Targeted GUCY2C chimeric antigen receptor with gastrointestinal homing effect and application thereof

Abstract

The invention relates to the technical field of bioengineering, in particular to a targeted GUCY2C chimeric antigen receptor with a gastrointestinal homing effect and application of the targeted GUCY2C chimeric antigen receptor. The invention provides a GUCY2C targeting chimeric antigen receptor with a gastrointestinal homing effect. The GUCY2C targeting chimeric antigen receptor comprises a signal peptide, a light chain of an anti-GUCY2C single-chain antibody, a heavy chain of the anti-GUCY2C single-chain antibody, a G4S linker, StrepII-G4S, a CD8 alpha hinge region, a 4-1BB intracellular region, a CD3 zeta intracellular region, a TLR5TIR sequence, T2A and a CCR9 chemotactic factor. The invention solves the problem of poor solid tumor effect of the existing CAR-T therapy, has wide application prospect and huge market value, and provides a method for treating cancer patients with cancer cells expressing GUCY2C and a method for preventing cancer with cancer cells expressing GUCY2C in patients identified as having increased risk.

Description

Technical Field

[0001] The present invention relates to the field of bioengineering technology, and particularly relates to a gut-homing effect-targeted GUCY2C chimeric antigen receptor and its application. Background Art

[0002] Gastrointestinal cancers (GIC) include gastric cancer, colorectal cancer, pancreatic cancer, esophageal cancer, etc. There are approximately 5 million new cases of GIC worldwide every year. GIC-related deaths account for 35% of all cancer-related deaths, and more than one-third of newly diagnosed gastrointestinal tumors globally occur in China. Colorectal cancer is a malignant lesion that occurs in the colonic mucosal epithelium under the action of various carcinogenic factors such as environment or genetics, and is one of the common malignant tumors. In recent years, the incidence and fatality rate of GIC in China have been on the rise. Currently, the traditional therapies for GIC include surgery, chemotherapy, radiotherapy, traditional Chinese medicine treatment, and targeted therapy. However, the overall survival rate of metastatic GIC patients remains very low, seriously affecting the quality of life of advanced patients. Immunotherapy is gradually playing an increasingly important role in colorectal cancer, but there are still many problems that lead to unsatisfactory curative effects, such as drugs being difficult to reach the tumor or enter the tumor, and the decreased expression of major histocompatibility complex (MHC) on the surface of tumor cells, which is prone to immune escape, etc.

[0003] Chimeric antigen receptor T cell (CAR-T) therapy is a new and relatively mature tumor immunotherapy. In vitro gene editing technology endows T cells with specific recognition of tumor antigens and powerful immune killing functions, enabling it to achieve remarkable results in clearing blood tumors. However, the curative effect of CAR-T cells on solid tumors is currently limited. The reasons hindering the successful treatment of solid tumors by CAR-T are multiple. For example, the lack of specific tumor antigens, tumor heterogeneity, or antigen loss can make CAR-T inefficient or dysfunctional; the hypoxic and immunosuppressive microenvironment of solid tumors, as well as the inability of T cells to migrate and infiltrate into solid tumors, etc., can all affect the treatment effect of CAR-T. Currently, the mainstream tumor antigens used in CAR-T therapy for GIC treatment mainly include CEA, HER2, EpCAM, etc. Although these molecules are highly expressed in epithelial cell tumors, they are also present in most epithelial cells, such as gastrointestinal, breast, liver, lung epithelial cells, etc., and are prone to cause systemic side effects; in addition, the traditional CAR structure design is relatively simple, and the commonly used intracellular domains such as CD28 and 4-1BB cannot effectively extend the functions of CAR-T in solid tumors, such as proliferation, migration, and infiltration, greatly limiting the application effect of CAR-T cell therapy in GIC treatment.

[0004] Guanylate cyclase C (abbreviated as GCC, or GUCY2C) is a membrane-bound receptor that, when activated by the hormone ligands guanylin or uroguanylin, induces the production of the second messenger cGMP, thereby regulating physiological processes such as intestinal homeostasis, tumorigenesis, and obesity. GUCY2C expression is restricted to the luminal surface of the intestinal epithelium and a subset of hypothalamic neurons. It is continuously expressed in more than 95% of colorectal cancer metastases and is ectopically expressed in tumors caused by intestinal metaplasia, including esophageal cancer, gastric cancer, oral cancer, salivary gland cancer, and pancreatic cancer. Due to the subcellular restriction of GUCY2C, it cannot reach the apical membrane of polarized epithelial tissues, which creates a therapeutic opportunity for targeting metastatic lesions of colorectal origin that have lost apical-basolateral polarization without concomitant systemic side effects. Researchers such as Snook have demonstrated in human xenograft models of immunodeficient mice that CAR-T cells targeting human GUCY2C can effectively inhibit the growth of metastatic colorectal cancer without intestinal toxicity. To date, there are only three CAR-T clinical studies targeting the GUCY2C target on the NIH clinical trials website, including phase I or pre-phase I clinical trials jointly carried out by Beijing Emmanet and Beijing Cancer Hospital, and Shanghai Stemirna Therapeutics jointly with the First Affiliated Hospital of Zhejiang University and Anhui Provincial Cancer Hospital. Among them, Shanghai Stemirna Therapeutics announced the results of the phase I clinical dose escalation trial of the novel therapy GCC19CART for the treatment of relapsed / refractory colorectal cancer at the 2022 ASCO annual meeting, showing good efficacy and safety.

[0005] Toll-like receptors (TLRs) have a TIR domain, a transmembrane domain, and leucine-rich repeats (LRRs). The signal cascades they regulate play a key role in the human immune system. The effect of TLRs on T cells has been shown to be an effective co-stimulatory signal and a promising strategy to improve the efficacy of cancer immunotherapy. TLR signaling is involved in enhancing cell division, survival, and cytotoxicity of CD4+ T or CD8+ T cells. In addition, the co-stimulatory effect of TLR signal transduction is also related to the ability to lower the activation threshold of TCR for weakly immunogenic tumor antigens. The innate immune receptor Toll-like receptor 2 (TLR2) is highly expressed on activated and memory T cells. The TLR2 signaling pathway can increase T cell proliferation and cytokine production, lower the activation threshold of co-stimulatory signals transmitted by antigen-presenting cells (APCs), promote the generation of memory CD8 + T cells, and directly trigger Th1 effector functions. In addition, TLR2-mediated signal transduction can eliminate the inhibitory ability of Treg cells. Studies have shown that CAR-T cells introducing the TIR domain of TLR2 can enhance the anti-tumor efficacy against leukemia and solid tumors.

[0006] Toll-like receptor 5 (TLR5) is a member of the Toll-like receptor family and is one of the most characteristic pattern recognition receptor molecules. It can recruit a variety of ligand proteins, stimulate signal transduction, and lead to the activation of some specific transcription factors. Studies have shown that the binding of TLR5 to dendritic cells can promote the stimulation of innate lymphocytes and T helper cells. The TLR5 ligand flagellin has been reported to regulate the activity of human Tregs. TLR5 is expressed on the surface of effector T cells and Tregs. High concentrations of flagellin co-stimulate the proliferation of TCR-stimulated responder T cells, while low concentrations of flagellin cause an increase in FOXP3 and the inhibitory ability of Tregs. Since naive human CD8 + T cells uniquely express functional TLR5, and existing studies have pointed out that it can act as an extremely effective co-stimulatory receptor.

[0007] Chemokine receptor 9 (CCR9) is an intestinal trophic chemokine receptor expressed on lymphocytes and dendritic cells. Chemokine CCL25 is the only ligand of CCR9, which is selectively expressed by myeloid dendritic cells and cortical epithelial cells in the thymus and small intestinal epithelium under normal circumstances. The CCL25 / CCR9 interaction is the key for CCR9 + T cell progenitors to home to the thymus. In addition to being an intestinal homing molecule, CCR9 inhibits Treg polarization, thereby inhibiting the development of Treg cells. Therefore, the CCL25 / CCR9 interaction plays an important role in the pathogenesis of inflammatory bowel disease. In addition, CCR9 + T cells have the potential to enhance activation and produce pro-inflammatory cytokines, and CCR9 + T helper cells can also promote the proliferation and survival of CD8 + T cells. Summary of the Invention

[0008] Aiming at the deficiencies of the prior art, the purpose of the present invention is to provide a gastrointestinal homing effect-targeted GUCY2C chimeric antigen receptor and its application, to solve the problem of poor efficacy of existing CAR-T therapies in treating solid tumors, and it has broad application prospects and great market value. The present invention provides methods for treating cancer patients with high expression of GUCY2C and methods for preventing diseases specifically expressing GUCY2C.

[0009] To achieve the object of the present invention, the present invention adopts the following technical solutions: A chimeric antigen receptor targeting GUCY2C with gastrointestinal homing effect, comprising a signal peptide, a light chain of a single-chain antibody against GUCY2C, a heavy chain of a single-chain antibody against GUCY2C, a G4S linker, StrepII-G4S, a CD8α hinge region, a 4-1BB intracellular region, a CD3ζ intracellular region, a TLR5 TIR sequence, T2A, and a CCR9 chemokine.

[0010] In some embodiments of the present invention, the amino acid sequence of the signal peptide is as shown in SEQ ID NO:1, the amino acid sequence of the light chain of the single-chain antibody against GUCY2C is as shown in SEQ ID NO:2, the amino acid sequence of the heavy chain of the single-chain antibody against GUCY2C is as shown in SEQ ID NO:3, the amino acid sequence of the G4S linker is as shown in SEQ ID NO:4, the amino acid sequence of StrepII-G4S is as shown in SEQ ID NO:6, the amino acid sequence of the CD8α hinge region is as shown in SEQ ID NO:7, the amino acid sequence of the 4-1BB intracellular region is as shown in SEQ ID NO:8, the amino acid sequence of the CD3ζ intracellular region is as shown in SEQ ID NO:9, the amino acid sequence of the TIR region of TLR5 is as shown in SEQ ID NO:10, the amino acid sequence of T2A is as shown in SEQ ID NO:11, and the amino acid sequence of the CCR9 chemokine is as shown in SEQ ID NO:12.

[0011] In the second aspect, the present invention also provides a therapeutic vector for treating diseases with high expression of GUCY2C, comprising any one of the following amino acid sequences: SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, SEQ ID NO:11, SEQ ID NO:12.

[0012] In some embodiments of the present invention, the therapeutic vector is a lentiviral vector, and the therapeutic vector further contains a replication origin; specifically, the replication origin is 3'LTR and 5'LTR.

[0013] In a third aspect, the present invention also provides a method for preparing CAR-modified immune cells targeting GUCY2C. The therapeutic vector targeting GUCY2C of the present invention is packaged using a four-plasmid lentiviral packaging system to form a lentiviral therapeutic vector containing pMD2G, pMDLg / pRRE, and pRSV-Rev. Using 293T cells as lentiviral packaging cells, through cell culture, transfection, collection of virus solution, virus concentration, titer determination, and then transfection of T cells, CAR-modified immune cells targeting GUCY2C can be obtained.

[0014] In a fourth aspect, the present invention also provides CAR-modified immune cells targeting GUCY2C prepared by the method for preparing CAR-modified immune cells targeting GUCY2C.

[0015] In some embodiments of the present invention, the CAR-modified immune cells targeting GUCY2C include CAR-T, CAR-NK, CAR-NKT, CAR-γδT, and CAR-Macrophage.

[0016] In a fifth aspect, the present invention also provides the application of the CAR-modified immune cells targeting GUCY2C in the preparation of a kit or biological product for treating gastrointestinal tumors or other related diseases with high expression of GUCY2C.

[0017] In a fifth aspect, the present invention also provides a host cell for treating diseases with high expression of GUCY2C, and the host cell includes any one of the following amino sequences; amino sequences: SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:9, SEQ ID NO:10, SEQ ID NO:11, SEQ ID NO:12.

[0018] The beneficial effects of the present invention are as follows:

[0019] (1) The present invention effectively combines the adaptive immune response brought about by CD3ζ chain activation, the innate immune response triggered by the TLR5 signaling domain, and the gastrointestinal homing receptor CCR9, enabling the modified immune cells to have a gastrointestinal homing effect and improve tumor killing ability, and can be applied to the treatment of patients with gastrointestinal tumors with high expression of the GUCY2C molecule.

[0020] (2) The present invention provides a novel CAR-T targeting GUCY2C with gastrointestinal homing effect, which solves the problem of poor efficacy of existing CAR-T therapies in treating solid tumors, has broad application prospects and great market value, and provides methods for treating cancer patients with cancer cells expressing GUCY2C and methods for preventing diseases expressing GUCY2C. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] The drawings are used to provide a further understanding of the present invention, and constitute a part of the specification. Together with the embodiments of the present invention, they are used to explain the present invention and do not constitute a limitation to the present invention. In the drawings:

[0022] Figure 1 It is a schematic diagram of the DNA fragment of GUCY2C-CAR in the embodiment;

[0023] Figure 2 It is the plasmid map of PTK-GUCY2C-CAR in the embodiment;

[0024] Figure 3 It is a schematic diagram of the detection of GUCY2C protein expression in human colorectal adenocarcinoma cell line SW480 and human colon adenocarcinoma lung metastasis cell T84 by flow cytometry in the embodiment;

[0025] Figure 4 It is a schematic diagram of the in vitro killing results of T cells and CAR-T cells on SW480 and T84 in the embodiment;

[0026] Figure 5 It is a schematic diagram of the in vitro killing results of CAR-T cells on SW480 and SW480-GUCY2C + cell lines in the embodiment. DETAILED DESCRIPTION OF THE EMBODIMENTS

[0027] The following will explain the solution of the present invention in combination with the embodiments. Those skilled in the art will understand that the following embodiments are only used to illustrate the present invention and should not be regarded as limiting the scope of the present invention. For those not specified in the embodiments regarding specific techniques or conditions, they shall be carried out according to the techniques or conditions described in the literature in the art or according to the product specifications. For reagents or instruments not specified by the manufacturer, they are all conventional products that can be obtained through commercial purchase.

[0028] Example 1: Construction of Chimeric Antigen Receptor Targeting GUCY2C

[0029] From a large-capacity GUCY2C phage antibody library prepared using GUCY2C as an antigen, a scFv with strong specificity and high affinity against GUCY2C was screened. After multiple rounds of screening, 1 scFv that could specifically recognize GUCY2C on the surface of tumor cells was obtained. Sequencing and analysis of its sequence showed that its amino acid sequence was as shown in SEQ ID NO:5.

[0030] Synthesize fragments separately. Select CD19 signal peptide, the light chain of the anti-GUCY2C single-chain antibody, the heavy chain of the anti-GUCY2C single-chain antibody, connect the light chain and the heavy chain with a G4S linker, StrepII-G4S, CD8α hinge region, 4-1BB intracellular region, CD3ζ intracellular region, the TIR region of TLR5, T2A, and CCR9 chemokine; the amino acid sequence of the CD19 signal peptide is as shown in SEQ ID NO:1, the amino acid sequence of the light chain of the anti-GUCY2C single-chain antibody is as shown in SEQ ID NO:2, the amino acid sequence of the heavy chain of the anti-GUCY2C single-chain antibody is as shown in SEQ ID NO:3, the amino acid sequence of the G4S linker is as shown in SEQ ID NO:4, the amino acid sequence of StrepII-G4S is as shown in SEQ ID NO:6, the amino acid sequence of the CD8α hinge region is as shown in SEQ ID NO:7, the amino acid sequence of the 4-1BB intracellular region is as shown in SEQ ID NO:8, the amino acid sequence of the CD3ζ intracellular region is as shown in SEQ ID NO:9, the amino acid sequence of the TIR region of TLR5 is as shown in SEQ ID NO:10, the amino acid sequence of T2A is as shown in SEQ ID NO:11, and the amino acid sequence of the CCR9 chemokine is as shown in SEQ ID NO:12. Use the Overlap PCR technique to splice the GUCY2C-CAR molecule sequentially from the N-terminus to the C-terminus. The schematic diagram is as Figure 1 shown.

[0031] Double-digest the PTK plasmid vector with EcoRI and BamHI. Separate the digestion products by 1% agarose gel electrophoresis, recover the target band, obtain the concentrations of the vector and the target fragment, and ligate and transform the two according to a molar ratio of 1:2. Extract the plasmid to obtain the schematic diagram of the structure of PTK-GUCY2C-CAR as Figure 2 shown.

[0032] Example 2: Preparation and titer detection of the PTK-GUCY2C-CAR lentiviral vector

[0033] 1. Preparation of the lentiviral vector

[0034] Mix the target plasmid (PTK-GUCY2C-CAR) in a ratio of 12:10:6:5 (asFigure 2 , 11954 bp), pMDLg / pRRE, pRSV-Rev and pMD2G were transfected into 293T cells. After culturing for 72 h, the cell culture supernatant was collected. The supernatant was centrifuged at 3000 rpm for 30 min, and the lentivirus supernatant was filtered through a 0.22-μm filter membrane, and then centrifuged at 30000 g for 3 h at 4 °C. The supernatant was removed, and the precipitate was resuspended in 500 μL of T cell medium. The lentivirus concentrate was aliquoted, labeled as Lenti3-PTK-GUCY2C-CAR, and stored at -80 °C for later use.

[0035] 2. Detection of the active titer of the lentiviral vector

[0036] 1) Take 293T cells in good growth state, digest and count them, and seed 5.0×105 cells per well into a 6-well cell culture plate, and culture for 6 - 10 h until the cells adhere to the wall;

[0037] 2) Add lentivirus concentrate with gradient concentrations to the cell culture supernatant in the 6-well plate, gently shake the 6-well plate to mix the virus solution and the culture medium evenly, and culture in an incubator at 37 °C with 5% CO2 for 48 h;

[0038] 3) After culturing for 48 h, digest and collect the cells, detect the percentage of positive cells by flow cytometry, calculate the virus titer, with the unit of TU / mL, and the results are shown in Table 1.

[0039] Table 1. Analysis results of the detection of the active titer of lentivirus

[0040] Sample Name Titer (TU / mL) PTK-GUCY2C-CAR <![CDATA[1.5×10 8 >

[0041] Example 3: Preparation of GUCY2C CAR-T cells

[0042] 1. Construction of CAR-T cells by lentiviral transduction of T cells

[0043] T cells were sorted from PBMCs, and after 24 h, the well-growing T cells to be transfected with the corresponding cell number were taken out; the lentivirus concentrate was added to the six-well plate at an MOI of 5, and Polybrene was added to make its final concentration 5 μg / mL; incubated at 37 °C in an inclined position for 4 h, and finally the complete T cell medium was added to the normal culture concentration. Continue to culture and expand for 10 - 20 days, observe and count every day, and replenish the liquid and expand the culture according to the counted cell number, always maintaining the cell culture density at 1.0 - 2.0×10 6 cells / mL.

[0044] 2. Detection of the transduction efficiency of CAR-T cells

[0045] Six days after lentiviral transduction, take 1.0×10 6CAR-T cells were used, and the percentage of positive cells was detected by flow cytometry to reflect the percentage of CAR-T cells in the total cells. The results are shown in Table 2.

[0046] Table 2 Detection results of CAR-T cell transduction efficiency

[0047] Sample Name Transduction Efficiency PTK-GUCY2C-CAR 33.6％

[0048] Example 4: In vitro function detection of GUCY2C CAR-T cells

[0049] The cytotoxicity of GUCY2C-CAR-T cells was detected by calcein release method. The tumor target cells used are shown in Table 3. After taking out the effector cells according to different effector-to-target ratios (25:1, 5:1, 1:1), they were co-incubated with the target cells in a U-bottom 96-well culture plate for 3 h. At the same time, a positive control group (adding boric acid lysis solution) and a negative control group (adding PBS with 5% fetal bovine serum) were set up. After 3 h, the 96-well plate was centrifuged at 500 g for 10 min at room temperature. 100 μL of the supernatant was taken from each well and transferred to a flat-bottom 96-well plate, and the absorbance of each well was measured by fluorescence colorimetry in an enzyme-linked immunosorbent assay (ELISA) reader.

[0050] Table 3 Selection of target cell lines for anti-GUCY2C CAR-T cells

[0051] Cancer Type Cell Line Colorectal Cancer SW480, T84

[0052] The cytotoxicity of effector cells in each group was calculated according to the following formula:

[0053] Specific lysis percentage = (fluorescence value of experimental group - fluorescence value of spontaneous release group) / (fluorescence value of maximum release group - fluorescence value of spontaneous release group) × 100%.

[0054] The detection of GUCY2C protein expression in human colorectal adenocarcinoma cell line SW480 and human colon adenocarcinoma lung metastasis cell line T84 by flow cytometry was as Figure 3 shown; the in vitro killing and lysis results of T cells and GUCY2C CAR-T cells against human colorectal adenocarcinoma cell line SW480 and human colon adenocarcinoma lung metastasis cell line T84 were respectively as Figure 4 shown; the in vitro killing and lysis results against the SW480-GUCY2C+ cell line constructed by GUCY2C lentiviral transfection were as Figure 5 shown.

[0055] The results showed that GUCY2C CAR-T cells had significant killing effects on the positive cell lines T84 and SW480-GUCY2C+, but had no lysis ability on the negative cell line SW480, indicating that it specifically recognized GUCY2C and had in vitro killing function against human colorectal cancer cell lines.

[0056] From the above in vitro tumor killing results, it can be seen that the constructed CAR-T cells targeting GUCY2C can be used for the treatment of colorectal tumors.

[0057] Although the embodiments of the present invention have been shown and described above, it can be understood that the above embodiments are exemplary and should not be construed as limiting the present invention. Those of ordinary skill in the art can make changes, modifications, substitutions, and variations to the above embodiments within the scope of the present invention.

Claims

1. A chimeric antigen receptor targeting GUCY2C with gastrointestinal homing effect, characterized in that, It includes a signal peptide, the light chain of an anti-GUCY2C single-chain antibody, the heavy chain of the anti-GUCY2C single-chain antibody, a G4S linker, StrepII-G4S, the CD8α hinge region, the 4-1BB intracellular region, the CD3ζ intracellular region, the TLR5 TIR sequence, T2A, and the CCR9 chemokine.

2. The chimeric antigen receptor targeting GUCY2C with gastrointestinal homing effect as claimed in claim 1, wherein The amino acid sequence of the signal peptide is as shown in SEQ ID NO:1, the amino acid sequence of the light chain of the anti-GUCY2C single-chain antibody is as shown in SEQ ID NO:2, the amino acid sequence of the heavy chain of the anti-GUCY2C single-chain antibody is as shown in SEQ ID NO:3, the amino acid sequence of the G4S linker is as shown in SEQ ID NO:4, the amino acid sequence of StrepII-G4S is as shown in SEQ ID NO:6, the amino acid sequence of the CD8α hinge region is as shown in SEQ ID NO:7, the amino acid sequence of the 4-1BB intracellular region is as shown in SEQ ID NO:8, the amino acid sequence of the CD3ζ intracellular region is as shown in SEQ ID NO:9, the amino acid sequence of the TIR region of TLR5 is as shown in SEQ ID NO:10, the amino acid sequence of T2A is as shown in SEQ ID NO:11, and the amino acid sequence of the CCR9 chemokine is as shown in SEQ ID NO:

12.

3. A therapeutic vector for treating diseases with high expression of GUCY2C, characterized in that, It contains any one of the amino acid sequences described in claim 2.

4. The therapeutic vector for treating diseases with high GUCY2C expression according to claim 3, wherein The therapeutic vector is a lentiviral vector, and the therapeutic vector further contains a replication origin, and the replication origin is 3’LTR and 5’LTR.

5. A method for preparing CAR-modified immune cells targeting GUCY2C, characterized in that, The therapeutic vector targeting GUCY2C described in claim 3 or 4 is packaged using a four-plasmid lentiviral packaging system to form a lentiviral therapeutic vector containing pMD2G, pMDLg / pRRE, and pRSV-Rev. Using 293T cells as lentiviral packaging cells, through cell culture, transfection, collection of virus solution, virus concentration, titer determination, and then transfection of T cells, CAR-modified immune cells targeting GUCY2C can be obtained.

6. A CAR-modified immune cell targeting GUCY2C prepared by the preparation method described in claim 6.

7. The CAR-modified immune cells targeting GUCY2C according to claim 6, characterized in that, It includes CAR-T, CAR-NK, CAR-NKT, CAR-γδT, and CAR-Macrophage.

8. Use of a CAR-modified immune cell targeting GUCY2C described in claim 7 in the preparation of a kit or biological product for treating gastrointestinal tumors or other related diseases with high expression of GUCY2C.

9. A host cell for treating diseases with high expression of GUCY2C, characterized in that, It contains any one of the amino acid sequences described in claim 2.

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