the heavy chain variable region contains HCDR3 of the sequence SEQ ID NO: 13.

[0012] The antibody or antigen-binding fragment thereof that specifically binds to MUC17 provided by the present invention comprises a light chain variable region, wherein the light chain variable region contains LCDR1 of the sequence SEQ ID NO: 6 or SEQ ID NO: 14; and / or, the light chain variable region contains LCDR2 of the sequence SEQ ID NO: 7 or SEQ ID NO: 15; and / or, the light chain variable region contains LCDR3 of the sequence SEQ ID NO: 8 or SEQ ID NO: 16.

[0013] In some embodiments, the light chain variable region contains LCDR1 of the sequence SEQ ID NO: 6; and / or, the light chain variable region contains LCDR2 of the sequence SEQ ID NO: 7; and / or, the light chain variable region contains LCDR3 of the sequence SEQ ID NO: 8.

[0014] In some embodiments, the light chain variable region contains LCDR1 of the sequence SEQ ID NO: 14; and / or, the light chain variable region contains LCDR2 of the sequence SEQ ID NO: 15; and / or, the light chain variable region contains LCDR3 of the sequence SEQ ID NO: 16.

[0015] The antibody or antigen-binding fragment thereof that specifically binds to MUC17 provided by the present invention comprises a heavy chain variable region (VH) and a light chain variable region (VL); wherein the heavy chain variable region contains HCDR1 of the sequence SEQ ID NO: 3 or SEQ ID NO: 11, HCDR2 of the sequence SEQ ID NO: 4 or SEQ ID NO: 12, and HCDR3 of the sequence SEQ ID NO: 5 or SEQ ID NO: 13; and the light chain variable region contains LCDR1 of the sequence SEQ ID NO: 6 or SEQ ID NO: 14, LCDR2 of the sequence SEQ ID NO: 7 or SEQ ID NO: 15, and LCDR3 of the sequence SEQ ID NO: 8 or SEQ ID NO: 16.

[0016] In some embodiments, the antibody or antigen-binding fragment thereof that specifically binds to MUC17 provided by the present invention comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises HCDR1 as shown in SEQ ID NO: 3, HCDR2 as shown in SEQ ID NO: 4, and HCDR3 as shown in SEQ ID NO: 5; and the light chain variable region comprises LCDR1 as shown in SEQ ID NO: 6, LCDR2 as shown in SEQ ID NO: 7, and LCDR3 as shown in SEQ ID NO: 8.

[0017] In some embodiments, the antibody or antigen-binding fragment thereof that specifically binds to MUC17 provided by the present invention comprises a heavy chain variable region and a light chain variable region, wherein the heavy chain variable region comprises HCDR1 as shown in SEQ ID NO: 11, HCDR2 as shown in SEQ ID NO: 12, and HCDR3 as shown in SEQ ID NO: 13; and the light chain variable region comprises LCDR1 as shown in SEQ ID NO: 14, LCDR2 as shown in SEQ ID NO: 15, and LCDR3 as shown in SEQ ID NO: 16.

[0018] The present invention provides an antibody or antigen-binding fragment that specifically binds to MUC17, wherein the antibody or antigen-binding fragment comprises a heavy chain variable region (VH) and / or a light chain variable region (VL), wherein the VH comprises an amino acid sequence having at least 90% identity with the sequence shown in SEQ ID NO: 1 or SEQ ID NO: 9, and the VL comprises an amino acid sequence having at least 90% identity with the sequence shown in SEQ ID NO: 2 or SEQ ID NO: 10.

[0019] In some embodiments, the antibody or antigen-binding fragment of the present invention comprises a heavy chain variable region as shown in SEQ ID NO: 1 or SEQ ID NO: 9, which includes HCDR1 as shown in SEQ ID NO: 3 or SEQ ID NO: 11, HCDR2 as shown in SEQ ID NO: 4 or SEQ ID NO: 12, and HCDR3 as shown in SEQ ID NO: 5 or SEQ ID NO: 13.

[0020] In some embodiments, the antibody or antigen-binding fragment of the present invention comprises a heavy chain variable region comprising an amino acid sequence as shown in SEQ ID NO: 1, or comprising an amino acid sequence having at least 90% identity with SEQ ID NO: 1, comprising HCDR1 as shown in SEQ ID NO: 3, HCDR2 as shown in SEQ ID NO: 4, and HCDR3 as shown in SEQ ID NO: 5, or the heavy chain variable region is composed of an amino acid sequence as shown in SEQ ID NO: 1.

[0021] In some embodiments, the antibody or antigen-binding fragment of the present invention comprises a heavy chain variable region comprising an amino acid sequence as shown in SEQ ID NO: 9, or comprising an amino acid sequence having at least 90% identity with SEQ ID NO: 9, comprising HCDR1 as shown in SEQ ID NO: 11, HCDR2 as shown in SEQ ID NO: 12, and HCDR3 as shown in SEQ ID NO: 13, or the heavy chain variable region is composed of an amino acid sequence as shown in SEQ ID NO: 9.

[0022] In some embodiments, the antibody or antigen-binding fragment of the present invention comprises a light chain variable region as shown in SEQ ID NO: 2 or SEQ ID NO: 10, which includes LCDR1 as shown in SEQ ID NO: 6 or SEQ ID NO: 14, LCDR2 as shown in SEQ ID NO: 7 or SEQ ID NO: 15, and LCDR3 as shown in SEQ ID NO: 8 or SEQ ID NO: 16.

[0023] In some embodiments, the antibody or antigen-binding fragment of the present invention comprises a light chain variable region comprising an amino acid sequence as shown in SEQ ID NO: 2, or comprising an amino acid sequence having at least 90% identity with SEQ ID NO: 2, comprising LCDR1 as shown in SEQ ID NO: 6, LCDR2 as shown in SEQ ID NO: 7, and LCDR3 as shown in SEQ ID NO: 8, or the light chain variable region is composed of an amino acid sequence as shown in SEQ ID NO: 2.

[0024] In some embodiments, the antibody or antigen-binding fragment of the present invention comprises a light chain variable region comprising an amino acid sequence as shown in SEQ ID NO: 10, or comprising an amino acid sequence having at least 90% identity with SEQ ID NO: 10, comprising LCDR1 as shown in SEQ ID NO: 14, LCDR2 as shown in SEQ ID NO: 15, and LCDR3 as shown in SEQ ID NO: 16, or the light chain variable region is composed of an amino acid sequence as shown in SEQ ID NO: 10.

[0025] In some embodiments, the antibody or antigen-binding fragment of the present invention comprises a heavy chain variable region as shown in SEQ ID NO: 1, which includes HCDR1 as shown in SEQ ID NO: 3, HCDR2 as shown in SEQ ID NO: 4, and HCDR3 as shown in SEQ ID NO: 5; and a light chain variable region as shown in SEQ ID NO: 2, which includes LCDR1 as shown in SEQ ID NO: 6, LCDR2 as shown in SEQ ID NO: 7, and LCDR3 as shown in SEQ ID NO: 8.

[0026] In some embodiments, the antibody or antigen-binding fragment of the present invention comprises a heavy chain variable region as shown in SEQ ID NO: 9, which includes HCDR1 as shown in SEQ ID NO: 11, HCDR2 as shown in SEQ ID NO: 12, and HCDR3 as shown in SEQ ID NO: 13; and a light chain variable region as shown in SEQ ID NO: 10, which includes LCDR1 as shown in SEQ ID NO: 14, LCDR2 as shown in SEQ ID NO: 15, and LCDR3 as shown in SEQ ID NO: 16.

[0027] In some embodiments, the antibody or antigen-binding fragment of the present invention comprises a heavy chain variable region as shown in SEQ ID NO: 1, and a light chain variable region as shown in SEQ ID NO: 2.

[0028] In some embodiments, the antibody or antigen-binding fragment of the present invention comprises a heavy chain variable region as shown in SEQ ID NO: 9, and a light chain variable region as shown in SEQ ID NO: 10.

[0029] In some embodiments, the antibody that specifically binds to MUC17 according to the present invention is a monoclonal antibody.

[0030] In some embodiments, the antibody or antigen-binding fragment thereof that specifically binds to MUC17 according to the present invention binds to human MUC17 or cells expressing human MUC17, for example, specifically to human MUC17 in immunohistochemical assays, cells expressing MUC17 in fluorescence-activated cell sorting assays, and fixed and permeabilized cells expressing MUC17; optionally, it does not bind to MUC1.

[0031] In some embodiments, the antibody described in this invention is an IgG, IgD, IgA, IgE, or IgM antibody. In some embodiments, the antibody described in this invention is an IgG antibody, such as IgG1, IgG2, IgG3, or IgG4 antibodies or modified forms thereof.

[0032] In some embodiments, the antibody or its antigen-binding fragment described in this invention is a murine antibody or a humanized antibody.

[0033] In some embodiments, the antigen-binding fragment of the antibody that specifically binds to MUC17 according to the present invention is, for example, Fab, Fab'-SH, Fv, scFv, or (Fab')2 fragment.

[0034] In some embodiments, the antibody or antigen-binding fragment thereof that specifically binds to MUC17 according to the present invention further comprises an antibody constant region. The constant region may be derived from germline antibodies of any suitable known species, for example, the antibody constant region may be derived from the constant region of a rodent (e.g., rat, mouse, rabbit) germline immunoglobulin sequence. In some embodiments, the antibody constant region comprises a heavy chain constant region and a light chain constant region. In some embodiments, the heavy chain constant region is, for example, derived from the heavy chain constant region of immunoglobulin IgG, such as the heavy chain constant region of IgG1, IgG2, IgG3, or IgG4. In some embodiments, the light chain constant region is derived from, for example, a light chain of kappa (κ) or lambda (λ).

[0035] In some embodiments, one or more amino acid modifications (e.g., insertion, deletion, or substitution) may be introduced into the Fc region of the antibody that specifically binds to MUC17 as described in this invention to generate Fc region variants. These Fc region variants have increased or decreased (e.g., lack) corresponding effector functions. In some embodiments, modification of the Fc region has altered the affinity of the Fc region for its ligand FcRn.

[0036] In some embodiments, the heavy chain constant region comprises the amino acid sequence shown in SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24 or SEQ ID NO:25, or comprises an amino acid sequence having at least 90% identity with SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24 or SEQ ID NO:25, or is composed of SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24 or SEQ ID NO:25.

[0037] In some embodiments, the light chain constant region comprises the amino acid sequence shown in SEQ ID NO:26, or comprises an amino acid sequence having at least 90% identity with SEQ ID NO:26, or is composed of SEQ ID NO:26.

[0038] In some embodiments, the antibody that specifically binds to MUC17 according to the present invention comprises a heavy chain, wherein the heavy chain comprises the amino acid sequence shown in SEQ ID NO:18, or comprises an amino acid sequence having at least 90% identity with SEQ ID NO:18, or is composed of SEQ ID NO:18.

[0039] In some embodiments, the antibody that specifically binds to MUC17 according to the present invention comprises a heavy chain, wherein the heavy chain comprises the amino acid sequence shown in SEQ ID NO:20, or comprises an amino acid sequence having at least 90% identity with SEQ ID NO:20, or is composed of SEQ ID NO:20.

[0040] In some embodiments, the antibody that specifically binds to MUC17 according to the present invention comprises a light chain, wherein the light chain comprises the amino acid sequence shown in SEQ ID NO:19, or comprises an amino acid sequence having at least 90% identity with SEQ ID NO:19, or is composed of SEQ ID NO:19.

[0041] In some embodiments, the antibody that specifically binds to MUC17 according to the present invention comprises a light chain, wherein the light chain comprises the amino acid sequence shown in SEQ ID NO:21, or comprises an amino acid sequence having at least 90% identity with SEQ ID NO:21, or is composed of SEQ ID NO:21.

[0042] In some embodiments, the antibody that specifically binds to MUC17 according to the present invention comprises a heavy chain and a light chain, wherein the heavy chain comprises the amino acid sequence shown in SEQ ID NO:18, or comprises an amino acid sequence having at least 90% identity with SEQ ID NO:18, or is composed of SEQ ID NO:18, and the light chain comprises the amino acid sequence shown in SEQ ID NO:19, or comprises an amino acid sequence having at least 90% identity with SEQ ID NO:19, or is composed of SEQ ID NO:19.

[0043] In some embodiments, the antibody that specifically binds to MUC17 according to the present invention comprises a heavy chain and a light chain, wherein the heavy chain comprises the amino acid sequence shown in SEQ ID NO:20, or comprises an amino acid sequence having at least 90% identity with SEQ ID NO:20, or is composed of SEQ ID NO:20, and the light chain comprises the amino acid sequence shown in SEQ ID NO:21, or comprises an amino acid sequence having at least 90% identity with SEQ ID NO:21, or is composed of SEQ ID NO:21.

[0044] As one aspect of the invention, the invention also relates to a polynucleotide molecule that encodes a binding fragment of any of the aforementioned antibodies or their antigens.

[0045] As one aspect of the invention, the invention also relates to a carrier comprising the aforementioned polynucleotide molecule.

[0046] As one aspect of the invention, the invention also relates to host cells that contain the aforementioned polynucleotide molecules or vectors through transformation or transfection.

[0047] In some embodiments, the present invention also relates to a method for producing the aforementioned antibody or antigen-binding fragment thereof, the method comprising culturing the aforementioned host cells under conditions that allow the antibody to be expressed and recovering the resulting antibody or antigen-binding fragment thereof from the culture.

[0048] As one aspect of the invention, the invention further relates to compositions comprising any of the aforementioned antibodies or their antigen-binding fragments.

[0049] As one aspect of the invention, the invention further relates to a detection system comprising any of the aforementioned antibodies or their antigen-binding fragments, or antibodies produced by the aforementioned methods.

[0050] As one aspect of the invention, the invention further relates to a diagnostic method comprising detecting the presence, distribution, or expression level of MUC17 in a sample from a subject using any of the aforementioned antibodies or their antigen-binding fragments, or antibodies, polynucleotide molecules, vectors, compositions, or detection systems produced by the aforementioned methods, and comparing the detection results with a reference value to determine whether the subject suffers from a disease associated with abnormal MUC17 expression. In some embodiments, the reference value is selected from (i) a predefined value for MUC17 expression; (ii) the MUC17 expression level determined in a control sample; or (iii) the MUC17 expression level determined in a sample obtained from the same source or subject at a previous time point.

[0051] As one aspect of this invention, the invention also relates to the use of any of the aforementioned antibodies or their antigen-binding fragments, polynucleotide molecules, carriers, compositions or detection systems in the preparation of diagnostic agents or diagnostic products used in the aforementioned diagnostic methods.

[0052] In some implementations, any of the aforementioned antibodies or their antigen-binding fragments, polynucleotide molecules, carriers, compositions, or detection systems are used for diagnostic purposes.

[0053] In some implementations, any of the aforementioned antibodies or their antigen-binding fragments, polynucleotide molecules, carriers, compositions, or detection systems are used as diagnostic agents or diagnostic products.

[0054] As one aspect of the invention, the invention further relates to one or more uses in which any of the aforementioned antibodies or antibodies, polynucleotide molecules, carriers, compositions or detection systems produced by the aforementioned methods are used in methods for detecting tumor growth.

[0055] In some embodiments, any of the aforementioned antibodies or their antigen-binding fragments, polynucleotide molecules, carriers, compositions, or detection systems are used in the preparation of diagnostic agents or diagnostic products, which are used in methods for detecting tumor growth.

[0056] In some implementations, any of the aforementioned antibodies or their antigen-binding fragments, polynucleotide molecules, carriers, compositions, or detection systems are used to detect tumor growth.

[0057] In some embodiments, any of the aforementioned antibodies or their antigen-binding fragments, polynucleotide molecules, carriers, compositions, or detection systems are used as diagnostic agents or diagnostic products in methods for detecting tumor growth.

[0058] In some implementations, the method includes determining the expression level of MUC17 in samples from patients suspected of having cancer and negative controls, and optionally positive controls.

[0059] In some embodiments, the method includes comparing MUC17 expression levels among these samples, optionally wherein the MUC17 expression levels in the positive and / or negative controls may be derived from stored data in at least one positive and / or negative control obtained in a method for detecting tumor growth.

[0060] In some embodiments, the expression levels of MUC17 in negative and / or positive control samples in the method may be derived from stored data of the average expression levels of more than one negative and / or more than one positive control sample obtained in the method for detecting tumor growth.

[0061] In some embodiments, the sample in the method is a solid tissue sample and / or a liquid tissue sample.

[0062] As one aspect of the present invention, the present invention provides a method for identifying MUC17-expressing cells, the method comprising contacting the cells with an antibody or composition described in the present invention and detecting the specific binding of the antibody or composition to the cells.

[0063] In some embodiments, the present invention provides a method for identifying the presence of MUC17 in a tissue sample, the method comprising contacting the tissue sample with an antibody or composition described in the present invention and detecting the specific binding of the antibody or composition to the tissue to identify tissue expressing MUC17.

[0064] In some embodiments, the present invention provides a method for identifying a subject who has or is at risk of having a disease associated with abnormal MUC17 expression, the method comprising contacting a tissue sample of the subject with an antibody or composition of the present invention and detecting specific binding of the antibody or composition to the tissue, thereby identifying a subject who has or is at risk of having a disease associated with abnormal MUC17 expression.

[0065] In some embodiments, the present invention relates to a method for detecting and / or quantifying MUC17 expression in a sample, the method comprising the following steps: (a) Using any of the antibodies described above according to the present invention or antibodies produced by the methods described above according to the present invention, or using the detection system described above according to the present invention, or using the composition described above according to the present invention, to determine the expression level of MUC17 in a sample. (b) Compare the MUC17 expression level determined in step (a) with: (i) a predefined value for MUC17 expression level; (ii) the MUC17 expression level determined in a control sample; or (iii) the MUC17 expression level determined in a sample obtained from the same source or subject at a previous time point.

[0066] In some embodiments, the present invention relates to a method for diagnosing neoplastic diseases associated with abnormal or increased MUC17 expression, or a method for diagnosing whether a subject has a tumor associated with abnormal or increased MUC17 expression, the method comprising the following steps: (a) Using any of the antibodies described above according to the present invention or antibodies produced by the methods described above according to the present invention, or using the detection system described above according to the present invention, or using the composition described above according to the present invention, to determine the expression level of MUC17 in a sample. (b) Compare the MUC17 expression level determined in step (a) with: (i) a predefined cutoff value indicating the absence of this disease, or (ii) the MUC17 expression level determined in a negative control sample representing the absence of this neoplastic disease, wherein a higher MUC17 expression level determined in step (a) compared to the predefined cutoff value in (i) or the MUC17 expression level determined in a negative control sample in (ii) indicates the presence of a disease associated with abnormal or increased MUC17 expression.

[0067] In some embodiments, the present invention relates to a method for monitoring the progression of diseases associated with or related to abnormal MUC17 expression or increased MUC17 expression levels, or a method for monitoring the response to treatment for diseases associated with or related to abnormal MUC17 expression or increased MUC17 expression levels, the method comprising the following steps: (a) Using any of the antibodies described above or antibodies produced according to the methods described above, or using the detection system described above, or using the composition described above, to determine the level of MUC17 expression in a sample obtained from a subject diagnosed with the disease at a first time point. (b) Using any of the antibodies described above according to the invention or antibodies produced according to the methods described above according to the invention, or using the detection system described above according to the invention, or using the composition described above according to the invention, to determine the level of MUC17 expression in a sample obtained from the subject at a second time point or after treatment; and (c) Compare the MUC17 expression level determined in step (a) with the MUC17 expression level determined in step (b); A higher MUC17 expression level in step (b) compared to the MUC17 expression level determined in step (a) indicates that the disease is progressing, and / or a lower MUC17 expression level in step (b) compared to the MUC17 expression level determined in step (a) indicates that the disease is entering a remission phase or that the disease is responding to the treatment.

[0068] In the aforementioned methods, the method for detecting the presence of MUC17 in the sample can be any suitable protein detection method in the art, whether for the expression level or quantity of MUC17. In some embodiments, the method is, for example, immunohistochemical detection, protein immunoblotting, fluorescence detection, flow cytometry, or quantitative PCR (e.g., real-time quantitative PCR).

[0069] In some embodiments, the present invention relates to the use of any of the aforementioned antibodies or their antigen-binding fragments, polynucleotide molecules, carriers, compositions or detection systems in the preparation of products for identifying cells expressing MUC17, or for detecting and / or quantifying MUC17 expression in a sample.

[0070] In some embodiments, the present invention relates to the use of any of the aforementioned antibodies or their antigen-binding fragments, polynucleotide molecules, carriers, compositions, or detection systems in the preparation of diagnostic reagents or products, said diagnostic reagents or products being used for: 1) Identify subjects who have or are at risk of having a disease associated with abnormal or increased MUC17 expression; 2) Used to diagnose neoplastic diseases associated with abnormal or increased MUC17 expression; 3) Used to monitor the progression of diseases associated with abnormal MUC17 expression or increased MUC17 expression; or 4) Used to monitor the response to treatment for diseases associated with abnormal MUC17 expression or increased MUC17 expression.

[0071] In some embodiments, any of the aforementioned antibodies or their antigen-binding fragments, polynucleotide molecules, vectors, compositions, or detection systems are used for: 1) To identify cells expressing MUC17, or to detect and / or quantify MUC17 expression in samples; 2) Identify subjects who have or are at risk of having diseases associated with MUC17 expression; 3) Diagnose neoplastic diseases associated with abnormal or increased MUC17 expression; 4) Monitor the progression of diseases associated with abnormal MUC17 expression or increased MUC17 expression; or 5) Used to monitor the response to treatment for diseases associated with abnormal MUC17 expression or increased MUC17 expression.

[0072] In some embodiments, the diagnostic methods and uses described above of the present invention utilize biological samples, preferably human biological samples, such as tissue samples or samples containing cultured cells. In some embodiments, the samples are solid tissue samples and / or liquid tissue samples. In some embodiments, the samples are processed for use in immunohistochemical detection methods (IHC methods).

[0073] In some embodiments, the diagnostic methods and uses described above of the present invention utilize samples obtained from human subjects, preferably from subjects suspected of having or suffering from diseases associated with abnormal or increased MUC17 expression, or from subjects who have received treatment for diseases associated with abnormal or increased MUC17 expression. In some embodiments, the sample is a solid tissue sample, a liquid tissue sample, a sample containing cultured cells, preferably a cell sample, a tissue sample (e.g., tissue sections and tissue microarrays), preferably a tissue section (e.g., paraffin sections or frozen sections), and more preferably a formaldehyde-fixed paraffin-embedded (FFPE) section.

[0074] In some embodiments, the diagnostic methods and uses described above in this invention refer to diseases selected from: gastric cancer, esophageal cancer, intestinal cancer, gastroesophageal cancer, gastroesophageal junction adenocarcinoma, colorectal cancer, pancreatic cancer, breast cancer, liver cancer, ovarian cancer, and bile duct cancer.

[0075] In some embodiments, any of the antibodies, polynucleotide molecules, carriers, compositions, or detection systems described above are used for the detection of MUC17 expression in an immunohistochemical (IHC) method, which includes the step of providing a tissue sample (including liquid biopsy material, such as cells obtained from an individual, human, or animal, or from a cell culture). In some embodiments, the sample material is typically provided on a support, such as a carrier like a glass or plastic slide. Optionally, the tissue may be fixed using a fixation medium known in the art, such as paraffin, ethanol, or acetone, or any other suitable medium that ensures fixation of the tissue or cells to the support. In some embodiments, the method further includes a subsequent sample preparation step, such as deparaffining, to allow the antibodies described in this invention to access the target MUC17. In some embodiments, the method further includes incubating the tissue sample with a sufficient amount of the antibodies or compositions disclosed in this invention to allow these antibodies or compositions to bind to MUC17 present in the tissue sample. After an incubation time that is generally known and reasonable, the reaction of the tissue sample with the antibody is typically stopped by washing the support and the tissue sample with a medium such as PBS. To examine the effects of incubation media, fixation media, incubation time, and temperature, positive control materials known to express the target MUC17 and negative controls known not to express MUC17 are typically used. These antibodies are added to the positive control material in a titration assay, thereby determining the concentration at which the antibodies preventing the specific and / or selective detection of MUC17 can be determined. Determining the correct amount of antibody for appropriate positive and negative controls is common knowledge known to those skilled in the art of immunocytochemistry and, particularly, immunohistochemistry. Once appropriate controls are established, test materials, such as those obtained from patients suspected of having a disease characterized by pathologically enhanced / distributed MUC17 expression (e.g., cancer), can be analyzed. As discussed further below, antibodies may carry a detectable marker, or they may be recognized by another conjugate carrying such a marker. These techniques are referred to by those skilled in the art as direct immunocytology / immunohistochemistry detection and indirect immunocytology / immunohistochemistry detection. Once the required incubation and washing steps have been performed on the tissue samples and controls, the tissues can be analyzed, for example, using an immunofluorescence-based detection system. The intensity and quantity of detectable markers can be determined and compared with positive and negative controls and / or with known standard values. Based on the comparison results, a conclusion can be drawn regarding whether the sample is positive for MUC17 expression.

[0076] In some embodiments, any of the antibodies, polynucleotide molecules, vectors, compositions or detection systems described above are used in a protein immunoblotting method (western blot or WB method) for detecting MUC17 expression.

[0077] In some embodiments, any of the antibodies, polynucleotide molecules, vectors, compositions or detection systems described above are used in real-time quantitative PCR (qPCR) for detecting MUC17 expression.

[0078] As one aspect of the invention, a kit is also provided comprising an antibody or antigen-binding fragment thereof that specifically binds to MUC17 as described above, or a combination thereof. In some embodiments, the kit is a diagnostic kit.

[0079] In some embodiments, the content of MUC17 in the sample refers to the amount detected by the MUC17 antibody after the sample is contacted with the MUC17 antibody of the present invention.

[0080] In some implementations, the presence or absence of MUC17 in the sample is determined by whether the MUC17 antibody described in this invention binds to the sample.

[0081] In some implementations, the content of MUC17 in the sample is determined by the amount of MUC17 antibody bound to the sample as described in this invention, which indicates the level or expression level of MUC17 in the sample.

[0082] This invention provides a first monoclonal antibody (or its antigen-binding fragment) for a detection system, comprising: (a) A heavy chain variable region as shown in SEQ ID NO: 1 or SEQ ID NO: 9, the heavy chain variable region comprising HCDR1 as shown in SEQ ID NO: 3 or SEQ ID NO: 11, HCDR2 as shown in SEQ ID NO: 4 or SEQ ID NO: 12, and HCDR3 as shown in SEQ ID NO: 5 or SEQ ID NO: 13; and a light chain variable region as shown in SEQ ID NO: 2 or SEQ ID NO: 10, the light chain variable region comprising LCDR1 as shown in SEQ ID NO: 6 or SEQ ID NO: 14, LCDR2 as shown in SEQ ID NO: 7 or SEQ ID NO: 15, and LCDR3 as shown in SEQ ID NO: 8 or SEQ ID NO: 16; or (b) A monoclonal antibody that binds to the same MUC17 protein as the antibody in (a) or that competes with the antibody in (a) for binding to MUC17.

[0083] This invention also provides monoclonal antibodies for the detection system: (a) Contains a heavy chain variable region as shown in SEQ ID NO: 1 or SEQ ID NO: 9; (b) Contains a light chain variable region as shown in SEQ ID NO: 2 or SEQ ID NO: 10; (c) Containing a heavy chain variable region as shown in SEQ ID NO: 1 or SEQ ID NO: 9 and a light chain variable region as shown in SEQ ID NO: 2 or SEQ ID NO: 10; or (d) The antibody that binds to the same MUC17 protein as the antibody in (c) or the monoclonal antibody that competes with the antibody in (c) for binding to MUC17.

[0084] The present invention also provides the use of the antibodies of the present invention in the following aspects or the use of the detection system of the present invention in the following aspects: 1. Detect MUC17 in the sample; 2. Quantitative analysis of MUC17 in the sample; 3. Diagnose diseases associated with abnormal or increased MUC17 expression; 4. Stratify patients diagnosed with diseases associated with abnormal or increased MUC17 expression; 5. Monitor the progression of diseases associated with abnormal or increased MUC17 expression; or 6. Monitor the response to treatment for diseases associated with abnormal or increased MUC17 expression.

[0085] The present invention also provides a method for detecting and / or quantifying MUC17 in a sample, the method comprising the following steps: (a) Determining the MUC17 content in a sample using the antibody of the present invention or the detection system of the present invention; and (b) Compare the MUC17 content determined in step (a) with: (i) a predefined value for the MUC17 content, (ii) the MUC17 content determined in a control sample, or (iii) the MUC17 content determined in a sample obtained from the same source or subject at a previous time point.

[0086] This invention provides a method for diagnosing diseases related to abnormal or increased MUC17 expression, the method comprising the following steps: (a) Determining the MUC17 content in a sample using the antibody, composition, or detection system of the present invention; and (b) Compare the MUC17 content determined in step (a) with: (i) a predefined cutoff value for the MUC17 content indicating the absence of the disease, or (ii) the MUC17 content determined in a control sample representing the absence of the disease, wherein a higher MUC17 content determined in step (a) compared to the predefined cutoff value in (i) or the MUC17 content determined in the control sample in (ii) indicates the presence of a disease associated with abnormal or increased MUC17 expression.

[0087] The present invention also provides a method for monitoring the progression of diseases associated with abnormal or increased MUC17 expression, or for monitoring the response to treatment of diseases associated with abnormal or increased MUC17 expression, the method comprising the following steps: (a) Using the antibody of the present invention or the detection system of the present invention, determine at a first time point the content of MUC17 in a biological sample obtained from a subject diagnosed with the disease; (b) Using the antibody of the present invention, or using the detection system of the present invention, determining the MUC17 content in a biological sample obtained from the subject at a second (later) time point or after treatment; and (c) Compare the MUC17 content determined in step (a) with the MUC17 content determined in step (b); A higher MUC17 content determined in step (a) compared to the MUC17 content determined in step (b) indicates that the disease is progressing, and / or a lower MUC17 content determined in step (a) compared to the MUC17 content determined in step (b) indicates that the disease is entering a remission phase or that the disease is responding to the treatment.

[0088] The antibody or antigen-binding fragment of the present invention that specifically binds to MUC17 comprises a heavy chain variable region having at least 75% or 80% homology with SEQ ID NO: 1 or SEQ ID NO: 9, preferably at least 85%, 90%, 91%, 92%, 93%, or 94%, and more preferably 95%, 96%, 97%, 98%, or 99% homology with an amino acid sequence.

[0089] The heavy chain variable region of the MUC17 antibody of the present invention includes HCDR1 as shown in SEQ ID NO: 3 or SEQ ID NO: 11, HCDR2 as shown in SEQ ID NO: 4 or SEQ ID NO: 12, and HCDR3 as shown in SEQ ID NO: 5 or SEQ ID NO: 13.

[0090] The antibody or antigen-binding fragment of the present invention that specifically binds to MUC17 comprises a light chain variable region having at least 75% or 80% homology with SEQ ID NO: 2 or SEQ ID NO: 10, preferably at least 85%, 90%, 91%, 92%, 93%, 94%, more preferably 95%, 96%, 97%, 98%, or 99% homology with the amino acid sequence.

[0091] The light chain variable region of the MUC17 antibody or its antigen-binding fragment described in this invention includes LCDR1 as shown in SEQ ID NO: 6 or SEQ ID NO: 14, LCDR2 as shown in SEQ ID NO: 7 or SEQ ID NO: 15, and LCDR3 as shown in SEQ ID NO: 8 or SEQ ID NO: 16.

[0092] The antibody or antigen-binding fragment of the present invention that specifically binds to MUC17 comprises a heavy chain variable region having at least 75% or 80% homology with SEQ ID NO: 1 or SEQ ID NO: 9, preferably at least 85%, 90%, 91%, 92%, 93%, 94%, more preferably 95%, 96%, 97%, 98%, or 99% homology with an amino acid sequence, comprising HCDR1 as shown in SEQ ID NO: 3 or SEQ ID NO: 11, HCDR2 as shown in SEQ ID NO: 4 or SEQ ID NO: 12, and HCDR3 as shown in SEQ ID NO: 5 or SEQ ID NO: 13.

[0093] The antibody or antigen-binding fragment of the present invention that specifically binds to MUC17 comprises a light chain variable region having at least 75% or 80% homology to SEQ ID NO: 2 or SEQ ID NO: 10, preferably at least 85%, 90%, 91%, 92%, 93%, 94%, more preferably 95%, 96%, 97%, 98%, or 99% homology, comprising LCDR1 as shown in SEQ ID NO: 6 or SEQ ID NO: 14, LCDR2 as shown in SEQ ID NO: 7 or SEQ ID NO: 15, and LCDR3 as shown in SEQ ID NO: 8 or SEQ ID NO: 16.

[0094] The antibody of the present invention specifically binds to MUC17, comprising...

[0095] (a) The heavy chain variable region as shown in SEQ ID NO: 1 or SEQ ID NO: 9; (b) The light chain variable region as shown in SEQ ID NO: 2 or SEQ ID NO: 10; (c) The heavy chain variable region as shown in SEQ ID NO: 1 or SEQ ID NO: 9 and the light chain variable region as shown in SEQ ID NO: 2 or SEQ ID NO: 10; (d) The antibody binds to the same MUC17 protein as the antibody in (c) or competes with the antibody in (c) for binding to MUC17.

[0096] The present invention provides a composition comprising the antibody described herein that specifically binds to MUC17.

[0097] This invention provides a detection system comprising: (a) The first monoclonal antibody combined with MUC17, and (b) A second monoclonal antibody that binds to MUC17. The binding of the first monoclonal antibody to MUC17 occurs in the presence of the second monoclonal antibody that is bound to MUC17, and / or the binding of the second monoclonal antibody to MUC17 occurs in the presence of the first monoclonal antibody that is bound to MUC17.

[0098] In some embodiments, in the detection system of the present invention, the first monoclonal antibody and / or the second monoclonal antibody comprise mouse heavy chain variable regions and / or mouse light chain variable regions.

[0099] In some embodiments, in the detection system of the present invention, the affinity (KD) of the first monoclonal antibody and / or the second monoclonal antibody for MUC17 is about ≤10. -7 M, ≤10 -8 M, ≤10 -9 M, or ≤10 -10 M.

[0100] In some embodiments, in the detection system of the present invention, the first monoclonal antibody comprises: (a) A heavy chain variable region as shown in SEQ ID NO: 1 or SEQ ID NO: 9, the heavy chain variable region comprising HCDR1 as shown in SEQ ID NO: 3 or SEQ ID NO: 11, HCDR2 as shown in SEQ ID NO: 4 or SEQ ID NO: 12, and HCDR3 as shown in SEQ ID NO: 5 or SEQ ID NO: 13; and a light chain variable region as shown in SEQ ID NO: 2 or SEQ ID NO: 10, the light chain variable region comprising LCDR1 as shown in SEQ ID NO: 6 or SEQ ID NO: 14, LCDR2 as shown in SEQ ID NO: 7 or SEQ ID NO: 15, and LCDR3 as shown in SEQ ID NO: 8 or SEQ ID NO: 16; (b) The antibody that binds to the same MUC17 protein as the antibody in (a) or the antibody that competes with the antibody in (a) for binding to MUC17.

[0101] The present invention provides a diagnostic kit comprising the MUC17 antibody, nucleic acid molecule, vector, host cell, or composition described in the present invention.

[0102] The present invention also provides a kit for a single-dose administration unit. The kit of the present invention may further contain a first acceptor comprising a dried / lyophilized MUC17 antibody of the present invention or an antigen-binding fragment thereof, and a second acceptor comprising an aqueous formulation.

[0103] In some implementations, the kit includes an instruction manual, which may be in printed or electronic form (such as a paper manual, an electronic document linked to a QR code, or an operation video / text document stored on a USB flash drive). The instruction manual contains at least one or more of the following: the purpose of the kit, the components of the kit, instructions for operation, and standards for interpreting results.

[0104] Terminology Definition

[0105] This invention provides an antibody that specifically binds to MUC17, or an antigen-binding portion or derivative thereof. Whenever the term "antibody" (i.e., an antibody that specifically binds to MUC17) is used in this invention, it encompasses an "antibody fragment." The definition and description of "antibody" (e.g., a monoclonal antibody that specifically binds to MUC17) provided below for this invention are similarly applicable to any chemically or enzymatically modified antibody, such as an antibody carrying a label (e.g., a fluorescent, radioactive, luminescent, or chromogenic label) or an enzyme capable of generating a detectable signal.

[0106] An antibody (sometimes also called an immunoglobulin) is a protein that binds specifically to its target. An antibody recognizes a unique target, known as an antigen, through its variable region. An antibody can be any immunoglobulin isotype, including IgG (including IgG1, IgG2, IgG3, and IgG4 subtypes), IgA (including IgA1 and IgA2 subtypes), IgM, and IgE. The term "antibody" can include, for example, monoclonal antibodies, chimeric antibodies, recombinant antibodies, deimmunized antibodies, affinity-matured antibodies, humanized antibodies, and human antibodies, as well as antibodies from other species such as rodents (e.g., rabbits, mice, rats, hamsters), goats, etc. Antibodies can be derived from a single source or can be "chimeric," meaning that different parts of the antibody (such as the CDR, frame region, variable region, constant region) can be derived from two different antibodies. The definition of "antibody" according to the invention includes full-length antibodies, as well as camel antibodies and other immunoglobulins generated by biotechnology or protein engineering methods or processes. Antibodies can also be produced in hybridomas.

[0107] A complete IgG antibody typically consists of two full-length heavy chains and two full-length light chains. The light chain comprises a variable region (“VL”) and a constant region (“CL”) with one domain. The variable region of the light chain is located at the amino terminus of the polypeptide. The light chain includes a κ chain and a λ chain. The heavy chain comprises a variable region (“VH”) and a constant region (“CH”) with three domains (in the case of a complete IgG antibody): CH1, CH2, and CH3. The VH domain is located at the amino terminus of the polypeptide, and the CH domain is located at the carboxyl terminus, with CH3 being closest to the carboxyl terminus of the polypeptide.

[0108] In classic full-length antibodies or immunoglobulins, each light (L) chain is linked to a heavy (H) chain by a covalent disulfide bond, while the two H chains are linked to each other by one or more disulfide bonds, depending on the H chain isotype. The heavy chain constant domain (CH) closest to the VH is usually named CH1. Constant domains do not directly participate in antigen binding but exhibit various effector functions, such as antibody-dependent cell-mediated cytotoxicity (ADCC) and complement activation (complement-dependent cytotoxicity, CDC). The Fc region of an antibody is the “tail” region of a classic antibody, which interacts with cell surface receptors called Fc receptors and some proteins in the complement system. In IgG, IgA, and IgD antibody isotypes, the Fc region consists of two identical protein fragments (Fc fragments) derived from the second and third constant domains (CH2 and CH3) of the two heavy chains of the antibody, which dimerize to produce the Fc region. The Fc regions of IgM and IgE contain three heavy-chain constant domains (CH2, CH3, and CH4) in each polypeptide chain. The Fc region also contains portions of a so-called "hinge" region held together by one or more disulfide bonds and non-covalent interactions. The Fc region of naturally occurring IgG has highly conserved N-glycosylation sites. Glycosylation of the Fc fragment is required for Fc receptor-mediated activity. Two Fc fragments in the Fc region can be identical or different from each other. Modifications can be introduced into the Fc fragments to alter the function of the Fc region or improve its stability.

[0109] The monoclonal antibody of the present invention may be an IgG, IgD, IgE, IgM, or IgA antibody. In some embodiments, the monoclonal antibody is an IgG antibody, such as an IgG1, IgG2, IgG3, or IgG4 antibody. The isotype and subtype of the antibody may be mouse, rabbit, hamster, etc. (e.g., mouse IgG, mouse IgG1, etc.).

[0110] In this invention, the term "variable" refers to those portions (i.e., "one or more variable regions") of an antibody or immunoglobulin domain that exhibit sequence variability and participate in determining the specificity and binding affinity of a particular antibody. Typically, the pairing of heavy chain variable regions (VH) and light chain variable regions (VL) together forms a single antigen-binding site. The variability is not uniformly distributed throughout the variable regions of the antibody; it includes highly variable regions called "hypervariable regions" or "complementarity-determining regions" (CDRs), and "framework regions" (FRs) whose sequences are more conserved (i.e., non-hypervariable) compared to the CDRs. The FRs provide a scaffold for the six CDRs in three-dimensional space to form an antigen-binding surface. The variable regions of naturally occurring antibody heavy and light chains each contain four FR regions (FR1, FR2, FR3, and FR4) that primarily employ a β-sheet configuration. The heavy chain variable regions or light chain variable regions contain the following sequences from the N-terminus to the C-terminus: FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4. The hypervariable regions in each chain are closely clustered together by the framework region and often form antigen-binding sites together with hypervariable regions from another chain.

[0111] The term "CDR" and its plural "CDRs," also known as "complementarity-determining regions," refers to regions within the variable domain of an antibody that are highly variable in sequence and form structurally defined loops ("hypervariant loops") and / or contain antigen-contacting residues ("antigen contact sites"). CDRs are primarily responsible for binding to antigenic epitopes. CDRs within the variable domain are typically referred to as CDR1, CDR2, and CDR3, numbered sequentially starting from the N-terminus. For example, the light chain variable domain contains LCHR1, LCDR2, and LCDR3, while the heavy chain variable domain contains HCHR1, HCDR2, and HCDR3. CDRs contain most of the residues responsible for the specific interaction between the antibody or its antigen-binding fragment and the antigen, and thus contribute to the functional activity of the antibody molecule: they are the primary determinants of antigen specificity. The precise definition of CDR boundaries and lengths is subject to different classification and numbering systems. Therefore, CDRs can be referenced using Kabat, Chothia, AbM, IMGT, or any other boundary definition (numbering system).

[0112] The term "monoclonal antibody" (mAb) or monoclonal antibody construct refers to an antibody obtained from a substantially homogeneous population of antibodies, meaning that the antibodies comprising that single population are identical except for possibly small amounts of naturally occurring mutations and / or post-translational modifications. Monoclonal antibodies exhibit high specificity against a single antigenic epitope or determinant on an antigen, compared to conventional (polyclonal) antibody formulations that typically comprise different antibodies targeting different determinants (or epitopes). In addition to their specificity, monoclonal antibodies have the advantage that they can be synthesized through hybridoma culture and are therefore not contaminated by other immunoglobulins. The modifier "monoclonal" indicates the characteristic of antibodies obtained from a substantially homogeneous population of antibodies and should not be construed as requiring the antibody to be produced by any particular method.

[0113] To prepare monoclonal antibodies, any technique that provides antibodies produced through continuous cell line culture can be used. For example, monoclonal antibodies or antigen-binding fragments thereof can be prepared by a hybridoma method first described by Koehler et al., Nature, 256:495 (1975), or by a recombinant DNA method (see, for example, U.S. Patent No. 4,816,567). Examples of other techniques for producing human monoclonal antibodies include tri-hybridoma technology, human B-cell hybridoma technology, and EBV-hybridoma technology.

[0114] Hybridomas can be screened using standard methods such as enzyme-linked immunosorbent assay (ELISA) and surface plasmon resonance (BIACORE) analysis to identify one or more hybridomas that produce antibodies or antigen-binding fragments that specifically bind to a specified antigen.

[0115] Exemplary methods for preparing antibodies or their antigen-binding fragments include screening protein expression libraries, such as phage display or ribosome display libraries. In addition to using display libraries, relevant antigens can be used to immunize non-human animals, such as rodents (e.g., mice, hamsters, rabbits, or rats).

[0116] Monoclonal antibodies can also be obtained from non-human animals and then modified using recombinant DNA techniques known in the art, such as humanization, deimmunization, and chimerism. Examples of modified antibodies, constructs, or antigen-binding fragments thereof include humanized variants of non-human antibodies / antibody constructs, “affinity-matured” antibodies, constructs or antigen-binding fragments thereof, and antibody variants or mutants with one or more altered effector functions.

[0117] The present invention also considers amino acid sequence modifications of the antibodies described herein. For example, it may be desirable to improve the binding affinity and / or other biological properties of the antibody. Amino acid sequence variants of the antibody are prepared by peptide synthesis or by introducing appropriate nucleotide alterations into the nucleic acid molecule encoding the antibody. All amino acid sequence modifications described below should produce antibodies that retain the biological activity of the unmodified parent molecule (binding to MUC17).

[0118] The term “amino acid” or “amino acid residue” typically refers to an amino acid having its recognized definition in the art, such as amino acids selected from the group consisting of: alanine (Ala or A); arginine (Arg or R); asparagine (Asn or N); aspartic acid (Asp or D); cysteine ​​(Cys or C); glutamine (Gln or Q); glutamic acid (Glu or E); glycine (Gly or G); histidine (His or H); isoleucine (Ile or I); leucine (Leu or L); lysine (Lys or K); methionine (Met or M); phenylalanine (Phe or F); proline (Pro or P); serine (Ser or S); threonine (Thr or T); tryptophan (Trp or W); tyrosine (Tyr or Y); and valine (Val or V), and modified, synthetic, or rare amino acids may be used as needed.

[0119] Amino acid modifications include, for example, the deletion, insertion, and / or substitution of residues within the amino acid sequence of a monoclonal antibody or its antigen-binding fragment. Any combination of deletions, insertions, and / or substitutions may be performed to obtain the final monoclonal antibody or its antigen-binding fragment, provided that the final antibody possesses the desired characteristics, such as the biological activity of the unmodified parent molecule (e.g., binding to MUC17). Amino acid changes can also alter the post-translational processes of the antibody, such as changing the number or location of glycosylation sites.

[0120] For example, 1, 2, 3, 4, 5, or 6 amino acids (depending on their respective lengths) may be inserted, deleted, and / or substituted in each CDR, while 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or 25 amino acids may be inserted, deleted, and / or substituted in each FR. Amino acid sequence insertions also include N-terminal and / or C-terminal amino acid additions, ranging in length from, for example, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 residues to polypeptides containing more than 10, for example, one hundred or more residues, as well as intra-sequence insertions of single or multiple amino acid residues.

[0121] The sites of most interest for amino acid modifications (particularly amino acid substitutions) include hypervariable regions, particularly the individual CDRs of the heavy and / or light chains; however, the invention also considers FR variations in the heavy and / or light chains. Substitutions can be conservative substitutions as described in this invention. Preferably, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids can be substituted in the CDR, while 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or 25 amino acids can be substituted in the frame region (FR), depending on the length of the CDR or FR, respectively. For example, if the CDR sequence covers 6 amino acids, it is conceivable that one, two, or three of these amino acids will be substituted. Similarly, if the CDR sequence covers 15 amino acids, it is conceivable that one, two, three, four, five, or six of these amino acids will be substituted. Generally, if an amino acid is substituted in one or all of the CDRs in the heavy chain and / or light chain / variable region, it is assumed that the "substituted" sequence obtained at that time has at least 60% or 65%, more preferably 70% or 75%, even more preferably 80% or 85%, and particularly preferably 90% or 95% identity / homology / similarity with the "original" or "parental" CDR sequence.

[0122] Conservative substitution (also known as conserved mutation or conserved replacement) is an amino acid substitution that changes a given amino acid to a different amino acid with similar biochemical properties (e.g., charge, hydrophobicity, size). Compared to non-conservative substitution, conserved substitutions in proteins generally have a smaller impact on protein function.

[0123] The term “percentage of nucleic acid sequence identity / homology / similarity (%)” for the nucleic acid sequence encoding the monoclonal antibody (antibody construct) identified in this invention is defined as the percentage of nucleotide residues in the candidate sequence that are identical to the nucleotide residues in the coding sequence of the antibody.

[0124] The terms "sequence identity" and "sequence homology" are used interchangeably herein, referring to the degree of sequence similarity on a nucleotide-by-nucleotide or amino acid-by-amino acid basis within a comparison window. The "sequence identity percentage" can be calculated by comparing two optimally aligned sequences within a comparison window, determining the number of positions in both sequences containing the same amino acid residue to obtain the number of matching positions, dividing the number of matching positions by the total number of positions in the comparison window (i.e., the window size), and multiplying the result by 100 to produce the sequence identity percentage. Optimal alignment for determining the sequence identity percentage can be performed in a variety of ways known in the art, for example, using publicly available computer software such as BLAST, BLAST-2, ALIGN, or Megalign (DNASTAR) software. Those skilled in the art can determine suitable parameters for aligning sequences, including any algorithms required to achieve maximum alignment across the full-length sequence being compared or within the target sequence region.

[0125] The term "antibody derivative" according to the invention may also comprise fragments of full-length antibodies, such as VH, VHH, VL, (s)dAb, Fv, light chain (VL-CL), Fd(VH-CH1), heavy chain, Fab, Fab', F(ab')2, or "rIgG" (a "half-antibody" composed of a heavy chain and a light chain). Antibody fragments can be generated by enzymatic or chemical cleavage of a complete antibody. Antibody constructs according to the invention may also comprise modified fragments of the antibody, also referred to as antibody variants or antibody derivatives.

[0126] The terms “binding domain” or “domain that binds to” or “antigen-binding fragment” are used interchangeably in this application to characterize a domain of the antibody of the present invention that specifically binds to / interacts with / recognizes a protein / epitope on a target or antigen (here: MUC17) that is recognized by a parent antibody derived from that domain. The structure and function of the binding domain are based on the structure and / or function of the antibody. A “binding domain” or “domain that binds to” may include the minimum structural requirements of an antibody that allows for immune-specific binding to a target. Such minimum structural requirements for a binding domain may be defined, for example, by the presence of at least three light chain CDRs (i.e., CDR1, CDR2, and CDR3 of the light chain variable region) and / or three heavy chain CDRs (i.e., CDR1, CDR2, and CDR3 of the heavy chain variable region), preferably all six CDRs. A “binding domain” (or “binding domain” or “antigen-binding fragment”) can typically contain both a variable region (VL) of the antibody light chain and a variable region (VH) of the antibody heavy chain; however, it does not have to contain both, but may contain only one of VH or VL.

[0127] The terms “binds (specifically or immune-specifically) with,” “recognizes (specifically or immune-specifically) with,” or “reacts (specifically or immune-specifically) with,” according to the present invention, mean that an antibody or its antigen-binding fragment interacts or (immuno-)specifically with a given amino acid residue / epitope on a target molecule (antigen) (MUC17). Such interaction or binding occurs more frequently and rapidly at amino acid residues / epitopes on a specific target than in alternative substances (non-target molecules), has a longer duration, greater affinity, or a combination thereof. However, due to sequence similarity between homologous proteins in different species, an antibody or binding domain that binds immune-specifically to its target (e.g., a human target) may cross-react with homologous target molecules from different species (e.g., from non-human primates, such as cynomolgus monkeys). Therefore, the term “specific / immunospecific binding” can include the binding of an antibody or binding domain to proteins / epitopes or structure-related proteins / epitopes in more than one species.

[0128] In this invention, the term "epitope" refers to a portion or region of an antigen that is recognized / immunely specifically by a binding domain, antibody, or its derivative. An "epitope" is antigenic, and therefore the term is sometimes also referred to as an "antigen structure" or "antigen determinant." The portion of the binding domain, antibody, or antibody construct that binds to the epitope is called a paratope. Specific binding is believed to be achieved through a specific motif in the amino acid sequence of the binding domain, antibody, or antibody construct and the antigen. Thus, binding is achieved as a result of their primary, secondary, and / or tertiary structures and potential secondary modifications of said structures. Specific interactions between the paratope and its antigenic determinant can lead to simple binding of said site to the antigen. In some cases, specific interactions can alternatively or additionally lead to signal initiation, for example, due to induced changes in antigen conformation, antigen oligomerization, etc.

[0129] The interaction between a monoclonal antibody and the amino acid residues / epitopes of the target antigen means that the variable region exhibits a significant or marked affinity for the epitope / target antigen (MUC17) and generally does not exhibit a significant affinity for proteins or antigens other than the target antigen. "Significant affinity" includes an antibody or its antigen-binding fragment having a KD (dissociation constant) of approximately ≤10. -6 The affinity of M for binding antigen. Preferably, the binding affinity, characterized by KD, is about ≤10. -7 M, ≤10 -8 M, ≤10 -9 M, or ≤10 -10In case M, the binding of antigen and antibody is considered specific. Therefore, the affinity (KD) of the monoclonal antibody (or antibody construct) of this invention for MUC17 is approximately ≤10. -7 M, ≤10 -8 M, ≤10 -9 M, or ≤10 -10 M. These values ​​are preferably measured in surface plasmon resonance determinations, such as Biacore determinations.

[0130] The test determines whether an antibody reacts or binds (immunospecifically) to a target, for example, by comparing the antibody's affinity for its target protein or antigen with its affinity for a non-target protein or antigen (here: proteins other than MUC17). Preferably, the antibodies of the present invention do not bind significantly to proteins or antigens other than MUC17 unless one or more other binding domains targeting another target are intentionally introduced into the antibody / antibody construct of the present invention. The term "not significantly binding" means that the monoclonal antibody (or antibody construct) of the present invention does not bind to proteins or antigens other than MUC17. Thus, the antibody construct exhibits ≤30%, preferably ≤20%, more preferably ≤10%, particularly preferably ≤9%, ≤8%, ≤7%, ≤6%, ≤5%, ≤4%, ≤3%, ≤2%, or ≤1% reactivity with proteins or antigens other than MUC17.

[0131] The antibodies of this invention can be "in vitro generated antibodies" and / or "recombinant antibodies". In this invention, the term "in vitro generated" refers to an antibody as defined above, wherein all or part of the variable region is generated in a non-immune cell selection process.

[0132] The antibodies of this invention may be "isolated" or "substantially pure" antibodies. When used to describe the antibodies of this invention, "isolated" or "substantially pure" means that the antibody has been identified, isolated, and / or recovered from the components of its production environment.

[0133] In some implementations, the antibody is derived from mice. The term "antibody" includes, respectively, an antibody having antibody-derived regions and a binding domain, such as variable and constant regions or domains that substantially correspond to germline immunoglobulin sequences known in the art.

[0134] Whether an antibody competes with another given antibody to bind to an antigen can be measured in competitive assays such as competitive ELISA.

[0135] Covalent modification of the antibodies of this invention is also included within the scope of this invention, and is generally, but not always, performed post-translational.

[0136] The "sample" referred to in this invention can be a biological sample. In some embodiments, the sample is a human sample, such as a human biological sample. The biological sample can be a (human) serum sample, plasma sample, blood sample, bone marrow sample, or tissue sample. The sample can also be a supernatant obtained from a cell culture of (human) bone marrow mononuclear cells or (human) peripheral blood mononuclear cells. The sample can be obtained from a subject, such as a subject suspected of having or diagnosed with a disease associated with abnormal or increased MUC17 expression, or a subject who has received treatment for a disease associated with abnormal or increased MUC17 expression.

[0137] The terms “reference sample,” “reference cell,” “reference tissue,” “control sample,” “control cell,” or “control tissue” refer to a sample, cell, tissue, or standard used for comparative purposes. In one embodiment, the reference sample, reference cell, reference tissue, control sample, control cell, or control tissue is obtained from the healthy and / or disease-free portion of the same subject or individual, and is healthy and / or disease-free tissue or cells. In yet another embodiment, the reference sample, reference cell, reference tissue, control sample, control cell, or control tissue is obtained from healthy tissue or cells of an individual who is not a subject.

[0138] According to the present invention, monoclonal antibodies are conjugated to detectable tags. In some embodiments, covalent modification of the monoclonal antibodies of the present invention includes the addition of one or more tags, such as detection tags. The tags or tag groups can be conjugated to the antibody via spacer arms of various lengths to reduce potential steric hindrance. Various methods for tagging proteins are known in the art and can be used in carrying out the present invention.

[0139] The term “marker” or “marker group” refers to any detectable mark.

[0140] This invention further provides a polynucleotide / nucleic acid molecule encoding the antibody of the present invention. Nucleic acid molecules are biopolymers composed of nucleotides. Polynucleotides are biopolymers composed of 13 or more nucleotide monomers covalently bonded in a chain. DNA (such as cDNA) and RNA (such as mRNA) are examples of polynucleotide / nucleic acid molecules with different biological functions. Nucleotides are organic molecules that act as monomers or subunits of nucleic acid molecules such as DNA or RNA. The nucleic acid molecule or polynucleotide of the present invention can be double-stranded or single-stranded, linear or circular. It is envisioned that the nucleic acid molecule or polynucleotide is contained in a vector. This vector is contained in a host cell. The host cell, for example, can express the antibody after transformation or transfection with the vector or polynucleotide / nucleic acid molecule of the present invention. For this purpose, the polynucleotide or nucleic acid molecule is operatively linked to a control sequence.

[0141] In some embodiments, the polynucleotide / nucleic acid molecule encoding the antibody of the present invention is in the form of a single molecule or two or more separate molecules. If the antibody construct of the present invention is single-chain, the polynucleotide / nucleic acid molecule encoding such a construct will most likely also be in the form of a single molecule. If the different components of the antibody (such as the heavy chain and light chain) are located on separate polypeptide chains, in this case, the polynucleotide / nucleic acid molecule is most likely to be in the form of two (or more) separate molecules.

[0142] This invention also provides vectors comprising the polynucleotide / nucleic acid molecules of this invention. A vector is a nucleic acid molecule used as a medium for transferring (foreign) genetic material into cells, typically for replication and / or expression. The term "vector" encompasses, but is not limited to, plasmids, viruses, granules, and artificial chromosomes. Some vectors are specifically designed for cloning (cloning vectors), while others are designed for protein expression (expression vectors). So-called transcription vectors are primarily used to amplify their inserts.

[0143] This invention provides host cells obtained by transformation or transfection with the polynucleotide / nucleic acid molecules or vectors of this invention. As used herein, the terms "host cell" or "recipient cell" are intended to include any single cell or cell culture that may be, or is already, a receptor for a vector, exogenous nucleic acid molecule, and / or a polynucleotide encoding an antibody of this invention, and / or a receptor for the antibody itself. The corresponding material may be introduced into cells via transformation, transfection, etc. The term "host cell" is also intended to include the progeny or potential progeny of a single cell.

[0144] The present invention provides a method for generating antibodies of the present invention, the method comprising culturing host cells of the present invention under conditions that allow expression of antibodies of the present invention and recovering the generated antibodies from the culture.

[0145] The term "culture" refers to the in vitro maintenance, differentiation, growth, proliferation, and / or reproduction of cells in a culture medium under suitable conditions. This involves growing and maintaining cells in a cell growth medium at appropriate temperatures and gas mixtures. Culture conditions vary considerably for each cell type. Typical growth conditions are a temperature of approximately 37°C, a CO2 concentration of approximately 5%, and a humidity of approximately 95%. The formulation of growth media can vary, for example, in terms of pH, the concentration of carbon sources (such as glucose), the nature and concentration of growth factors, and the presence of other nutrients (such as amino acids or vitamins). Growth factors used to supplement the culture medium are often derived from animal blood serum, such as fetal bovine serum (FBS), calf serum (FCS), horse serum, and porcine serum. Cells can be grown in suspension or as adherent cultures. Cell lines that have been modified to survive in suspension cultures also exist, allowing them to grow at higher densities than would be permitted under adherent conditions.

[0146] The term “expression” includes any step involved in generating the antibodies of the present invention, including but not limited to transcription, post-transcriptional modification, translation, folding, post-translational modification, targeting of specific subcellular or extracellular sites, and secretion.

[0147] The term "recovery" refers to a series of processes aimed at isolating antibodies from cell cultures. A "recovery" or "purification" process can separate the protein and non-protein fractions of a cell culture and ultimately isolate the desired antibody from all other peptides and proteins. Separation steps typically utilize differences in protein size, physicochemical properties, binding affinity, and biological activity. Preparative purification aims to produce relatively large quantities of purified protein for subsequent use, while analytical purification produces relatively small quantities of protein for various research or analytical purposes. The antibodies of this invention prepared from host cells can be recovered or purified using, for example, hydroxyapatite chromatography, gel electrophoresis, dialysis, and affinity chromatography.

[0148] This invention provides a composition or formulation comprising an antibody of the present invention or comprising an antibody produced by a method according to the present invention. The composition is preferably a diagnostic composition. As used herein, the term "diagnostic composition" refers to a composition suitable for use in a diagnostic kit or detection system. One possible diagnostic composition of the present invention comprises one or more antibodies of the present invention, preferably in an amount suitable for detecting MUC17 in a sample.

[0149] The present invention provides a detection system comprising: a) a first monoclonal antibody (or a derivative thereof) that binds to MUC17, and b) a second monoclonal antibody (or a derivative thereof) that optionally binds to MUC17, wherein the binding of the first monoclonal antibody (or a derivative thereof) to MUC17 occurs in the presence of the second monoclonal antibody (or a derivative thereof) that binds to MUC17.

[0150] A "detection system" is a kit or tool (or diagnostic kit / tool) containing reagents for performing analytical assays. In this invention, the system detects and / or quantifies the presence of MUC17 in a sample (typically a tissue sample). The detection system contains an antibody that binds to MUC17. Typically, the detection system involves using a solid support (such as a microtiter plate or membrane) that acts as a surface to immobilize the antigen to be detected (e.g., in the case of a "direct ELISA") or a (monoclonal) antibody ("capture antibody") that binds to MUC17, or a "second antibody" (e.g., an anti-Fc antibody) that binds to the antibody that binds to MUC17 (capture antibody). Typically, this immobilization occurs nonspecifically (by adsorption onto a surface) or specifically (by capture by an antibody, such as a second antibody). Furthermore, the detection system may contain a (monoclonal) detection antibody that binds to MUC17 (optionally conjugated to an enzyme, a detectable label, or a reporter group), and optionally a second antibody (e.g., an anti-Fc antibody) that binds to the detection antibody and is conjugated to an enzyme, a detectable label, or a reporter group.

[0151] The detection system may be for an ELISA assay, which may be used for the purposes of this invention. An ELISA is used to detect antigens in a sample or to quantify unknown amounts of antigens. These steps may include: providing a surface for binding to a known amount of a “capture antibody.” This binding can occur directly by adsorbing the capture antibody onto the surface, or by a second antibody adsorbed onto the surface and binding to the capture antibody. Any non-specific binding sites on the surface are blocked. A sample containing the antigen is applied to the surface, and the antigen is captured (bound) by the antibody. The plate is washed to remove unbound antigen. A “detection antibody” is added, and the “detection antibody” binds to the antigen. This detection antibody may be coupled (e.g., covalently linked) to an enzyme, a detectable label, or a reporter group. If this is not the case, a second antibody coupled to an enzyme, a detectable label, or a reporter group and binding to the detection antibody is applied. The plate is washed to remove any unbound antibody. A chemical substrate is added, which is converted into a detectable form, such as an optical signal or an electrochemical signal. The absorbance or fluorescence or electrochemical signal of the plate wells or surface is measured to determine the presence and / or amount of the antigen.

[0152] Traditional ELISA typically involves chromogenic reporter molecules and substrates, which produce observable color changes to indicate the presence of antigens. Newer ELISA-like technologies use fluorescence, electrochemiluminescence, and quantitative PCR reporter molecules to generate quantifiable signals.

[0153] This detection system can be used in qualitative or quantitative form. Qualitative results provide a simple positive or negative (yes or no) result for the sample. In quantitative form, the optical density (OD) or electrochemical signal of the sample is compared with a standard curve to obtain the specific content of the analyte.

[0154] The terms "subject," "individual," or "patient" are used interchangeably and refer to mammals. Mammals include, but are not limited to, domesticated animals (e.g., cows, sheep, cats, dogs, and horses), primates (e.g., human and non-human primates such as monkeys), rabbits, and rodents (e.g., mice and rats). Preferably, the subject is a human. In one embodiment, the subject includes both those already suffering from a disease and those with a disease to be prevented.

[0155] The term "diagnosis" or "medical diagnosis" is the process of determining which disorder or condition can explain a subject's symptoms and signs. Typically, one or more diagnostic procedures, such as diagnostic tests or medical tests, are performed in this process.

[0156] The term "monitoring" refers to observing a disease, symptom, or one or more medical parameters over a period of time.

[0157] The term "improvement" refers to any improvement in the disease state of a patient with a disease as specified in this invention, achieved by administering an antibody construct to a patient in need. Such improvement can also be viewed as a slowing or halting of disease progression in the patient.

[0158] The term “prevention” means preventing the onset or recurrence of a patient with a tumor or cancer or metastatic cancer as specified in this invention by administering an antibody construct to a patient in need.

[0159] The term "disease" refers to any condition that would benefit from treatment with the antibody construct or pharmaceutical composition described in this invention.

[0160] The terms "diseases and / or conditions associated with abnormal MUC17 expression," "diseases and / or conditions associated with increased MUC17," and "diseases and / or conditions associated with increased MUC17 expression" are used interchangeably in this application to refer to disease states caused or promoted by abnormal MUC17 expression (e.g., overexpression or aberrant activation). In one specific embodiment of the invention, the term "diseases and / or conditions associated with abnormal MUC17 expression" refers to cancer or tumors associated with abnormal MUC17 expression.

[0161] The term "tumor" can be benign, potentially malignant (precancerous), or malignant. Malignant tumors are commonly referred to as cancer. They typically invade and destroy surrounding tissues and can form metastases, meaning they spread to other parts of the body, tissues, or organs.

[0162] The terms "subject in need" or "patient in need of treatment" include subjects who already have a disease as well as subjects with a disease to be prevented. Subjects in need or "patients" include people and other mammalian subjects who are receiving preventive or therapeutic treatment.

[0163] The term “amount” or “content” for MUC17 may be used interchangeably with the terms “level,” “quantity,” or “concentration” for MUC17. A “predefined value for MUC17 content” may be a predetermined “cutoff value.” This value may, for example, indicate a certain level of MUC17 in a sample that indicates a disease or MUC17-positive tumor associated with abnormal or increased MUC17 expression.

[0164] The term "pharmaceutical composition" refers to a composition suitable for administration to a patient, preferably a human patient. The pharmaceutical composition comprises, preferably, a therapeutically effective amount of one or more antibodies or antigen-binding fragments thereof of the present invention. The composition may be administered to a subject in a suitable dose.

[0165] The term "effective dose" or "effective amount" is defined as an amount sufficient to achieve, or at least partially achieve, the desired effect. Specifically, it refers to the amount or dose of the antibody or composition of the present invention, which, when administered to a patient in a single or multiple doses, produces the intended effect in a patient requiring treatment or prevention. The effective amount can be readily determined by a physician skilled in the art by considering a variety of factors, such as: the species of the mammal; weight, age, and general health condition; the specific disease involved; the degree or severity of the disease; the individual patient's response; the specific antibody administered; the administration modality; the bioavailability characteristics of the administered formulation; the chosen dosing regimen; and the use of any concomitant therapies.

[0166] The term "therapeutic effective dose" is defined as an amount sufficient to cure or at least partially stop the disease and its complications in a patient with an existing disease. Specifically, the term "therapeutic effective dose" refers to the amount that effectively achieves the desired therapeutic outcome at the required dose and for the required duration. Therapeutic effective doses of antibodies, antibody fragments, or compositions can vary depending on various factors such as disease state, individual age, sex, weight, and the ability of the antibody or antibody fraction to elicit the desired response in the individual. A therapeutic effective dose is also a dose in which any toxic or harmful effects of the antibody, antibody fragment, or composition are less than the beneficial therapeutic effect. Relative to an untreated subject, a "therapeutic effective dose" preferably inhibits a measurable parameter (e.g., tumor growth rate, tumor volume, etc.) by at least about 20%, more preferably at least about 40%, even more preferably at least about 50%, 60%, or 70%, and still more preferably at least about 80% or 90%. The ability of a compound to inhibit measurable parameters (e.g., cancer) can be evaluated in animal model systems that predict efficacy in human tumors.

[0167] The term "kit" means two or more components (one of which corresponds to the MUC17 antibody of the present invention or its antigen-binding fragment) packaged together in a container, receiver, or other packaging. A kit can be described as a set of products and / or devices sufficient to achieve a particular objective, and can be sold as a single unit. A kit may include one or more receivers having any suitable shape, size, and material, which contain an appropriate dose of the antibody construct or pharmaceutical composition of the present invention. The kit may additionally include instructions for administering the antibody of the present invention and its antigen-binding fragment, for reconstructing the antibody of the present invention and its antigen-binding fragment, and / or for diluting the antibody of the present invention and its antigen-binding fragment.

[0168] The term “about” or “approximately” means within ±20%, preferably within ±15%, more preferably within ±10%, and most preferably within ±5% of a given value or range.

[0169] When the term “and / or” is used in conjunction with multiple options, it should be understood to mean any one of the options or any combination of two or more of the options.

[0170] As used herein, the terms “comprising” or “including” mean to include the stated elements, integers, or steps, but do not exclude any other elements, integers, or steps. In this document, when the terms “comprising” or “including” are used, unless otherwise specified, they also cover situations consisting of the mentioned elements, integers, or steps. For example, when referring to an antibody variable region “comprising” a specific sequence, it is also intended to cover the antibody variable region consisting of that specific sequence. Attached Figure Description

[0171] Figure 1 The results of verifying the specific recognition of MUC17 by the antibodies Ab-6 and Ab8 of the present invention using a protein immunoblotting method are shown, where bands 5 and 6 show the results of Ab-6, and bands 9 and 10 show the results of Ab-8.

[0172] Figure 2 shows the immunohistochemical (IHC) detection results of the antibodies of the present invention on cells. Figure 2A The results of immunohistochemical staining of the antibody Ab-6 of the present invention in the MUC17 overexpressing cell line CHOK1-HU-positive are shown. Figure 2B The results of IHC staining of the antibody Ab-6 of the present invention in CHOKI-negative cells are shown. Figure 2C The image shows the IHC staining results of the antibody Ab8 of the present invention in the MUC17 overexpressing cell line CHOK1-HU-positive. Figure 2DThe results of IHC staining of the antibody Ab8 of the present invention in CHOKI-negative cells are shown.

[0173] Figure 3 shows the immunohistochemical detection results of the antibody of the present invention on gastric cancer cells. Figure 3A The image shows the IHC staining results of the antibody Ab-6 of this invention on the gastric cancer cell line GSU. Figure 3B The results of IHC staining of the gastric cancer cell line GSU with the antibody Ab8 of the present invention are shown. Figure 3C The image shows the staining of the positively labeled antibody in the gastric cancer cell line GSU.

[0174] Figure 4 shows the immunohistochemical detection results of the antibody of the present invention on gastric cancer cells. Figure 4A The results of IHC staining of the gastric cancer cell line NUGC-4 with the antibody Ab-6 of the present invention are shown. Figure 4B The results of IHC staining of the gastric cancer cell line NUGC-4 with the antibody Ab-8 of the present invention are shown. Figure 4C The results of IHC staining of the gastric cancer cell line NUGC-4 with the positive control antibody are shown.

[0175] Figure 5 shows the immunohistochemical detection results of the antibody of the present invention on a gastric cancer tissue sample. Figure 5A The results of IHC staining of the antibody Ab-6 of the present invention in gastric cancer tissue sample F190061 are shown. Figure 5B The results of IHC staining of the antibody Ab-8 of the present invention in gastric cancer tissue sample F190061 are shown. Figure 5C The results of IHC staining of the antibody Ab-6 of the present invention in gastric cancer tissue sample F190562 are shown. Figure 5D The results of IHC staining of the antibody Ab-8 of the present invention in gastric cancer tissue sample F190562 are shown.

[0176] Figure 6 shows the immunohistochemical detection results of the antibody of the present invention on different cancer PDX samples. Figure 6A The results of IHC staining of the antibody Ab-6 of the present invention in colorectal cancer PDX are shown. Figure 6B The results of IHC staining of the antibody Ab-6 of the present invention in PDX of colon cancer are shown. Figure 6C The results of IHC staining of the antibody Ab-6 of the present invention in rectal cancer PDX are shown.

[0177] Figure 7 shows the immunohistochemical detection results of the antibody of the present invention on different cancer tissue samples. Figure 7A This shows the IHC staining results of Ab-6 antibody in tumor tissue of colorectal cancer sample F190343. Figure 7B The image shows the IHC staining results of Ab-6 antibody in the adjacent normal tissue region of colorectal cancer sample F190343. Figure 7CThis shows the IHC staining results of Ab-6 antibody in tumor tissue from gastric cancer tissue sample F221429. Figure 7D The image shows the IHC staining results of Ab-6 antibody in the adjacent normal tissue region of gastric cancer tissue sample F221429. Figure 7E This shows the IHC staining results of Ab-6 in the tumor tissue of pancreatic cancer tissue sample F232624. Figure 7F This shows the IHC staining results of Ab-6 antibody in the adjacent normal tissue region of pancreatic cancer tissue sample F232624. Detailed Implementation

[0179] The following embodiments further illustrate the present invention; however, it should be understood that the embodiments are described in an illustrative rather than limiting manner, and various modifications can be made by those skilled in the art. Where specific conditions are not specified in the embodiments, conventional conditions or conditions recommended by the manufacturer are followed. Reagents or instruments whose manufacturers are not specified are all commercially available conventional products.

[0180] In some embodiments, the present invention provides a method for preparing an anti-MUC17 antibody, wherein the method includes culturing a host cell containing a nucleic acid encoding an anti-MUC17 antibody or an expression vector containing said nucleic acid under conditions suitable for expressing a nucleic acid encoding said anti-MUC17 antibody, and optionally isolating said anti-MUC17 antibody. In one embodiment, the method further includes recovering the anti-MUC17 antibody from said host cell (or host cell culture medium).

[0181] To recombinantly generate the anti-MUC17 antibody of the present invention, the nucleic acid encoding the anti-MUC17 antibody of the present invention is first isolated, and said nucleic acid is inserted into a vector for further cloning and / or expression in host cells. Such nucleic acids are easily isolated and sequenced using conventional procedures, for example, by using oligonucleotide probes capable of specifically binding to the nucleic acid encoding the anti-MUC17 antibody of the present invention.

[0182] The anti-MUC17 antibody of the present invention, prepared as described herein, can be purified using known prior art techniques such as high-performance liquid chromatography (HPLC), ion-exchange chromatography, gel electrophoresis, affinity chromatography, and size exclusion chromatography. The actual conditions used to purify a specific protein also depend on factors such as net charge, hydrophobicity, and hydrophilicity, which are obvious to those skilled in the art. The purity of the anti-MUC17 antibody of the present invention can be determined by any of a variety of well-known analytical methods, including size exclusion chromatography, gel electrophoresis, and HPLC.

[0183] Example 1: Preparation of anti-MUC17 antibody

[0184] 1.1 Construction and identification of cell lines overexpressing human Mucin17

[0185] Construction of the CHO-K1 (ECACC) cell line overexpressing human Mucin17 (hereinafter referred to as huMucin17-CHO K1): The coding nucleic acid sequence of the extracellular region fragment of human Mucin17 protein (NP_001035194.1) SEQ ID NO:17 was constructed into a proprietary lentiviral vector (Genscript Biotech). Then, the lentiviral plasmid was transfected into HEK293T cells (Clontech, 632180) for viral packaging. After 4 days of culture, the cell culture supernatant was collected. The harvested lentiviral supernatant was used to transfect CHO-K1 cells, using Puromycin as a selection pressure. After 3 days of selection, antigen expression in the cell pool was screened using flow cytometry. After obtaining a positive cell pool, single-cell clones were selected from the positive cell pool using limiting dilution or flow cytometry sorting. After single-clone selection, the cells were cultured for another week for antigen expression identification.

[0186] Flow cytometry identification of antigen expression in the cell line: The above-mentioned cell line in logarithmic growth phase was digested and plated into 96-well plates. After washing with FACS buffer (99% DPBS + 1% FBS), primary antibody Tab 1 (10 ng / ml) was added, and the cells were incubated at 4°C for 60 min. After washing, the prepared fluorescent secondary antibody PE Goat anti-mouse IgG antibody (BioLegend, Cat. NO: 410708) was added, and the cells were incubated at 4°C for 20 min. Finally, the cells were detected by flow cytometry (BD FACS Celesta). The results showed that the huMucin17-CHO K1 cell line with high expression of human Mucin17 was obtained.

[0187] 1.2 Animal Immunization and Hybridoma Cell Screening

[0188] Mucin17-His or Mucin17-Mouse Fc protein antigens, Mucin17 expression plasmid-DNA antigens, and multiple immunogens overexpressing Mucin17 CHO-K1 cells were used, alone or alternately, to immunize mice of different species (Balb / c, SJL, A / J). Depending on the selected antigen, mice were immunized subcutaneously / intraperitoneally or via gene gun, with immunizations every two weeks for a total of 3-5 immunizations. One week after the last immunization, mouse blood was collected for immunotiter testing. Finally, a booster immunization was administered using the same antigens.

[0189] Dilute the antigen to 0.5 µg / mL with coating buffer, mix well, and add 100 µL / well to each well of a microplate. Incubate overnight at 4°C for 16 h. Discard the coating buffer, wash the plate once, and blot dry. Add 150 µL of blocking buffer (1% BSA) to each well and incubate at 37°C (Shanghai Yiheng Science Co., Ltd.) for 1 h. Remove the microplate and discard the blocking buffer. Serially dilute the obtained mouse serum and add 100 µL / well to each well of the microplate. Incubate at 37°C for 1 h. Remove the microplate, discard the internal solution, and wash the plate three times. Add 100 µL of diluted enzyme-labeled secondary antibody to each well and incubate at 37°C for 0.5 h. Remove the microplate, discard the internal solution, wash the plate three times, blot dry, add 100 µL of chromogenic reagent, and react at 25°C for 15 minutes. The reaction was terminated by adding 50 μL of 1M HCl, and the OD reading was taken at 450 nm using a microplate reader (Thermo Fisher Scientific (Shanghai) Instruments Co., Ltd., Multiskan FC). Mice with an immunotiter greater than 512000 were selected for hybridoma fusion.

[0190] Prepare a suspension of myeloma cells (SP2 / 0, Cell Bank of Chinese Academy of Sciences) and calculate the total number of SP2 / 0 cells using a hemocytometer. Prepare a suspension of spleen cells and count the total number of cells using a hemocytometer. Mix myeloma cells and spleen cells at a 1:2 ratio for electrofusion. After cell fusion, mix the fused cells in HAT (Sigma) medium to prepare a cell suspension, and then seed the cells into 96-well cell culture plates using a multichannel pipette. Incubate the fused 96-well cell culture plates in a 5.5% CO2 incubator at 37°C. After hybridoma cell culture, collect the supernatant for maternal clone screening, and screen the protein and cellular levels of antigen-binding ability of the cell culture supernatant.

[0191] After the first round of maternal clone selection, the selected maternal clone cell lines were retained. Cells in the subcloning wells were gently pipetted and mixed. 20 µl of the cell suspension was added to 20 µl of trypan blue solution and mixed well. 20 µl of this solution was then used for hemocytometer counting. Using the limiting dilution method, cells were seeded into 96-well cell culture plates with 1×HT replacing feeder cell culture medium. The plates were labeled and incubated in a CO2 incubator. The subcloned 96-well plates were then incubated at 37°C with 5.5% CO2. After 7 days of culture, the supernatant was used for single-clone selection.

[0192] After subcloning, the selected antibodies were sequenced to obtain several candidate antibodies. Among them, antibodies Ab-6 and Ab-8 showed significant selectivity and specificity for huMUC17, as shown in the flow cytometry results in the table below, and also exhibited good specificity for detecting the protein in tumor tissue. The sequences of the Ab-6 and Ab-8 antibodies are shown in the sequence listing in the specification.

[0193]

[0194] Example 2: Specificity verification of anti-MUC17 antibodies Ab-6 and Ab-8

[0195] This embodiment evaluated the specific binding of several antibodies, including Ab-6 and Ab-8 antibodies, to MUC17.

[0196] Protein sample extraction and preparation: 1. Cell lysis: Collect cells by centrifugation, wash twice with pre-chilled PBS; remove PBS, add pre-chilled lysis buffer (Thermo Fisher, #89900), and add the appropriate amount of lysis buffer RIPA according to the instructions based on the different culture plates and the actual number of cells. Mix well, lyse on ice for 5 min, and repeatedly pipette until the cells are completely lysed; centrifuge at 12000 rpm for 10 min at 4°C, and the supernatant is the protein-containing solution, which should be stored at -80°C.

[0197] 2. Electrophoresis: Proteins in the solution were quantified using a BCA kit (Tiangen, catalog number PA115-01). Sample preparation for electrophoresis was as follows: Add an appropriate amount of 5x loading buffer (weigh 15.1g Tris, 94g Glycine, and 5g SDS; add approximately 800mL of deionized water to a beaker, stir thoroughly to dissolve, and bring the volume to 1L), boil at 100℃ for 10min, and centrifuge at 12000 rpm for 2min. Electrophoresis was performed using standard loading methods: the protein antigen loading volume was 3ug. The SDS-PAGE electrophoresis plate gap thickness was selected according to the sample volume; generally, for a 0.75mm gap 15-well plate, the loading volume was <15uL / well, and for a 1mm gap 10-well plate, the loading volume was <30uL / well. The precast gel was purchased from GenScript, catalog number M00652.

[0198] 3. Transfer and Development: Wet transfer was used. The transfer buffer was pre-cooled on ice. A suitable PVDF membrane was cut according to the size of the SDS-PAGE gel and activated by soaking in methanol. A transfer "sandwich" was prepared by arranging the transfer clip, sponge, filter paper, SDS-PAGE gel, VDF membrane, filter paper, sponge, and transfer clip in that order. The membrane and gel were placed on the positive and negative electrodes respectively in the "sandwich". The pre-cooled transfer buffer (2.9g Glycine, 5.8g Tris, and 0.37g SDS in 800mL water and 200mL methanol) was added. The tank was placed in an ice box to keep it cool, and the entire electrophoresis tank was buried in ice. Electrophoresis was performed at a constant voltage of 90V for 120min. To prevent interference from non-specific proteins on the membrane, the PVDF membrane after transfer was placed in 5% skim milk blocking buffer (TBST, 8.8g NaCl, 20ml 1M Tris-HCl (pH 8.0) in 800mL deionized water, 0.5ml L-ween 20 added, mixed thoroughly, and then diluted to 1L with deionized water). The membrane was blocked on a shaker at room temperature for 1-2 hours. After blocking, the membrane was slowly washed three times with 1×TBST on a shaker for 10 minutes each time. The corresponding primary antibody was diluted with 5% skim milk blocking buffer (TBST) according to the appropriate dilution ratio. The blocked and cleaned membrane was then incubated overnight on a shaker at 4°C. The primary antibody could be recovered to -20°C. The membrane was then slowly washed three times with 1×TBST on a shaker for 10 minutes each time. Select the appropriate species (goat anti-mouse / goat anti-rabbit) secondary antibody (HRP-labeled) based on the species of origin of the primary antibody (mouse / rabbit). Dilute the secondary antibody according to the instructions. Immerse the membrane in the secondary antibody dilution solution and incubate on a shaker at room temperature for 1-2 hours. Wash slowly three times on a 1×TBST shaker for 10 minutes each time. Add an equal volume of ECL developing solution to the membrane, develop it using a chemiluminescence imaging system, and save the image data (ECL kit purchased from Millipore, catalog number WBKLS0100).

[0199] Figure 1 Ab-6 (bands 5 and 6) and Ab8 (bands 9 and 10) showed strong specificity. No fluorescence was observed in the negative cell bands. The intensity of the β-actin (commercially available routine antibody reagent, CST, Lot: 4967S) bands in both positive and negative cell lines was comparable. The other tested antibodies did not show specificity and therefore will not be discussed separately.

[0200] Example 3: Immunohistochemical assay to identify MUC17-expressing positive and negative cell lines

[0201] Immunohistochemistry (IHC) is a technique that uses colorimetric antibodies to locate, qualitatively and quantitatively determine target antigens in tissue cells. IHC has been widely used in tumor pathology diagnosis. By examining the distribution, location, and abundance of tumor biomarkers in clinicopathological samples, IHC can provide information on multiple aspects of tumors, such as stage and grade, benign or malignant nature, and can be used to predict a patient's clinical responsiveness to biomarker-targeted therapy. A key element in this type of pathological diagnosis is the availability of validated and reliable IHC antibodies for detecting biomarker-positive cases.

[0202] When preparing samples for immunohistochemical analysis, some epitopes of antigens expressed on the membrane surface of the sample may be masked or destroyed, preventing some antibodies from recognizing them. Furthermore, even if the epitope remains, different antibodies to the same antigen can exhibit significantly different detection performance due to factors such as the epitope's location and membrane surface binding activity. Therefore, monoclonal antibodies used for IHC often require specialized screening and validation.

[0203] In this invention, the method for preparing IHC sections derived from cells and animal tumors uses conventional paraffin embedding methods, which mainly include conventional tissue fixation, tissue processing, tissue embedding, and tissue sectioning.

[0204] The IHC staining method is as follows: 1. Obtain cell or tissue sections using conventional paraffin embedding method, and bake the sections at 60℃ for 30 min; 2. Staining: Place the baked sections into a Leica Bond III fully automated immunohistochemical staining machine and stain according to the staining program. After staining, remove the slides and immerse them in 95% ethanol, anhydrous ethanol I, anhydrous ethanol II, xylene I, and xylene II for 5 min each; 3. Mounting: Use a dropper to apply an appropriate amount of neutral resin to the tissue section, then gently cover it with a coverslip, avoiding air bubbles. Place the mounted slide in a fume hood to allow the neutral resin to cure naturally; 4. Viewing the slides: Observe the staining results under a microscope and take pictures. The staining program is shown in the table below:

[0205] To determine whether anti-MUC17 antibodies Ab-6 and Ab8 can detect MUC17 in cancer cells, immunohistochemical assays were first performed on CHOK1-HU-positive cells and CHOK1-negative cells (CHO-K1 cell line derived from ECACC) expressing MUC17 using Ab-6, Ab-8, and the positive control Ab122184 (source: Abcam) at an antibody concentration of 10 μg / ml.

[0207] The method for constructing the CHOK1-hu-positive cell line is as described in Example 1. In Example 1, a huMucin17-CHO K1 cell line with high expression of human Mucin17 was obtained. In this example, it is referred to as the CHOK1-HU-positive cell line.

[0208] Figure 2A and 2B The staining of Ab-6 in CHOK1-HU-positive and CHOK1-negative cells is shown, with Ab-6 being positive in CHOK1-HU-positive cells and negative in CHOK1-negative cells.

[0209] Figure 2C and 2D The staining of Ab-8 in CHOK1-HU-positive and CHOK1-negative cells is shown, with Ab-8 being positive in CHOK1-HU-positive cells and negative in CHOK1-negative cells.

[0210] The results showed that Ab-6 and Ab-8 stained highly on CHOK1-HU-positive cells that highly expressed human MUC17, but not on negative cell lines, indicating that the staining of Ab-6 and Ab-8 antibodies is specific and can specifically recognize cells expressing human MUC17.

[0211] Example 4: IHC assay of GSU gastric cancer cell line expressing MUC17

[0212] To clarify whether anti-MUC17 antibodies Ab-6 and Ab-8 can detect MUC17 in cancer cells, this embodiment uses immunohistochemistry (see Example 3 for details): using 10 μg / ml Ab-6 and Ab-8 as detection antibodies and 0.16 μg / ml of commercially available positive antibody Ab122184 (Abcam) as a reference, the expression of MUC17 in gastric cancer cells GSU (source: Riken, RC2278) was detected.

[0213] Figure 3A The image shows Ab-6 antibody staining in the gastric cancer cell line GSU. Figure 3B The image shows Ab-8 antibody staining in the gastric cancer cell line GSU. Figure 3C The results show that the positive marker antibody stained the gastric cancer cell line GSU. Both Ab-6 and Ab-8 can specifically stain MUC17 positive cells, and the positive marker antibody ab122184 was positive as a positive quality control.

[0214] Example 5: IHC assay of NUGC-4, a positive gastric cancer cell line expressing MUC17

[0215] To further clarify the detection efficacy of anti-MUC17 antibodies Ab-6 and Ab-8 against MUC17 in cancer cells, this embodiment uses immunohistochemistry (see Example 3 for details): using 10 μg / ml antibodies Ab-6 and Ab-8 as detection antibodies and 0.16 μg / ml ab122184 as a reference, the expression of MUC17 in gastric cancer cells NUGC-4 (source: Nanjing Kebai Biotechnology Co., Ltd.) was detected.

[0216] Figure 4A The image shows Ab-6 antibody staining in the gastric cancer cell line NUGC-4. Figure 4B The staining of Ab-8 antibody in the gastric cancer cell line NUGC-4 is shown. Figure 4C The results show the staining of positive marker antibodies in the gastric cancer cell line NUGC-4. Both Ab-6 and Ab-8 can specifically stain in MUC17-positive cells, and the positive marker antibody Ab122184 was positive as a positive quality control.

[0217] Experimental Example 6: IHC determination of MUC17 expression in PDX models of colorectal cancer, colon cancer, and rectal cancer

[0218] In this embodiment, immunohistochemistry (see Example 3 for specific methods) was used to detect the detection efficacy of anti-MUC17 antibodies Ab-6 and Ab-8 against MUC17 in different PDX models.

[0219] The expression of MUC17 in colorectal cancer PDX models, colon cancer PDX models, and rectal cancer PDX models (source: Shanghai Lidi Biotechnology Co., Ltd.) was detected using 0.1 μg / ml Ab-6.

[0220] Figure 6A The results of Ab-6 staining in PDX (LD1-2012-362829) of colorectal cancer are shown. Figure 6B The results of Ab-6 staining in PDX (F22-1929) colon cancer are shown. Figure 6C The results of Ab-6 staining in a rectal cancer PDX model (F22-0288) are shown.

[0221] The results showed that Ab-6 could specifically recognize MUC17 expressed in various colorectal cancer models.

[0222] Experimental Example 7: Immunohistochemical identification of MUC17 expression in gastric cancer tissue samples by antibody

[0223] In this embodiment, immunohistochemistry (see Example 3 for specific methods) was used to detect the detection efficacy of anti-MUC17 antibodies Ab-6 and Ab-8 in gastric cancer tissue samples.

[0224] Immunohistochemical assays were performed on MUC17-positive gastric cancer tissue samples using Ab-6 and Ab-8 at antibody concentrations of 0.5 μg / ml.

[0225] Figure 5A The image shows staining of Ab-6 antibody in a gastric cancer tissue sample F190061. Figure 5B This shows the staining of Ab-8 in the gastric cancer tissue sample.

[0226] Figure 5C This shows the staining of Ab-6 antibody in another gastric cancer tissue sample, F190562. Figure 5D This shows Ab-8 staining in the same gastric cancer tissue sample.

[0227] The results showed that Ab-6 and Ab-8 antibodies could stain gastric cancer tissue samples and specifically recognize MUC17.

[0228] Experimental Example 8: Specificity identification of antibodies against various cancer tissue samples

[0229] This embodiment uses immunohistochemistry (see Example 3 for the specific method) to detect the specificity of the exemplary antibody of the present invention for recognizing a variety of cancer samples.

[0230] Immunohistochemical staining was performed on tissue samples of MUC17-positive colorectal cancer, gastric cancer, and pancreatic cancer using Ab-6 at an antibody concentration of 0.2–0.5 μg / ml.

[0231] Figure 7A This demonstrates the specific staining of Ab-6 antibody in a colorectal cancer tissue sample F190343. Figure 7B This shows the parallel staining results of Ab-6 antibody on adjacent normal tissue areas in the same colorectal cancer sample; Figure 7C The image shows staining of Ab-6 antibody in a gastric cancer tissue sample F221429. Figure 7D The results of parallel staining of adjacent normal tissue areas with Ab-6 antibody in the same gastric cancer sample are shown. Figure 7E The image shows staining of Ab-6 antibody in a pancreatic cancer tissue sample F232624. Figure 7F This shows the parallel staining results of Ab-6 antibody on adjacent normal tissue areas in the same pancreatic cancer sample.

[0232] The results showed that Ab-6 could stain in a variety of cancer tissue samples and identify abnormally expressed MUC17 in the cancer tissue, while it did not specifically stain adjacent normal tissue. This indicates that the antibody of the present invention has excellent tumor tissue specificity recognition and provides a highly specific tool for the pathological detection and diagnosis of MUC17 positive cancer.

[0233] sequence list

Claims

1. An antibody or its antigen-binding fragment that binds to human MUC17 binds to the MUC17 protein expressed on the cell surface, wherein, The antibody or its antigen-binding fragment comprises a heavy chain variable region (VH) and a light chain variable region (VL), wherein the heavy chain variable region contains HCDR1 of the sequence SEQ ID NO: 3 or SEQ ID NO: 11, HCDR2 of the sequence SEQ ID NO: 4 or SEQ ID NO: 12, and HCDR3 of the sequence SEQ ID NO: 5 or SEQ ID NO: 13, and the light chain variable region contains LCDR1 of the sequence SEQ ID NO: 6 or SEQ ID NO: 14, LCDR2 of the sequence SEQ ID NO: 7 or SEQ ID NO: 15, and LCDR3 of the sequence SEQ ID NO: 8 or SEQ ID NO:

16.

2. The antibody or its antigen-binding fragment according to claim 1, wherein, 1) The heavy chain variable region contains HCDR1 of sequence SEQ ID NO: 3, HCDR2 of sequence SEQ ID NO: 4, and HCDR3 of sequence SEQ ID NO: 5; the light chain variable region contains LCDR1 of sequence SEQ ID NO: 6, LCDR2 of sequence SEQ ID NO: 7, and LCDR3 of sequence SEQ ID NO: 8; or, 2) The heavy chain variable region contains HCDR1 of SEQ ID NO: 11, HCDR2 of SEQ ID NO: 12, and HCDR3 of SEQ ID NO: 13, and the light chain variable region contains LCDR1 of SEQ ID NO: 14, LCDR2 of SEQ ID NO: 15, and LCDR3 of SEQ ID NO:

16.

3. The antibody or antigen-binding fragment thereof according to claim 1 or 2, wherein the antibody or antigen-binding fragment thereof comprises VH and VL, wherein: 1) The VH comprises the amino acid sequence shown in SEQ ID NO: 1, or comprises an amino acid sequence having at least 90% identity with SEQ ID NO: 1, or is composed of SEQ ID NO: 1; the VL comprises the amino acid sequence shown in SEQ ID NO: 2, or comprises an amino acid sequence having at least 90% identity with SEQ ID NO: 2, or is composed of SEQ ID NO: 2; or 2) The VH contains an amino acid sequence as shown in SEQ ID NO: 9, or contains an amino acid sequence that is at least 90% identical to SEQ ID NO: 9, or is composed of SEQ ID NO: 9, and the VL contains an amino acid sequence as shown in SEQ ID NO: 10, or contains an amino acid sequence that is at least 90% identical to SEQ ID NO: 10, or is composed of SEQ ID NO:

10.

4. The antibody or antigen-binding fragment thereof according to any one of claims 1 to 3, wherein the antibody is a monoclonal antibody.

5. The antibody or antigen-binding fragment thereof according to any one of claims 1 to 4, wherein the antigen-binding fragment is selected from Fab, Fab'-SH, Fv, scFv or (Fab')2 fragment.

6. The antibody or antigen-binding fragment thereof according to any one of claims 1 to 5, wherein the antibody or antigen-binding fragment thereof further comprises a heavy chain constant region and a light chain constant region, preferably, the heavy chain constant region and the light chain constant region are derived from the constant regions of rodent germline immunoglobulin sequences.

7. The antibody or antigen-binding fragment thereof according to claim 6, wherein the heavy chain constant region comprises the amino acid sequence shown in SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24 or SEQ ID NO:25, or comprises an amino acid sequence having at least 90% identity with SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24 or SEQ ID NO:25, or is composed of SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24 or SEQ ID NO:25, and the light chain constant region comprises the amino acid sequence shown in SEQ ID NO:26, or comprises an amino acid sequence having at least 90% identity with SEQ ID NO:26, or is composed of SEQ ID NO:

26.

8. The antibody or antigen-binding fragment thereof according to any one of claims 1 to 7, comprising a heavy chain and a light chain, wherein: 1) The heavy chain comprises the amino acid sequence shown in SEQ ID NO:18, or comprises an amino acid sequence having at least 90% identity with SEQ ID NO:18, or is composed of SEQ ID NO:18; the light chain comprises the amino acid sequence shown in SEQ ID NO:19, or comprises an amino acid sequence having at least 90% identity with SEQ ID NO:19, or is composed of SEQ ID NO:19; or 2) The heavy chain contains the amino acid sequence shown in SEQ ID NO:20, or contains an amino acid sequence that has at least 90% identity with SEQ ID NO:20, or is composed of SEQ ID NO:20, and the light chain contains the amino acid sequence shown in SEQ ID NO:21, or contains an amino acid sequence that has at least 90% identity with SEQ ID NO:21, or is composed of SEQ ID NO:

21.

9. A polynucleotide molecule encoding an antibody or an antigen-binding fragment thereof according to any one of claims 1 to 8.

10. An expression vector comprising the polynucleotide molecule according to claim 9.

11. A host cell comprising the polynucleotide molecule of claim 9 or the expression vector of claim 10.

12. A method for producing an anti-MUC17 antibody or an antigen-binding fragment thereof, comprising culturing a host cell according to claim 11 under conditions that allow expression of said antibody or an antigen-binding fragment thereof, and recovering the resulting antibody or antigen-binding fragment thereof from the culture.

13. A kit comprising an antibody or an antigen-binding fragment thereof as described in any one of claims 1 to 8, or comprising an antibody or an antigen-binding fragment thereof produced by the method according to claim 12.

14. A composition comprising the antibody or antigen-binding fragment thereof of any one of claims 1 to 8, or comprising the antibody or antigen-binding fragment thereof produced by the method according to claim 12.

15. A detection system comprising an antibody or antigen-binding fragment thereof according to any one of claims 1 to 8, or comprising an antibody or antigen-binding fragment thereof produced by the method according to claim 12, a kit according to claim 13, or a composition according to claim 14.

16. The use of the antibody or antigen-binding fragment thereof according to any one of claims 1 to 8, the antibody or antigen-binding fragment thereof produced by the method according to claim 12, the kit according to claim 13, the composition according to claim 14, or the detection system according to claim 15 in the preparation of a product, wherein the product is used for: 1) Identify cells expressing MUC17; 2) Detect whether MUC17 is expressed in the sample; or 3) Quantitative analysis of MUC17 expression in samples.

17. The use of the antibody or antigen-binding fragment thereof according to any one of claims 1 to 8, the antibody or antigen-binding fragment thereof produced by the method according to claim 12, the kit according to claim 13, the composition according to claim 14, or the detection system according to claim 15 in the preparation of a diagnostic reagent or diagnostic product, wherein the diagnostic reagent or diagnostic product is used for: 1) To identify subjects who have or are at risk of having diseases associated with abnormal or increased MUC17 expression; 2) Diagnose diseases associated with abnormal or increased MUC17 expression; 3) Monitor the progression of diseases associated with abnormal MUC17 expression or increased MUC17 expression; 4) Stratify patients diagnosed with diseases associated with abnormal or increased MUC17 expression; 5) Monitor subjects’ response to treatment for diseases associated with abnormal MUC17 expression or increased MUC17 expression.

18. A method for identifying or detecting whether MUC17 is expressed in a sample, or for quantifying the expression of MUC17 in a sample, comprising: 1) Contact the sample with the antibody or antigen-binding fragment thereof according to any one of claims 1 to 8, the antibody or antigen-binding fragment thereof produced by the method according to claim 12, the kit according to claim 13, the composition according to claim 14, or the detection system according to claim 15. 2) Detect the presence of a complex formed by the antibody or its antigen-binding fragment and MUC17, and / or determine the expression level of MUC17 in the sample; Optionally, the antibody is directly or indirectly labeled with a detectable marker. Optionally, the detection is a quantitative detection.

19. The method of claim 18, wherein the sample is a solid tissue sample, a liquid tissue sample, a sample containing cultured cells, preferably a cell sample, a tissue sample (e.g., tissue sections and tissue microarrays), preferably, the sample is a tissue section (e.g., paraffin section or frozen section), more preferably, the sample is a formaldehyde-fixed paraffin-embedded (FFPE) section, and even more preferably, the sample is a sample from a human subject.

20. A method for diagnosing whether a subject suffers from a disease associated with abnormal or increased MUC17 expression, comprising: (a) Using the antibody or antigen-binding fragment thereof according to any one of claims 1 to 8, the antibody or antigen-binding fragment thereof produced by the method according to claim 12, the kit according to claim 13, the composition according to claim 14, or the detection system according to claim 15, to detect the expression level of MUC17 and / or thereof in a biological sample from a subject; (b) Compare the MUC17 expression level determined in step (a) with the following: (i) A predefined cutoff value indicating the level of MUC17 expression in which the disease is absent; (ii) The level of MUC17 expression determined in healthy individual control samples. (iii) The level of MUC17 expression determined in negative control samples, representing the absence of this neoplastic disease, or (iv) The level of MUC17 expression determined in samples obtained from the same source or subject at a previous time point. If the MUC17 expression level determined in step (a) is higher than the MUC17 expression level determined in any of (i)-(iv), it indicates that the subject has a disease associated with abnormal or increased MUC17 expression.

21. A method for monitoring the progression of a disease associated with abnormal MUC17 expression in a subject, or a method for monitoring a subject's treatment response to a disease associated with abnormal MUC17 expression or increased MUC17 expression, comprising: (a) Using the antibody or antigen-binding fragment thereof according to any one of claims 1 to 8, the antibody or antigen-binding fragment thereof produced by the method according to claim 12, the kit according to claim 13, the composition according to claim 14, or the detection system according to claim 15, to detect the level of MUC17 expression in a sample obtained from a subject at a first time point; (b) Detect the expression level of MUC17 in samples obtained from the subject at a second time point or after treatment; and (c) Compare the MUC17 content determined in step (a) with the MUC17 content determined in step (b). If the MUC17 content determined in step (b) is lower than the MUC17 content determined in step (a), it indicates that the progression of the disease associated with abnormal MUC17 expression or increased MUC17 expression in the subject has been controlled or alleviated, or that the subject has responded to treatment for the disease associated with abnormal MUC17 expression or increased MUC17 expression.

22. The method of claim 20 or 21, wherein the sample is a solid tissue sample and / or a liquid tissue sample, a sample containing cultured cells, preferably a cell sample, a tissue sample (e.g., tissue sections and tissue microarrays), preferably, the sample is a tissue section (e.g., paraffin section or frozen section), more preferably, the sample is a formaldehyde-fixed paraffin-embedded (FFPE) section, and more preferably, the sample is from a human subject suspected of having or having a disease associated with abnormal or increased MUC17 expression, or from a human subject who has received treatment for a disease associated with abnormal or increased MUC17 expression.

23. The use according to claim 17 or the method according to any one of claims 20 to 22, wherein the disease associated with abnormal or increased MUC17 expression is selected from: gastric cancer, esophageal cancer, intestinal cancer, gastroesophageal cancer, gastroesophageal junction adenocarcinoma, colorectal cancer, pancreatic cancer, breast cancer, liver cancer, ovarian cancer, and bile duct cancer.

24. The method of any one of claims 17-21, wherein the detection is performed by immunohistochemical (IHC) staining.

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