Use of new immune checkpoint GRHL3 in diagnosis and treatment of cancer and evaluation of curative effect of immunotherapy

Abstract

The present invention belongs to the field of biomedicines, and particularly relates to use of a new immune checkpoint GRHL3 in the diagnosis and treatment of cancer and the evaluation of the curative effect of immunotherapy. On one hand, the present invention provides use of the new immune checkpoint GRHL3 in the evaluation of the killing ability of T cells, the diagnosis of cancer, the prediction of prognosis of a patient, and the prediction of the curative effect of immunotherapy; and on the other hand, the present invention provides use of an inhibitor of the new immune checkpoint GRHL3 in the improvement of the killing ability of T cells, the treatment of cancer, and the improvement of the curative effect of immunotherapy.

Description

Application of the new immune checkpoint GRHL3 in cancer diagnosis and treatment and evaluation of immunotherapy efficacy Technical Field

[0001] The present invention belongs to the field of biomedicine, and specifically relates to the application of the new immune checkpoint GRHL3 in cancer diagnosis and treatment and immunotherapy efficacy evaluation. Background Art

[0002] The recent rise of tumor immunotherapy differs from previous treatment strategies in that it targets the body's own immune system rather than tumor cells and tissues. This therapy aims to modulate and activate the immune system, relying on its own immune function to eliminate all tumor cells in the body, including those that have metastasized to distant sites. Currently, there are various approaches to tumor immunotherapy, but the use of immune checkpoint inhibitors, which block immunosuppressive pathways and thereby relieve the body's immune tolerance, reactivate the immune system, and enhance its ability to recognize and eliminate tumor cells, ultimately killing them, is a proven and effective treatment.

[0003] Immune checkpoints are proteins expressed on the surface of T cells or tumor cells that inhibit T cell proliferation and activation, allowing tumor cells to evade immune surveillance. Immune checkpoint inhibitors (ICIs) regulate T cell proliferation and activation through immune checkpoints, thereby inducing tumor cell death. ICIs have shown remarkable promise in the treatment of tumors such as malignant melanoma, non-small cell lung cancer (NSCLC), small cell lung cancer, and urothelial carcinoma. Currently, there are two main categories of ICIs used in clinical practice: one is anti-programmed cell death protein 1 (PD-1) / programmed cell death protein ligand 1 (PD-L1) antibodies, and the other is anti-cytotoxic T lymphocyte associated antigen-4 (CTLA-4) antibodies. In addition to CTLA-4 and PD-1, new target drugs such as TIM-3, LAG3, KIR, GITR, VISTA, IDO1, 4-1BB, and TDO2 are also being explored and developed, and many related drugs have entered the clinical trial stage. As research deepens, it is believed that more and more new immune checkpoints will be discovered.

[0004] Summary of the Invention

[0005] The main purpose of the present invention is to provide the application of a new immune checkpoint GRHL3 in cancer diagnosis and treatment and immunotherapy efficacy evaluation.

[0006] application

[0007] In one aspect, the present invention provides the use of a reagent for detecting GRHL3 expression in the preparation of a product for evaluating the traction force, killing ability, and infiltration ability (infiltration characteristics) of T cells.

[0008] Preferably, the T cells are CD8 + T cells (cytotoxic T lymphocytes / CTL cells).

[0009] Preferably, the T cells include CD8 T cells isolated from the blood (preferably peripheral blood) of the subject. + T cells.

[0010] Preferably, the subject is a cancer patient.

[0011] Preferably, the subject is a patient with gastric adenocarcinoma, hepatocellular carcinoma, colorectal cancer, lung cancer, breast cancer, glioblastoma multiforme, lung squamous cell carcinoma or cutaneous melanoma.

[0012] In one aspect, the present invention provides the use of a reagent for detecting GRHL3 expression in the preparation of a product for diagnosing cancer and predicting the prognosis of a patient, wherein the patient is a cancer patient receiving immunotherapy.

[0013] Preferably, the GRHL3 expression level is in the peripheral blood CD8 + Increased expression on T cells.

[0014] Preferably, the prognostic indicators include objective response rate (ORR), overall survival rate (OS), progression-free survival (PFS), time to progression (TTP), disease-free survival (DFS), time to treatment failure (TTF), response rate (RR), complete response (CR), and partial response (PR).

[0015] More preferably, the predicting of patient prognosis refers to predicting the patient's survival period.

[0016] Preferably, the cancer diagnosis is diagnosis of gastric adenocarcinoma, hepatocellular carcinoma or colorectal cancer.

[0017] Preferably, the predicted patient prognosis is the predicted prognosis of patients with colon cancer, lung cancer, breast cancer, rectal adenocarcinoma, glioblastoma multiforme, lung squamous cell carcinoma, skin melanoma and gastric adenocarcinoma.

[0018] In one aspect, the present invention provides the use of a reagent for detecting GRHL3 expression in the preparation of a product for predicting the efficacy of immunotherapy.

[0019] More specifically, the immunotherapy is for cancer patients.

[0020] Preferably, the cancer is gastric adenocarcinoma, hepatocellular carcinoma, colorectal cancer, lung cancer, breast cancer, glioblastoma multiforme, lung squamous cell carcinoma or skin melanoma.

[0021] Preferably, the immunotherapy is a treatment method that hinders the progression of cancer through T cells.

[0022] Preferably, the immunotherapy comprises treatment by administering immune checkpoint inhibitors. The immune checkpoints include PD1, CTLA4, VISTA, IDO, CD137, TIGIT, TIM3, BTLA, CD27L, CD40, LAG-3, CD270, GITR, SIRPα, etc.

[0023] Specifically, the immune checkpoint inhibitors include PD-1 / L1 immune checkpoint inhibitors, such as antibodies targeting PD-1 / L1, specifically pembrolizumab (Keytruda), nivolumab (Opdivo), atezolizumab (TECENTRIQ), durvalumab (IMFINZI), avelumab (BAVENCIO), cemiplimab (Libtayo), toyitriptyline, dabigatran, erikacitabine, and tislelizumab; and also include the CTLA4 immune checkpoint inhibitor Ipilimumab (trade name Yervoy).

[0024] More preferably, the immunotherapy is treatment with a PD-1 antibody.

[0025] In one aspect, the present invention provides the use of a GRHL3 inhibitor in the preparation of a drug for enhancing the traction force, killing ability, and tumor infiltration ability of T cells.

[0026] In one aspect, the present invention provides the use of GRHL3 inhibitors in the preparation of cancer treatment drugs and drugs for improving the efficacy of immunotherapy.

[0027] Preferably, the immunotherapy is a treatment method that hinders cancer progression through T cells.

[0028] Preferably, the immunotherapy comprises treatment by administering immune checkpoint inhibitors. The immune checkpoints include PD1, CTLA4, VISTA, IDO, CD137, TIGIT, TIM3, BTLA, CD27L, CD40, LAG-3, CD270, GITR, SIRPα, etc.

[0029] Specifically, the immune checkpoint inhibitors include PD-1 / L1 immune checkpoint inhibitors, such as antibodies targeting PD-1 / L1, specifically pembrolizumab (Keytruda), nivolumab (Opdivo), atezolizumab (TECENTRIQ), durvalumab (IMFINZI), avelumab (BAVENCIO), cemiplimab (Libtayo), toyitriptyline, dabigatran, erikacitabine, and tislelizumab; and also include the CTLA4 immune checkpoint inhibitor Ipilimumab (trade name Yervoy).

[0030] method

[0031] In one aspect, the present invention provides a method for evaluating the traction force, killing ability, and tumor infiltration ability of T cells, comprising the step of detecting the expression level of GRHL3 in T cells.

[0032] Preferably, the T cells include cells in the human body, cells isolated from the body, or commercial cell lines.

[0033] Preferably, the T cells are CD8 + T cells.

[0034] Preferably, the T cells include CD8 + T cells.

[0035] Preferably, the subject is a cancer patient.

[0036] Preferably, the subject is a patient with gastric adenocarcinoma, hepatocellular carcinoma, colorectal cancer, lung cancer, breast cancer, glioblastoma multiforme, lung squamous cell carcinoma or cutaneous melanoma.

[0037] In one aspect, the present invention provides a method for diagnosing cancer and / or predicting the prognosis of a patient, wherein the patient is a cancer patient receiving immunotherapy, the method comprising the step of detecting the expression level of GRHL3 in a sample from the subject.

[0038] Preferably, the sample is CD8 isolated from the blood (preferably peripheral blood) of the subject. + T cells.

[0039] Preferably, the GRHL3 expression level is in the peripheral blood CD8 + Increased expression on T cells.

[0040] Preferably, the subject is a person suspected of having cancer and awaiting diagnosis.

[0041] Preferably, the subject is a patient who has been confirmed to be a cancer patient and whose prognosis needs to be determined.

[0042] In one aspect, the present invention provides a method for predicting the efficacy of immunotherapy, comprising the step of detecting the expression level of GRHL3 in a sample from a subject, wherein the subject is a cancer patient who has received immunotherapy.

[0043] Preferably, the sample is CD8 isolated from the blood (preferably peripheral blood) of a subject. + T cells.

[0044] In a preferred embodiment, the evaluation method and prediction method of the present invention include the following additional step: comparing the measured nucleic acid or protein expression level with a control level. The state, nature, amount, and condition of the control level can be adjusted as necessary. Preferably, a healthy or benign control level can be used. As used herein, the term "comparison" refers to any suitable method for evaluating, calculating, assessing, or processing data.

[0045] In one aspect, the present invention provides a method for enhancing the traction force, killing ability, and tumor infiltration ability of T cells, comprising contacting T cells with a GRHL3 inhibitor.

[0046] Preferably, the T cells include CD8 + T cells;

[0047] Preferably, the T cells include CD8 + T cells;

[0048] Preferably, the T cells comprise isolated in vitro cells.

[0049] Preferably, the subject is a cancer patient.

[0050] In one aspect, the present invention provides a method for treating cancer or enhancing the efficacy of immunotherapy, comprising administering a GRHL3 inhibitor to a patient.

[0051] Specifically, the inhibitor of GRHL3 can be administered to the patient by any method that can make the GRHL3 inhibitor reach the target cell. These methods include but are not limited to oral, rectal, nasal, external application, intradermal, subcutaneous, intravenous, intramuscular, intratracheal and intraperitoneal administration. The inhibitor can be dissolved in aqueous vehicles such as sodium chloride, Ringer's solution, glucose, glucose and sodium chloride, water, saline, Ringer's solution, glucose solution for administration, and can also be dissolved in liposomes and non-aqueous vehicles such as fixed oil, cottonseed oil, sesame oil or peanut oil and esters for administration. In addition, various additives that can enhance the stability, sterility and isotonicity of the composition can also be added, including antimicrobial preservatives, antioxidants, chelating agents and buffers. However, any vehicle, diluent or additive used must all have biocompatibility and be compatible with the inhibitor of the present invention.

[0052] The GRHL3 inhibitors of the present invention can be formulated into various dosage forms as needed, and a physician can determine the dosage that is beneficial to the patient based on factors such as patient type, age, weight, general disease condition, and route of administration. The dosage of the GRHL3 inhibitor can vary depending on the intended recipient, route of administration, or pharmaceutical formulation, but is premised on ensuring that the pharmaceutical composition achieves an effective blood concentration in a mammal. The pharmaceutical composition can be in any dosage form and can be administered in any manner.

[0053] product

[0054] In one aspect, the present invention provides a drug combination composition for treating cancer, wherein the drug combination composition comprises a GRHL3 inhibitor and an immune checkpoint inhibitor (ICIs).

[0055] Preferably, the immune checkpoints include PD1, CTLA4, VISTA, IDO, CD137, TIGIT, TIM3, BTLA, CD27L, CD40, LAG-3, CD270, GITR, and SIRPα.

[0056] Specifically, the immune checkpoint inhibitors include PD-1 / L1 immune checkpoint inhibitors and CTLA4 immune checkpoint inhibitors.

[0057] More specifically, the immune checkpoint inhibitor is a PD-1 / L1 immune checkpoint inhibitor.

[0058] In a specific embodiment, the dosage forms of the GRHL3 inhibitor and the immune checkpoint inhibitor in the drug combination composition are the same or different.

[0059] In a specific embodiment, the GRHL3 inhibitor and the immune checkpoint inhibitor in the drug combination composition are administered simultaneously or sequentially, specifically, at intervals of 0, 1, 2, 3, 4, 5, 6, 7 or more days.

[0060] In a specific embodiment, the dosage ratio of the GRHL3 inhibitor and the immune checkpoint inhibitor in the drug combination composition is 1:0.1-2; specifically including 3:1, 2:1, 1:1, 1:2, and 1:3.

[0061] General Concepts

[0062] As used herein, "cancer" and "tumor" are interchangeable terms that refer to any abnormal cell or tissue growth or proliferation in an animal. As used herein, the terms "cancer" and "tumor" encompass solid tumor cancers and non-solid tumor cancers, and also encompass malignant, pre-malignant, and benign growths, such as dysplasia.

[0063] Preferably, the solid tumor cancer includes melanoma, prostate cancer, bladder cancer, kidney cancer, liver cancer, breast cancer, ovarian cancer, lung cancer, pancreatic cancer, brain tumor, cervical cancer, uterine cancer, endometrial cancer, colon cancer, rectal cancer, hemangioendothelioma, testicular cancer, skin cancer, human immunodeficiency virus-associated solid tumors, head and neck cancer, glioma, skin cancer;

[0064] Preferably, the solid tumor cancer includes cell carcinoma and adenocarcinoma.

[0065] Preferably, the non-solid tumor cancer includes blood cancer, for example, leukemia: including acute lymphocytic leukemia (ALL), more specifically including B cell ALL, T cell ALL; acute myeloid leukemia (AML), more specifically including B cell AML, T-cell AML; chronic myeloid leukemia (CML), more specifically including B cell CML, T cell CML; and chronic lymphocytic leukemia (CLL), more specifically including B cell CLL, T cell CLL.

[0066] Preferably, the non-solid tumor cancer includes lymphoma: including Hodgkin lymphoma (HL), more specifically including B cell HL, T cell HL; non-Hodgkin lymphoma (NHL), more specifically including B cell NHL.

[0067] As used herein, the term "inhibitor" refers to a substance that targets, reduces or inhibits at least one activity of a target protein. Specifically, the target protein of interest in the present invention is GRHL3.

[0068] Specifically, the inhibitors of the present invention include polypeptides, polynucleotides or small molecules that specifically target GRHL3; for example, antibodies, antisense oligonucleotides, low molecular weight molecules (LMW), siRNA, aptamers, small molecule compounds, etc. that bind to GRHL3, and also include compounds that can reduce the expression of target proteins.

[0069] Preferably, the inhibitor is artificially synthesized or naturally occurring.

[0070] More specifically, as shown in the specific embodiments of the present invention, a specific GRHL3-targeting shRNA is used. More specifically, its target sequence is as shown in the specific embodiments of the present invention. Selecting a specific target sequence and designing a synthetic shRNA are conventional techniques well known in the art.

[0071] The detection of the present invention includes evaluating the presence of marker expression in a sample from a subject and measuring the amount (which may be an effective amount) of the marker to derive a qualitative result or a quantitative concentration level. The markers of interest in the present invention include GRHL3.

[0072] Preferably, the reagent for detecting the GRHL3 expression level includes a reagent for detecting the GRHL3 protein expression level and / or the GRHL3 mRNA expression level.

[0073] Preferably, the reagents for detecting the expression level of GRHL3 protein include reagents used in the following methods: hematoxylin-eosin staining (HE staining), safranin O-fast green staining, protein blotting (Western Blot method), enzyme-linked immunosorbent assay (ELISA), radioimmunoassay (RIA), sandwich assay, immunohistochemistry (Immunohistochemistry) staining method, mass spectrometry, immunoprecipitation analysis, complement fixation analysis, flow cytometry fluorescence resolution technology and protein chip method.

[0074] Preferably, the reagents for detecting GRHL3 mRNA expression include reagents used in the following methods: PCR-based detection method, Southern hybridization method, Northern hybridization method, dot hybridization method, fluorescence in situ hybridization method, DNA microarray method, ASO method, and high-throughput sequencing platform method.

[0075] As used herein, the term "treat" refers to the complete or partial alleviation or reduction of a disease, condition, or disorder or the symptoms, adverse effects, consequences, or phenotypes associated therewith. The effects of treatment include, but are not limited to, preventing the occurrence or recurrence of the disease, alleviating symptoms, reducing any direct or indirect pathological consequences of the disease, preventing metastasis, slowing the rate of disease progression, improving or alleviating the disease state, and alleviating or improving prognosis. The term does not imply a complete cure of the disease or the complete elimination of any symptom or effect or all symptoms or consequences.

[0076] The "drug" of the present invention can be a composition, which, in addition to containing a GRHL3 inhibitor, can also contain other cancer therapeutic drugs and / or pharmaceutically acceptable carriers or diluents. Specifically, the carrier is solid, gel or liquid, the solid carrier is lactose, white clay, sucrose, talc, gelatin, agar, pectin, gum arabic, magnesium stearate, stearic acid or a degradable polymer, the liquid carrier is phosphate buffered saline solution, syrup, oil, water, emulsion, wetting agent or sterile solution, and the diluent is distilled water, physiological saline, Ringer's solution, glucose solution, PBS solution or Hank's solution. The dosage form of the drug can be tablets, pills, powders, granules, capsules, lozenges, syrups, emulsions, suspensions, controlled release preparations, aerosols, films, injections, intravenous drips, transdermal absorption preparations, ointments, lotions, adhesive preparations, suppositories, nasal preparations, pulmonary preparations or eye drops. BRIEF DESCRIPTION OF THE DRAWINGS

[0077] FIG1 is a graph showing the results of a traction force assay on GRHL3 knockdown T cells.

[0078] FIG2 is a flow cytometry result of co-culture of GRHL3 knockdown T cells and B16F10 cells.

[0079] Figure 3 shows the peripheral blood CD8 + Figure 2 shows the detection results of GRHL3 expression in T cells.

[0080] Figure 4 is a peripheral blood CD8 + Figure 2 shows the detection results of T cell traction force.

[0081] FIG5 is a graph showing the correlation between GRHL3 expression and survival in cancer patients receiving PD-1 antibody treatment.

[0082] Figure 6 shows the expression of GRHL3 and the expression of peripheral blood CD8 in patients with colon cancer, lung cancer and breast cancer. + Results of the T cell tumor infiltration correlation diagram.

[0083] FIG7 is a graph showing the relationship between GRHL3 expression and overall survival of patients after immunotherapy.

[0084] Figure 8 shows the relationship between GRHL3 expression and CD8 + Figure 3. Correlation analysis of T cell infiltration characteristics in patients with rectal adenocarcinoma, glioblastoma multiforme, and lung squamous cell carcinoma.

[0085] Figure 9 shows the relationship between GRHL3 expression and CD8 + Figure 1. Correlation of T cell infiltration characteristics between patients with cutaneous melanoma and gastric adenocarcinoma. DETAILED DESCRIPTION

[0086] The present invention will be further described below with reference to specific embodiments, but the scope of protection of the present invention is not limited thereto. Any technician familiar with the technical field, within the technical scope disclosed by the present invention, who makes equivalent replacements or changes based on the technical solutions and inventive concepts of the present invention, should be covered by the scope of protection of the present invention.

[0087] Unless otherwise specified, the materials and reagents used in the following examples can be obtained from commercial sources.

[0088] Example 1: Knockdown of GRHL3 (Grhl3) leads to enhanced T cell traction and cytotoxicity

[0089] Sense and antisense sequences were synthesized for the shRNA targets shown in SEQ ID NO.1 and SEQ ID NO.4, and Grhl3-shRNAs lentiviral vectors were constructed. T cells were transduced to obtain Grhl3 knockdown mouse CD8+ T cells.

[0090] Table 1. Sequences related to knockdown of GRHL3

[0091] 1) T cell traction force detection and results

[0092] The glass bottom of the confocal dish was pretreated with APTES (Sigma-Aldrich). After polymerization, the gel surface was covered with a fluorescent bead solution for 15 minutes to fix it. The gel was activated with sulfamethoxazole (Sanpah) and coated with polylysine (PLL) overnight at 4°C. Then, 5 μg / mL anti-CD3ε antibody was applied at 37°C for 60 minutes. Cells were incubated on the PAA gel for 30 minutes to ensure activation. Phase contrast and fluorescent bead fluorescence images of the cells were taken to measure the displacement field generated during T cell activation.

[0093] The results showed that CD8 + T cells significantly enhanced traction force (Figure 1).

[0094] 2) Cytotoxicity assay and results

[0095] GRHL3 knockdown mice CD8 + T cells were co-cultured with B16F10 cells (mouse melanoma cell line, purchased from ATCC) at a ratio of 20:1 for 4 hours, and CD45 - Tumor cell apoptosis.

[0096] The results showed that knocking down GRHL3 levels enhanced T cell cytotoxicity and promoted tumor cell apoptosis (Figure 2).

[0097] Example 2: Peripheral blood CD8 of cancer patients + GRHL3 expression and traction force in T cells

[0098] Peripheral blood was collected from 6 healthy volunteers, 10 patients with colorectal cancer (CRC), 6 patients with gastric adenocarcinoma (STAD), and 6 patients with hepatocellular carcinoma (HCC). CD8+ T cells were isolated from the peripheral blood of healthy blood donors and cancer patients using the RosetteSep human CD8+ T cell isolation kit, and the expression of GRHL3 was detected by immunofluorescence assay.

[0099] The results showed that peripheral blood CD8 + GRHL3 expression was elevated in T cells ( Figure 3 ).

[0100] The traction force was tested according to the traction force detection method shown in Example 1. The results showed that compared with the control group, the peripheral blood CD8 + T cell traction force was significantly reduced (Figure 4).

[0101] Example 3: GRHL3 expression is negatively correlated with cancer patient survival and T cell tumor infiltration

[0102] To perform survival analysis, we collected GRHL3 gene-related data and patient survival information from the Kaplan-Meier Plotter dataset. + To correlate T cell infiltration signatures, we used RNA-seq data from the Timer2.0 dataset.

[0103] Kaplan-Meier Plotter data showed that GRHL3 expression was negatively correlated with survival in patients with various cancers who received PD-1 antibody treatment (n=896). GRHL3 expression was also negatively correlated with peripheral blood CD8 + There was a negative correlation between the infiltration of T cells into the tumor and the infiltration of T cells into the tumor (Figures 5-6).

[0104] High expression of GRHL3 shortens the overall survival of cancer patients responsive to all immunotherapies (Figure 7); GRHL3 expression is closely related to CD8 + T cell infiltration characteristics were negatively correlated in patients with rectal adenocarcinoma, glioblastoma multiforme, lung squamous cell carcinoma, skin melanoma, and gastric adenocarcinoma (Figures 8-9).

Claims

1. Application of reagents for detecting GRHL3 expression in the preparation of products for evaluating the traction, killing, and infiltration capabilities of T cells; Preferably, the T cells are CD8 + T cells; Preferably, the T cells include CD8 + T cells; Preferably, the T cells include CD8 + T cells; Preferably, the subject is a cancer patient; Preferably, the cancer includes solid tumor cancer and non-solid tumor cancer; Preferably, the solid tumor cancer includes melanoma, prostate cancer, bladder cancer, kidney cancer, liver cancer, breast cancer, ovarian cancer, lung cancer, pancreatic cancer, brain tumor, cervical cancer, uterine cancer, endometrial cancer, colon cancer, rectal cancer, hemangioendothelioma, testicular cancer, skin cancer, human immunodeficiency virus-associated solid tumors, head and neck cancer, glioma, skin cancer; Preferably, the non-solid tumor cancer includes blood cancer and lymphoma; Preferably, the subject is a patient with gastric adenocarcinoma, hepatocellular carcinoma, colorectal cancer, lung cancer, breast cancer, glioblastoma multiforme, lung squamous cell carcinoma or cutaneous melanoma.

2. A method for diagnosing cancer and / or predicting the prognosis of a patient, wherein the patient is a cancer patient receiving immunotherapy, the method comprising the step of detecting the expression level of GRHL3 in a sample from the subject; Preferably, the T cells comprise CD8 T cells isolated from the blood of a subject. + T cells; Preferably, the T cells include CD8 T cells isolated from the peripheral blood of the subject. + T cells; Preferably, the GRHL3 expression level is in the blood CD8 + Increased expression on T cells; Preferably, the predicting of patient prognosis refers to predicting the patient's survival period; Preferably, the cancer includes solid tumor cancer and non-solid tumor cancer; Preferably, the solid tumor cancer includes melanoma, prostate cancer, bladder cancer, kidney cancer, liver cancer, breast cancer, ovarian cancer, lung cancer, pancreatic cancer, brain tumor, cervical cancer, uterine cancer, endometrial cancer, colon cancer, cancer, colorectal cancer, hemangioendothelioma, testicular cancer, skin cancer, human immunodeficiency virus-associated solid tumors, head and neck cancer, glioma, skin cancer; Preferably, the non-solid tumor cancer includes blood cancer and lymphoma; Preferably, the cancer diagnosis is gastric adenocarcinoma, hepatocellular carcinoma or colorectal cancer; Preferably, the predicted patient prognosis is the predicted prognosis of patients with colon cancer, lung cancer, breast cancer, rectal adenocarcinoma, glioblastoma multiforme, lung squamous cell carcinoma, skin melanoma and gastric adenocarcinoma.

3. A method for predicting the efficacy of immunotherapy, comprising the step of detecting the expression level of GRHL3 in a sample from a subject, wherein the subject is a cancer patient receiving immunotherapy; Preferably, the T cells comprise CD8 T cells isolated from the blood of a subject. + T cells; Preferably, the T cells include CD8 T cells isolated from the peripheral blood of the subject. + T cells; Preferably, the cancer includes solid tumor cancer and non-solid tumor cancer; Preferably, the solid tumor cancer includes melanoma, prostate cancer, bladder cancer, kidney cancer, liver cancer, breast cancer, ovarian cancer, lung cancer, pancreatic cancer, brain tumor, cervical cancer, uterine cancer, endometrial cancer, colon cancer, rectal cancer, hemangioendothelioma, testicular cancer, skin cancer, human immunodeficiency virus-associated solid tumors, head and neck cancer, glioma, skin cancer; Preferably, the non-solid tumor cancer includes blood cancer and lymphoma; Preferably, the cancer is gastric adenocarcinoma, hepatocellular carcinoma, colorectal cancer, lung cancer, breast cancer, glioblastoma multiforme, lung squamous cell carcinoma or skin melanoma; Preferably, the immunotherapy is a treatment method that hinders cancer progression through T cells; Preferably, the immunotherapy comprises treatment by administering immune checkpoint inhibitors; the immune checkpoints include PD1, CTLA4, VISTA, IDO, CD137, TIGIT, TIM3, BTLA, CD27L, CD40, LAG-3, CD270, GITR, SIRPα; Specifically, the immune checkpoint inhibitors include PD-1 / L1 immune checkpoint inhibitors and CTLA4 immune checkpoint inhibitors; More preferably, the immunotherapy is treatment with a PD-1 antibody.

4. The method according to any one of claims 1 to 3, wherein the GRHL3 expression level comprises the GRHL3 protein expression level and / or the GRHL3 mRNA expression level; Preferably, the method for detecting the expression level of GRHL3 protein comprises: Hematoxylin-eosin staining, Safranin O-Fast Green staining, Western blotting, enzyme-linked immunosorbent assay, radioimmunoassay, sandwich assay, immunohistochemical staining, mass spectrometry, immunoprecipitation analysis, complement fixation assay, flow cytometry fluorescence resolution technology, and protein microarray; Preferably, the method for detecting the GRHL3 mRNA expression level includes: a PCR-based detection method, a Southern hybridization method, a Northern hybridization method, a dot hybridization method, a fluorescence in situ hybridization method, a DNA microarray method, an ASO method, and a high-throughput sequencing platform method.

5. A method for improving the traction, killing, and tumor infiltration capabilities of T cells, the method comprising contacting T cells with a GRHL3 inhibitor; Preferably, the T cells are CD8 + T cells; Preferably, the T cells include CD8 + T cells; Preferably, the T cells include CD8 + T cells; Preferably, the subject is a cancer patient; Preferably, the cancer includes solid tumor cancer and non-solid tumor cancer; Preferably, the solid tumor cancer includes melanoma, prostate cancer, bladder cancer, kidney cancer, liver cancer, breast cancer, ovarian cancer, lung cancer, pancreatic cancer, brain tumor, cervical cancer, uterine cancer, endometrial cancer, colon cancer, rectal cancer, hemangioendothelioma, testicular cancer, skin cancer, human immunodeficiency virus-associated solid tumors, head and neck cancer, glioma, skin cancer; Preferably, the non-solid tumor cancer includes blood cancer and lymphoma; Preferably, the subject is a patient with gastric adenocarcinoma, hepatocellular carcinoma, colorectal cancer, lung cancer, breast cancer, glioblastoma multiforme, lung squamous cell carcinoma or cutaneous melanoma.

6. A method for treating cancer or improving the efficacy of immunotherapy, comprising administering a GRHL3 inhibitor to a patient; Preferably, the patient is a cancer patient; Preferably, the cancer includes solid tumor cancer and non-solid tumor cancer; Preferably, the solid tumor cancer includes melanoma, prostate cancer, bladder cancer, kidney cancer, liver cancer, breast cancer, ovarian cancer, lung cancer, pancreatic cancer, brain tumor, cervical cancer, uterine cancer, endometrial cancer, colon cancer, rectal cancer, hemangioendothelioma, testicular cancer, skin cancer, human immunodeficiency virus-associated solid tumors, head and neck cancer, glioma, skin cancer; Preferably, the non-solid tumor cancer includes blood cancer and lymphoma.

7. The method according to claim 6, wherein the immunotherapy is a treatment method that inhibits cancer progression through T cells; Preferably, the immunotherapy comprises treatment by administering immune checkpoint inhibitors, wherein the immune checkpoints include PD1, CTLA4, VISTA, IDO, CD137, TIGIT, TIM3, BTLA, CD27L, CD40, LAG-3, CD270, GITR, and SIRPα; Specifically, the immune checkpoint inhibitors include PD-1 / L1 immune checkpoint inhibitors and CTLA4 immune checkpoint inhibitors; More preferably, the immunotherapy is treatment with a PD-1 antibody.

8. The method of claim 5 or 6, wherein the inhibitor comprises a polypeptide, polynucleotide or small molecule that specifically targets GRHL3; Preferably, the inhibitor comprises an antibody, an antisense oligonucleotide, a low molecular weight molecule, siRNA, shRNA, an aptamer, or a compound that binds to GRHL3.

9. A drug combination composition for treating cancer, comprising a GRHL3 inhibitor and an immune checkpoint inhibitor; Preferably, the immune checkpoints include PD1, CTLA4, VISTA, IDO, CD137, TIGIT, TIM3, BTLA, CD27L, CD40, LAG-3, CD270, GITR, and SIRPα; Specifically, the immune checkpoint inhibitors include PD-1 / L1 immune checkpoint inhibitors and CTLA4 immune checkpoint inhibitors; More specifically, the immune checkpoint inhibitor is a PD-1 / L1 immune checkpoint inhibitor; Preferably, the inhibitor comprises a polypeptide, polynucleotide or small molecule that specifically targets GRHL3; Preferably, the inhibitor comprises an antibody, an antisense oligonucleotide, a low molecular weight molecule, siRNA, shRNA, an aptamer, or a compound that binds to GRHL3.

10. Application of reagents for detecting GRHL3 expression in the preparation of products for evaluating the traction, killing, and infiltration capabilities of T cells; Preferably, the T cells are CD8 + T cells; Preferably, the T cells comprise CD8 T cells isolated from the blood of a subject. + T cells; Preferably, the T cells include CD8 T cells isolated from the peripheral blood of the subject. + T cells; Preferably, the subject is a cancer patient; Preferably, the cancer includes solid tumor cancer and non-solid tumor cancer; Preferably, the solid tumor cancer includes melanoma, prostate cancer, bladder cancer, kidney cancer, liver cancer, breast cancer, ovarian cancer, lung cancer, pancreatic cancer, brain tumor, cervical cancer, uterine cancer, endometrial cancer, colon cancer, rectal cancer, hemangioendothelioma, testicular cancer, skin cancer, human immunodeficiency virus-associated solid tumors, head and neck cancer, glioma, skin cancer; Preferably, the non-solid tumor cancer includes blood cancer and lymphoma; Preferably, the subject is a patient with gastric adenocarcinoma, hepatocellular carcinoma, colorectal cancer, lung cancer, breast cancer, glioblastoma multiforme, lung squamous cell carcinoma or cutaneous melanoma.

11. Use of a reagent for detecting GRHL3 expression in the preparation of a product for diagnosing cancer and predicting the prognosis of a patient undergoing immunotherapy; Preferably, the GRHL3 expression level is in the peripheral blood CD8 + Increased expression on T cells; More preferably, the predicting of the patient's prognosis refers to predicting the patient's survival period; Preferably, the cancer includes solid tumor cancer and non-solid tumor cancer; Preferably, the solid tumor cancer includes melanoma, prostate cancer, bladder cancer, kidney cancer, liver cancer, breast cancer, ovarian cancer, lung cancer, pancreatic cancer, brain tumor, cervical cancer, uterine cancer, endometrial cancer, colon cancer, rectal cancer, hemangioendothelioma, testicular cancer, skin cancer, human immunodeficiency virus-associated solid tumors, head and neck cancer, glioma, skin cancer; Preferably, the non-solid tumor cancer includes blood cancer and lymphoma; Preferably, the cancer diagnosis is gastric adenocarcinoma, hepatocellular carcinoma or colorectal cancer; Preferably, the predicted patient prognosis is the predicted prognosis of patients with colon cancer, lung cancer, breast cancer, rectal adenocarcinoma, glioblastoma multiforme, lung squamous cell carcinoma, skin melanoma and gastric adenocarcinoma.

12. Application of reagents for detecting GRHL3 expression in the preparation of products for predicting the efficacy of immunotherapy; More specifically, the immunotherapy is for cancer patients; Preferably, the cancer includes solid tumor cancer and non-solid tumor cancer; Preferably, the solid tumor cancer includes melanoma, prostate cancer, bladder cancer, kidney cancer, liver cancer, breast cancer, ovarian cancer, lung cancer, pancreatic cancer, brain tumor, cervical cancer, uterine cancer, endometrial cancer, colon cancer, rectal cancer, hemangioendothelioma, testicular cancer, skin cancer, human immunodeficiency virus-associated solid tumors, head and neck cancer, glioma, skin cancer; Preferably, the non-solid tumor cancer includes blood cancer and lymphoma; Preferably, the cancer is gastric adenocarcinoma, hepatocellular carcinoma, colorectal cancer, lung cancer, breast cancer, glioblastoma multiforme, lung squamous cell carcinoma or skin melanoma; Preferably, the immunotherapy is a treatment method that hinders cancer progression through T cells; Preferably, the immunotherapy comprises treatment by administering immune checkpoint inhibitors; the immune checkpoints include PD1, CTLA4, VISTA, IDO, CD137, TIGIT, TIM3, BTLA, CD27L, CD40, LAG-3, CD270, GITR, SIRPα; Specifically, the immune checkpoint inhibitors include PD-1 / L1 immune checkpoint inhibitors and CTLA4 immune checkpoint inhibitors; More preferably, the immunotherapy is treatment with a PD-1 antibody.

13. The use according to any one of claims 10 to 12, wherein the reagent for detecting GRHL3 expression comprises a reagent for detecting GRHL3 protein expression and / or GRHL3 mRNA expression; Preferably, the reagents for detecting the expression level of GRHL3 protein include reagents used in the following methods: hematoxylin-eosin staining, safranin O-fast green staining, Western blotting, enzyme-linked immunosorbent assay, radioimmunoassay, sandwich assay, immunohistochemical staining method, mass spectrometry, immunoprecipitation analysis, complement fixation analysis, flow cytometry fluorescence resolution technology and protein chip method; Preferably, the reagents for detecting GRHL3 mRNA expression include reagents used in the following methods: PCR-based detection method, Southern hybridization method, Northern hybridization method, dot hybridization method, fluorescence in situ hybridization method, DNA microarray method, ASO method, and high-throughput sequencing platform method.

14. Application of GRHL3 inhibitors in the preparation of drugs that enhance the traction, killing, and tumor infiltration capabilities of T cells; Preferably, the T cells are CD8 + T cells; Preferably, the T cells include CD8 + T cells; Preferably, the T cells include CD8 + T cells; Preferably, the subject is a cancer patient; Preferably, the cancer includes solid tumor cancer and non-solid tumor cancer; Preferably, the solid tumor cancer includes melanoma, prostate cancer, bladder cancer, kidney cancer, liver cancer, breast cancer, ovarian cancer, lung cancer, pancreatic cancer, brain tumor, cervical cancer, uterine cancer, endometrial cancer, colon cancer, rectal cancer, hemangioendothelioma, testicular cancer, skin cancer, human immunodeficiency virus-associated solid tumors, head and neck cancer, glioma, skin cancer; Preferably, the non-solid tumor cancer includes blood cancer and lymphoma; Preferably, the subject is a patient with gastric adenocarcinoma, hepatocellular carcinoma, colorectal cancer, lung cancer, breast cancer, glioblastoma multiforme, lung squamous cell carcinoma or cutaneous melanoma.

15. Application of GRHL3 inhibitors in the preparation of cancer treatment drugs and drugs to enhance the efficacy of immunotherapy; Preferably, the cancer includes solid tumor cancer and non-solid tumor cancer; Preferably, the solid tumor cancer includes melanoma, prostate cancer, bladder cancer, kidney cancer, liver cancer, breast cancer, ovarian cancer, lung cancer, pancreatic cancer, brain tumor, cervical cancer, uterine cancer, endometrial cancer, colon cancer, rectal cancer, hemangioendothelioma, testicular cancer, skin cancer, human immunodeficiency virus-associated solid tumors, head and neck cancer, glioma, skin cancer; Preferably, the non-solid tumor cancer includes blood cancer and lymphoma.

16. The use according to claim 15, wherein the immunotherapy is a treatment method that inhibits cancer progression through T cells; Preferably, the immunotherapy comprises treatment by administering immune checkpoint inhibitors, wherein the immune checkpoints include PD1, CTLA4, VISTA, IDO, CD137, TIGIT, TIM3, BTLA, CD27L, CD40, LAG-3, CD270, GITR, and SIRPα; Specifically, the immune checkpoint inhibitors include PD-1 / L1 immune checkpoint inhibitors and CTLA4 immune checkpoint inhibitors; More preferably, the immunotherapy is treatment with a PD-1 antibody.

17. The use according to claim 14 or 15, wherein the inhibitor comprises a polypeptide, polynucleotide or small molecule that specifically targets GRHL3; Preferably, the inhibitor comprises an antibody, an antisense oligonucleotide, a low molecular weight molecule, siRNA, shRNA, an aptamer, or a compound that binds to GRHL3.

18. A drug combination composition for treating cancer, comprising a GRHL3 inhibitor and an immune checkpoint inhibitor; Preferably, the immune checkpoints include PD1, CTLA4, VISTA, IDO, CD137, TIGIT, TIM3, BTLA, CD27L, CD40, LAG-3, CD270, GITR, and SIRPα; Specifically, the immune checkpoint inhibitors include PD-1 / L1 immune checkpoint inhibitors and CTLA4 immune checkpoint inhibitors; More specifically, the immune checkpoint inhibitor is a PD-1 / L1 immune checkpoint inhibitor; Preferably, the inhibitor comprises a polypeptide, polynucleotide or small molecule that specifically targets GRHL3; Preferably, the inhibitor comprises an antibody, an antisense oligonucleotide, a low molecular weight molecule, siRNA, shRNA, an aptamer, or a compound that binds to GRHL3.

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