Application of PSMB5 in diagnosis and treatment of bladder cancer

Abstract

The invention discloses application of PSMB5 in diagnosis and treatment of bladder cancer, and belongs to the technical field of biological medicine. The research finds that the expression of the PSMB5 gene in bladder cancer tissues is obviously higher than that in para-carcinoma tissues, and the high-expression PSMB5 is related to poor prognosis of bladder cancer patients. The invention verifies and finds that proliferation, invasion, migration and EMT transformation of the PSMB5 on bladder cancer are possibly caused by mediation of a Wnt / beta-catenin signal pathway. The invention aims to clearly inhibit the action mechanism of PSMB5 expression in a Wnt / beta-catenin signal channel for regulating and controlling the bladder cancer, and provides a new therapeutic target for treatment of the bladder cancer.

Description

Technical Field

[0001] The present invention belongs to the field of biomedicine technology, and specifically relates to an application of PSMB5 in diagnosing and treating bladder cancer. Background Art

[0002] The pathogenesis of bladder cancer (BLCA) is still unclear. Studies have found that smoking and occupational exposure to carcinogens such as aromatic amines are the main risk factors for bladder cancer. The most common histological type of BLCA is urothelial carcinoma, accounting for more than 90%, while squamous cell carcinoma, adenocarcinoma, small cell carcinoma and sarcoma are relatively rare. According to pathological histological classification, BLCA can be divided into non-muscle invasive bladder cancer (NMIBC) and muscle-invasive bladder cancer (MIBC). Although radical surgery and radiotherapy are effective, 25% of MIBC patients still have a poor prognosis. In addition, approximately 75%-80% of bladder cancer patients are initially diagnosed with NMIBC, of ​​which approximately 50% will relapse within five years and have a poor prognosis. The treatment of MIBC patients is usually radical cystectomy combined with regional pelvic lymph node dissection, followed by cisplatin-based chemotherapy. However, cisplatin-based chemotherapy can cause serious toxic side effects and has a relatively low anticancer effect, with only 40%-60% of patients with metastatic bladder cancer showing clinical responses. Despite significant progress in combined therapies including surgery, chemotherapy, and radiotherapy, the high recurrence and metastasis rates of BLCA in the past few decades have resulted in disease-free survival and 5-year survival rates of BLCA patients below 10%. Therefore, it is necessary to conduct in-depth studies on the mechanism of action of BLCA in order to find molecular biomarkers and therapeutic targets related to BLCA progression and provide new ideas for the diagnosis and treatment of BLCA.

[0003] The ubiquitin-proteasome is responsible for the selective removal of misfolded, damaged and unassembled polypeptide chains and regulatory proteins in cells. It plays an important role in cell cycle progression, signal transduction, proliferation, apoptosis, regulation of surface receptors, and regulation of tumor suppressor proteins. The proteasome is composed of a 2.2 MDa barrel-shaped multi-catalytic protease complex, the 26S proteasome. The 26S proteasome is the core component of the ubiquitin-proteasome, consisting of a 20S core complex responsible for protein hydrolysis and a 19S regulatory complex that recognizes polyubiquitinated substrates. The 20S proteasome is a cylindrical stack composed of four rings, two outer rings composed of 7 α subunits and two inner rings composed of 7 β subunits. Among these subunits, PSMB5 (proteasome β5 subunit) with chymotrypsin activity appears to be essential for the rate-limiting step of protein hydrolysis. It plays an important role in the process of antigen presentation and oxidative stress. Overexpression of PSMB5 can increase the resistance of cells to hydrogen peroxide-mediated cytotoxicity and protein oxidation. In addition, PSMB5 is the target of bortezomib, which has significant clinical activity in multiple myeloma. Mutations in the PSMB5 gene or upregulation of PSMB5 protein expression will lead to bortezomib resistance. PSMB5 mutations can cause conformational changes or spatial barriers in the drug binding site of multiple myeloma, thereby impairing the catalytic function of chymotrypsin in the 20S proteasome, indicating that PSMB5 point mutations can serve as a regulatory mechanism for drug resistance and tumor evolution in multiple myeloma. In addition, PSMB5 is significantly upregulated in prostate cancer. Overexpression of PSMB5 can enhance the proteomic degradation of regulatory proteins such as tumor suppressors and apoptosis regulators. Overexpression of miR-127-3p can inhibit the invasion and migration of prostate cancer cells in vitro by targeting PSMB5, ​​indicating that PSMB5 plays an important role in the invasion, metastasis and drug resistance of prostate cancer. Overexpression of PSMB5 in triple-negative breast cancer is associated with poor prognosis in patients. Downregulation of PSMB5 can induce apoptosis of triple-negative breast cancer cells and significantly increase the sensitivity of cancer cells to the chemotherapy drugs bortezomib and paclitaxel. The above research results show that PSMB5 can be used as a biomarker and therapeutic target for a variety of tumors. However, there are no reports on the expression of PSMB5 in bladder cancer in relevant literature at home and abroad.

[0004] In view of this, the following technical solution is proposed. Summary of the invention

[0005] A deep understanding of the molecular mechanism involved in the development of PSMB5 in bladder cancer is crucial to the development of new methods for the treatment of bladder cancer. The present invention found that the expression of the PSMB5 gene in bladder cancer tissue was significantly higher than that in adjacent tissues, and high expression of PSMB5 was associated with poor prognosis in bladder cancer patients. The effects of PSMB5 on the proliferation, invasion, migration and EMT transformation (epithelial-mesenchymal transformation) of bladder cancer may be mediated by the Wnt / β-catenin signaling pathway. Therefore, the present invention proposes a scientific hypothesis that PSMB5 mediates the proliferation and EMT transformation of bladder cancer cells through the Wnt / β-catenin signaling pathway.

[0006] The present invention includes the following technical solutions: In a first aspect, the present invention provides a use of a PSMB5 inhibitor in the preparation of a drug for bladder cancer mediated by the Wnt / β-catenin signaling pathway, wherein the drug has at least one of the following functions: a1) Inhibit the proliferation of bladder cancer cells; a2) Inhibit bladder cancer cell migration; a3) Inhibit bladder cancer cell invasion; a4) Inhibit spheroidization of bladder cancer cells; a5) Inhibit EMT transformation of bladder cancer cells.

[0007] The PSMB5 inhibitor includes a substance that reduces the expression of PSMB5 protein, the content of PSMB5 protein, or the activity of the protein, or a substance that inhibits the expression of the PSMB5 gene, including but not limited to one or more of nucleic acid molecules, small molecule compounds, antibodies, peptides, proteins, gene editing systems, lentiviruses, or adeno-associated viruses.

[0008] In some embodiments of the present invention, the PSMB5 inhibitor is a molecule that binds to the PSMB5 protein, such as one or a combination of two or more of an antibody, a peptide, or a small molecule, including but not limited to a humanized monoclonal antibody against the PSMB5 protein, a receptor antagonist with PSMB5 as a ligand, and the inhibitor can reduce the content or activity of the PSMB5 protein, or interfere with the binding of PSMB5 to its receptor.

[0009] In some embodiments of the present invention, the PSMB5 inhibitor is a short hairpin RNA (shRNA) or small interfering RNA (siRNA) targeting the PSMB5 gene; an adenovirus or lentivirus that implements PSMB5 gene knockout or low expression; a small molecule compound that interferes with the PSMB5 gene, and the inhibitor can reduce the expression of the gene encoding the PSMB5 protein.

[0010] In some embodiments of the present invention, the PSMB5 inhibitor is a gene editing system that can specifically knock out the PSMB5 gene, and the gene editing system is selected from the CRISPR-Cas system.

[0011] In a specific embodiment of the present invention, the PSMB5 inhibitor is selected from the short hairpin RNA (shRNA) shown in SEQ ID NO.1 or SEQ ID NO.2.

[0012] In a second aspect, the present invention provides a use of a biomarker PSMB5 and / or a substance for detecting PSMB5 in at least one of the following: b1) Use in the preparation and / or screening of products for diagnosis or auxiliary diagnosis of bladder cancer; b2) Application in the preparation and / or screening of products for evaluating or assisting in evaluating the therapeutic efficacy of bladder cancer.

[0013] The products include but are not limited to reagents, test kits, chips, test strips, membrane strips or detection platforms.

[0014] The substance for detecting PSMB5 includes any reagent required for detecting the protein expression amount or gene expression level of PSMB5 by RT-PCR method, RT-qPCR method, biochip detection method, Southern blotting method, protein blotting method (Western Blot), in situ hybridization method, immunohistochemistry (IHC), enzyme-linked immunosorbent assay (ELISA), and spatial transcriptome technology.

[0015] In some embodiments of the present invention, the substance for detecting PSMB5 is a reagent for detecting PSMB5 protein by immunohistochemistry.

[0016] In some embodiments of the present invention, the substance for detecting PSMB5 is a reagent for detecting PSMB5 protein by Western Blot.

[0017] In some embodiments of the present invention, the substance for detecting PSMB5 is a reagent for detecting the expression of nucleic acid related to PSMB5 protein, including but not limited to antibodies, polypeptides, proteins or nucleic acid molecules that bind to PSMB5 protein.

[0018] In a third aspect, the present invention provides a use of PSMB5 as a target in at least one of the following: c1) Use in the preparation and / or screening of drugs for preventing bladder cancer; c2) Application in the preparation and / or screening of drugs for the treatment or adjuvant treatment of bladder cancer.

[0019] In a fourth aspect, the present invention provides a use of a PSMB5 inhibitor in at least one of the following: d1) Use in the preparation and / or screening of drugs for preventing bladder cancer; d2) Use in the preparation and / or screening of drugs for the treatment or adjuvant treatment of bladder cancer.

[0020] The PSMB5 inhibitor includes a substance that reduces the expression of PSMB5 protein, the content of PSMB5 protein, or the activity of the protein, or a substance that inhibits the expression of the PSMB5 gene, including but not limited to one or more of nucleic acid molecules, small molecule compounds, antibodies, peptides, proteins, gene editing systems, lentiviruses, or adeno-associated viruses.

[0021] In some embodiments of the present invention, the PSMB5 inhibitor is a molecule that binds to the PSMB5 protein, such as one or a combination of two or more of an antibody, a peptide, or a small molecule, including but not limited to a humanized monoclonal antibody against the PSMB5 protein, a receptor antagonist with PSMB5 as a ligand, and the inhibitor can reduce the content or activity of the PSMB5 protein, or interfere with the binding of PSMB5 to its receptor.

[0022] In some embodiments of the present invention, the PSMB5 inhibitor is a short hairpin RNA (shRNA) or small interfering RNA (siRNA) targeting the PSMB5 gene; an adenovirus or lentivirus that implements PSMB5 gene knockout or low expression; a small molecule compound that interferes with the PSMB5 gene, and the inhibitor can reduce the expression of the gene encoding the PSMB5 protein.

[0023] In some embodiments of the present invention, the PSMB5 inhibitor is a gene editing system that can specifically knock out the PSMB5 gene, and the gene editing system is selected from the CRISPR-Cas system.

[0024] In a specific embodiment of the present invention, the PSMB5 inhibitor is selected from the short hairpin RNA (shRNA) shown in SEQ ID NO.1 or SEQ ID NO.2.

[0025] In a fifth aspect, the present invention provides a pharmaceutical composition comprising an effective amount of the PSMB5 inhibitor described in the present invention.

[0026] Preferably, the PSMB5 inhibitor is a short hairpin RNA (shRNA) targeting the PSMB5 gene.

[0027] Specifically, the PSMB5 inhibitor is selected from the short hairpin RNA (shRNA) shown in SEQ ID NO.1 or SEQ ID NO.2.

[0028] Furthermore, the pharmaceutical composition also includes pharmaceutically acceptable excipients.

[0029] The term "pharmaceutically acceptable" as used herein means that the substance or composition must be chemically and / or toxicologically compatible with the other ingredients of the formulation and / or the mammal to be treated with it. Preferably, the term "pharmaceutically acceptable" means approved by a federal regulatory agency or a national government or listed in the United States Pharmacopoeia or other generally recognized pharmacopoeia for use in animals, especially humans. The pharmaceutically acceptable excipients described herein may include any solvent, solid excipient, diluent or other liquid excipient, etc., suitable for a specific target dosage form.

[0030] The pharmaceutical composition of the present invention can be used for the treatment of diseases and can also be used for in vitro cell culture experiments. When used for the treatment of diseases, the pharmaceutical composition generally refers to a unit dosage form and can be prepared by any of the methods well known in the pharmaceutical field. All methods include the step of combining the active ingredient with an excipient that constitutes one or more auxiliary ingredients.

[0031] The disease treatment refers to administering a therapeutically effective amount of the PSMB5 inhibitor or pharmaceutical composition of the present invention to a subject in need thereof.

[0032] Furthermore, the disease treatment also includes administering the PSMB5 inhibitor or pharmaceutical composition of the present invention in combination with other conventional agents or conventional treatment methods in the art.

[0033] In a sixth aspect, the present invention provides a use of a biomarker PSMB5 and / or a substance for detecting PSMB5 in the preparation and / or screening of products for evaluating the prognosis and survival of bladder cancer patients.

[0034] The product includes but is not limited to a reagent, a test kit, a chip, a test paper, a membrane strip or a detection platform. The substance for detecting PSMB5 is as described in the second aspect of the present invention.

[0035] The present invention aims to clarify the mechanism of action of inhibiting PSMB5 expression in regulating bladder cancer in the Wnt / β-catenin signaling pathway, so that the proliferation cycle of bladder cancer cells stagnates at a certain period or mediates bladder cancer cell apoptosis, thereby preventing the growth of bladder cancer cells and delaying the further development of bladder cancer, thereby providing a new therapeutic target for the treatment of bladder cancer. BRIEF DESCRIPTION OF THE DRAWINGS

[0036] Figure 1 Scientific hypothesis diagram of Wnt / β-catenin signaling pathway.

[0037] Figure 2 The expression of genes related to the Wnt / β-catenin signaling pathway, cell cycle and EMT in cancer and adjacent normal tissues using different immune protein antibodies.

[0038] Figure 3 Cell mechanism experiments showed that after PSMB5 knockdown, the Wnt / β-Catenin pathway signal was significantly weakened.

[0039] Figure 4 Correlation of PSMB5 expression with clinical characteristics of bladder cancer patients; (A) Expression of PSMB5 in bladder cancer and adjacent tissues; (BJ) Expression levels of PSMB5 in patients with different clinical characteristics: T stage (B), N stage (C), radiotherapy (D), initial treatment (E), tissue grade (F), classification (G), OS events (H), DSS events (I), PFI events (J); (K) Area under the ROC curve of PSMB5 in bladder cancer.

[0040] Figure 5 The expression of PSMB5 protein in bladder cancer and adjacent tissues, and the tissue morphology of normal urothelium and bladder cancer.

[0041] Figure 6 The expression of PSMB5 protein in bladder cancer and adjacent tissues, the expression of PSMB5 in non-muscle invasive bladder cancer and adjacent tissues.

[0042] Figure 7 Expression of PSMB5 protein in bladder cancer and adjacent tissues; Expression of PSMB5 in muscle-invasive bladder cancer and adjacent tissues.

[0043] Figure 8 WB experiment detected the expression level of PSMB5; (A) shows the expression level of PSMB5 in each cell line; (B) shows that the expression level of PSMB5 in 5637 cells was significantly decreased after shRNA knockdown.

[0044] Fig. 9 Cell proliferation experiment; (A) CCK-8 results showed that after knocking down PSMB5, ​​the dynamic proliferation ability of cells was weakened at an absorbance of 450; (B) EDU showed that the proportion of EDU+ fluorescent cells decreased significantly after knocking down PSMB5; (C) is the quantification of (B); (D) The clone formation experiment suggested that the cell sphere-forming ability was reduced; (E) is the quantification of (D); (*p<0.05, **p<0.01, ***p<0.001).

[0045] Fig.10 Transwell invasion assay of 5637 cell line after knockdown of PSMB5.

[0046] Fig.11 Flow cytometry experiment of 5637 cell line after knockdown of PSMB5.

[0047] Fig.12 Kaplan-Meier survival analysis was performed on the prognosis of PSMB5 expression in bladder cancer: (A) Overall survival rate of PSMB5 expression in bladder cancer; (BH) Survival rate of PSMB5 expression in different subgroups of bladder cancer: T2 / T3 stage (B), T1 / T3 stage (C), pathological stage (D), N stage (E), gender (F), tissue grade (G), and smoking history (H). DETAILED DESCRIPTION

[0048] The technical solutions in the embodiments of the present invention are described clearly and completely below. Obviously, the described embodiments are only some embodiments of the present invention, not all. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0049] The clinical studies involved in this invention were approved by the Human Subjects Ethics Committee, the research protocols followed the guidelines set by the Declaration of Helsinki, and written informed consent was obtained from all subjects. The present invention proposes the following research conclusions: 1. PSMB5 mediates bladder cancer progression through the Wnt / β-Catenin signaling pathway In terms of mechanism, the present invention uses the TCGA database to perform GO function enrichment and KEGG pathway enrichment on PSMB5-related genes. The results show that PSMB5 is significantly enriched in the Wnt signaling pathway and is in a positive regulatory relationship. In addition, we performed GSEA data enrichment analysis on the above gene set, and the results further verified that PSMB5 is significantly enriched in the Wnt signaling pathway. The present invention uses immunohistochemistry experiments to confirm that PSMB5 is significantly correlated with the Wnt / β-catenin signaling pathway, cell cycle, and bladder cancer cell EMT. After knocking down PSMB5, ​​it was found that the Wnt / β-Catenin pathway signal was significantly weakened, and it is considered that PSMB5 regulates the biological process of bladder cancer through the Wnt / β-Catenin axis ( Figure 1 ).

[0050] 1.1 Immunohistochemical phenotype markers of Wnt / β-catenin signaling pathway, cell cycle and EMT-related genes 90 tissue samples were selected, including 45 cancer tissues and 45 paracancerous tissues, and immunohistochemical labeling was performed on the β-catenin gene, C-myc, E-Cadherin in the Wnt / β-catenin signaling pathway, and cell cycle Cyclin D1, Cyclin B1, CDK1, CDK2, EMT, N-Car, E-Car, Snail, and Vimentin to show the role of PSMB5 in the occurrence and development of bladder cancer. According to strong positive, moderately positive, weakly positive, and negative, the scores were divided into 3 points, 2 points, 1 point, and 0 points for statistical analysis (Table 1). By analyzing the data of different antibodies in cancer tissues and adjacent tissues, it was found that the expression of β-catenin, C-myc, E-Cadherin, Cyclin D1, Cyclin B1, CDK1, CDK2, EMT, N-Car, E-Car, Snail, and Vimentin in cancer was better than that in adjacent tissues, and the difference was statistically significant (P<0.05) ( Figure 2 ). The above data showed that PSMB5 was significantly correlated with the Wnt / β-catenin signaling pathway, cell cycle, and EMT of bladder cancer cells.

[0051] Table 1 Expression of different antibodies in cancer and adjacent tissues 1.2 Changes in the Wnt pathway in 5637 cells after knockdown of PSMB5 The results are as follows Figure 3 As shown in the figure, after PSMB5 knockdown, the Wnt / β-Catenin pathway signal was significantly weakened, which confirmed that PSMB5 regulates the biological process of bladder cancer through the Wnt / β-Catenin axis.

[0052] PSMB5 as a biomarker for bladder cancer diagnosis The inventors found that the expression of PSMB5 mRNA in bladder cancer was significantly higher than that in adjacent tissues. Immunohistochemistry was used to detect the expression of PSMB5 in 61 cases of bladder cancer and normal tissues. The results showed that the expression of PSMB5 gene in bladder cancer was significantly higher than that in adjacent tissues (P<0.01). Therefore, PSMB5 can be used as a biomarker for the diagnosis of bladder cancer.

[0053] 2.1 High expression of PSMB5 in bladder cancer patients The expression level of PSMB5 mRNA in bladder cancer patients was further evaluated using the TCGA database and compared with that in adjacent paracancerous tissues. The results further confirmed that the expression of PSMB5 in bladder cancer was significantly higher than that in adjacent paracancerous tissues (P<0.01). Figure 4A). Subsequently, we analyzed the expression level of PSMB5 mRNA in different clinical categories in the TCGA database. The results showed that high expression of PSMB5 was significantly correlated with T stage, N stage, radiotherapy, initial treatment results, tissue grade, typing, OS events, DSS events, and PFI events in bladder cancer patients (P<0.01) ( Figure 4 BJ). Analyzing the ROC curve, the AUC is 0.836 ( Figure 4 K), the results showed that PSMB5 has a high accuracy in diagnosing bladder cancer.

[0054] 2.2 Correlation between PSMB5 and clinical characteristics of bladder cancer patients In this study, 61 patients with radical bladder urothelial carcinoma who underwent surgical resection and were pathologically diagnosed with radical bladder urothelial carcinoma and signed the informed consent form were selected for PSMB5 immunohistochemical expression detection, and the correlation between their clinical pathological characteristics was summarized (Table 2).

[0055] Table 2 Correlation between PSMB5 expression and clinical characteristics of bladder cancer 2.3 Immunohistochemistry confirmed that the expression of PSMB5 protein in bladder cancer was higher than that in adjacent tissues Radical specimens from 61 patients with bladder cancer, including bladder cancer tissue and adjacent tissue, were fixed with 4% paraformaldehyde and stained with eosin and hematoxylin ( Figure 5 ). Immunohistochemistry was used to detect the expression of PSMB5 in bladder cancer and adjacent tissues. The results showed that the expression of PSMB5 gene in bladder cancer was significantly higher than that in adjacent tissues, including 28 cases in the high expression group and 33 cases in the low expression group. The immunohistochemical expression test results were consistent with the results in the database ( Figure 6 , Figure 7 ).

[0056] 2.4 WB detection of PSMB5 expression in various bladder cancer cell lines The expression of PSMB5 in bladder cancer cell lines 5637, J82, T24 and UM-UC-3 and the control group ureteral epithelial cell SV-HUC-1 was detected. The study found that compared with normal cell lines, the expression of PSMB5 was generally higher, especially in 5637 and J82 ( Figure 8 A). PSMB5 in 5637 cells was knocked down by shRNA shown in SEQ ID NO.1 and SEQ ID NO.2. The results are shown in Figure 8 As shown in B, the expression level of PSMB5 decreased significantly.

[0057] 3. PSMB5 can be used as a target for bladder cancer treatment The expression of PSMB5 in various bladder cancer cell lines was detected by WB, and it was found that the expression of PSMB5 was generally high, especially in 5637 and J82. Subsequently, the CCK-8 experiment found that after knocking down PSMB5, ​​the dynamic proliferation ability of cells was weakened at 450nm absorbance; the EdU experiment showed that the proportion of EDU+ fluorescent cells decreased significantly after knocking down PSMB5, ​​and the bulge formation experiment indicated that the cell sphere formation ability decreased. This shows that PSMB5 can be used as a target for the treatment of bladder cancer.

[0058] 3.1 Changes in proliferation ability of 5637 cells after knockdown of PSMB5 The results are as follows Fig. 9 As shown, after knocking down PSMB5, ​​the dynamic proliferation ability of cells was weakened and the cell sphere-forming ability was decreased.

[0059] 3.2 Transwell invasion assay for detecting PSMB5 gene After knocking down 5637, the number of cell migration in the two knockdown groups was significantly reduced, indicating that knocking down PSMB5 significantly inhibited the motility of bladder cancer cell lines. This result is similar to the bioinformatics prediction ( Fig.10 ).

[0060] 3.3 Detection of PSMB5 gene flow cytometry After knocking down PSMB5, ​​the number of apoptotic cells in bladder cancer cell line 5637 increased significantly, indicating that the expression of PSMB5 maintains the survival of tumor cells ( Fig.11 ).

[0061] 4. PSMB5 can be used as a marker for the prognosis and survival of bladder cancer patients The present invention found that high expression of PSMB5 was associated with poor prognosis of bladder cancer patients through Kaplan-Meier survival analysis, and high expression of PSMB5 was significantly correlated with poor prognosis of bladder cancer, such as T stage, N stage, pathological grade, and histological grade. Single and multivariate CoX regression analysis showed that there was a correlation between high expression of PSMB5 and poor OS in bladder cancer patients. These data show that high expression of PSMB5 is an independent risk prognostic factor for OS in bladder cancer patients.

[0062] 4.1 High expression of PSMB5 is an independent factor for prognosis and survival in bladder cancer patients To determine whether PSMB5 expression affects patient survival, we divided bladder cancer patients in the TCGA database into high PSMB5 expression group and low PSMB5 expression group, and performed Kaplan-Meier survival analysis. The results showed that high expression of PSMB5 was associated with poor prognosis in bladder cancer patients (HR=1.44, p=0.015) ( Fig.12A). In addition, subgroup analysis showed that high expression of PSMB5 was significantly associated with poor prognosis of bladder cancer, including T stage, N stage, pathological grade, histological grade, smoking history, and gender ( Fig.12 BH). Single and multivariate CoX analysis showed that high expression of PSMB5 was associated with poor OS in patients with bladder cancer. These data suggest that high expression of PSMB5 is an independent prognostic factor for OS in patients with bladder cancer (Table 3).

[0063] Table 3 Univariate and multivariate CoX regression analysis of clinical characteristics associated with overall survival in bladder cancer Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the aforementioned embodiments, or replace some or all of the technical features therein with equivalents. However, these modifications or replacements do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. Use of a PSMB5 inhibitor in the preparation of a drug for bladder cancer mediated by the Wnt / β-catenin signaling pathway, characterized in that: The drug has at least one of the following functions: a1) Inhibit the proliferation of bladder cancer cells; a2) Inhibit bladder cancer cell migration; a3) Inhibit bladder cancer cell invasion; a4) Inhibit spheroidization of bladder cancer cells; a5) Inhibit EMT transformation of bladder cancer cells.

2. The use according to claim 1, characterized in that: The PSMB5 inhibitor includes a substance that reduces the expression amount of PSMB5 protein or the content of PSMB5 protein or the activity of the protein, or a substance that inhibits the expression of the PSMB5 gene; the substance includes one or more of nucleic acid molecules, small molecule compounds, antibodies, polypeptides, proteins, gene editing systems, lentiviruses or adeno-associated viruses.

3. Use of the biomarker PSMB5 and / or a substance for detecting PSMB5 in at least one of the following: b1) Use in the preparation and / or screening of products for diagnosis or auxiliary diagnosis of bladder cancer; b2) Application in the preparation and / or screening of products for evaluating or assisting in evaluating the therapeutic efficacy of bladder cancer.

4. The use according to claim 3, characterized in that: The substances for detecting PSMB5 include reagents required for detecting the protein expression amount or gene expression level of PSMB5 by RT-PCR method, RT-qPCR method, biochip detection method, Southern blotting method, protein blotting method, in situ hybridization method, immunohistochemistry method, enzyme-linked immunosorbent assay, and spatial transcriptome technology.

5. Use of PSMB5 as a target in at least one of the following: c1) Use in the preparation and / or screening of drugs for preventing bladder cancer; c2) Application in the preparation and / or screening of drugs for the treatment or adjuvant treatment of bladder cancer.

6. Use of a PSMB5 inhibitor in at least one of the following: d1) Use in the preparation and / or screening of drugs for preventing bladder cancer; d2) Use in the preparation and / or screening of drugs for the treatment or adjuvant treatment of bladder cancer.

7. The use according to claim 6, characterized in that: The PSMB5 inhibitor includes a substance that reduces the expression amount of PSMB5 protein or the content of PSMB5 protein or the activity of the protein, or a substance that inhibits the expression of the PSMB5 gene; the substance includes one or more of nucleic acid molecules, small molecule compounds, antibodies, polypeptides, proteins, gene editing systems, lentiviruses or adeno-associated viruses.

8. A pharmaceutical composition, characterized in that The pharmaceutical composition comprises an effective amount of the PSMB5 inhibitor according to claim 1.

9. The pharmaceutical composition according to claim 8, characterized in that The PSMB5 inhibitor is a short hairpin RNA targeting the PSMB5 gene.

10. Use of the biomarker PSMB5 and / or a substance for detecting PSMB5 in the preparation and / or screening of products for evaluating the prognosis and survival of bladder cancer patients.

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