Application of PTPRG as marker and therapeutic target for diagnosing colorectal cancer

Abstract

The invention discloses application of PTPRG as a marker and a therapeutic target for diagnosing colorectal cancer, and belongs to the technical field of biological medicines, reagents for detecting the expression level of PTPRG comprise a reagent for detecting the expression level of PTPRG gene mRNA and / or a reagent for detecting the expression level of PTPRG protein. The invention also provides application of PTPRG as a regulation target spot in preparation of medicines for treating colorectal cancer. The application has the beneficial effects that a new therapeutic target for colorectal cancer is provided, a new thought is provided for treatment of colorectal cancer, it is revealed that liquiritin inhibits miR-23b to promote expression of PTPRG in a targeted mode, then the effect of preventing and treating colorectal cancer is achieved, and the application prospect and economic value are wide.

Description

Technical Field

[0001] The present invention belongs to the technical field of biomedicine, and particularly relates to the use of PTPRG as a marker for diagnosing colorectal cancer and a therapeutic target. Background Art

[0002] Colorectal cancer is one of the most common malignant tumors worldwide and ranks second in cancer-related deaths, accounting for approximately 10% of global cancer deaths. Despite significant progress in the diagnosis and treatment of colorectal cancer in modern medicine, its high incidence and high mortality remain a major challenge to global public health. The occurrence of colorectal cancer is closely related to genetic susceptibility and environmental factors. Studies have shown that unhealthy lifestyle factors such as high-fat diet, lack of exercise, obesity, smoking, and alcohol consumption are the main inducements for colorectal cancer. Traditionally, this disease mainly affects people over 50 years old, and the incidence is higher in men than in women. However, in recent years, the incidence of colorectal cancer has shown a trend of younger age, especially in countries with rapid economic development, where the incidence has increased significantly, suggesting that changes in the environment and lifestyle may be the key factors. Although surgery, radiotherapy, chemotherapy, and targeted therapy have become the main treatment strategies for colorectal cancer, due to the atypical early symptoms, many patients are diagnosed at an advanced stage or have distant metastases, resulting in increased treatment difficulty. In addition, traditional chemotherapy drugs (such as 5-FU, oxaliplatin, etc.) have significant toxic side effects and are prone to inducing drug resistance, seriously affecting the prognosis and quality of life of patients. Therefore, finding new treatment strategies with low toxicity and high efficiency, especially targeting key molecules that regulate the progression of colorectal cancer, has become a hot topic in current research.

[0003] Protein tyrosine phosphatase receptor G (PTPRG) is a member of the protein tyrosine phosphatase family and mainly participates in regulating biological processes such as cell growth, proliferation, differentiation, and tumor transformation, and acts as a tumor suppressor factor in various cancers such as renal cancer, gastric cancer, and breast cancer.

[0004] Liquiritin is an important monomeric active ingredient in licorice flavonoids and has various pharmacological effects such as antioxidant and anti-HIV effects. Currently, there is no report on the inhibition of miR-23b targeting to promote the expression of PTPRG by liquiritin and further preventing and treating colorectal cancer. Summary of the Invention

[0005] The technical problem to be solved by the present invention is how to propose a new therapeutic target for colorectal cancer and a new use of liquiritin.

[0006] The present invention solves the above technical problem by the following technical means:

[0007] The first aspect of the present invention proposes the use of a reagent for detecting the expression level of PTPRG in the preparation of a product for diagnosing colorectal cancer.

[0008] Preferably, the reagent for detecting the expression level of PTPRG includes a reagent for detecting the mRNA expression level of the PTPRG gene and / or a reagent for detecting the protein expression level of PTPRG.

[0009] Preferably, the reagent for detecting the protein expression level of PTPRG includes an antibody against the PTPRG protein.

[0010] Preferably, the product includes a kit, a chip or a test strip.

[0011] The second aspect of the present invention provides the application of PTPRG as a regulatory target in the preparation of drugs for treating colorectal cancer.

[0012] The third aspect of the present invention provides the application of a reagent for promoting the expression of PTPRG in the preparation of drugs for treating colorectal cancer.

[0013] The fourth aspect of the present invention provides the application of liquiritin in the preparation of drugs for preventing and / or treating colorectal cancer by inhibiting miR-23b to target and promote the expression of PTPGR.

[0014] Preferably, the drug further contains a pharmaceutically acceptable carrier.

[0015] Preferably, the pharmaceutically acceptable carrier is selected from one or more of diluents, excipients, fillers, binders, wetting agents, disintegrants, absorption promoters, surfactants, adsorption carriers and lubricants.

[0016] Preferably, the drug is made into a pharmaceutically acceptable dosage form.

[0017] The beneficial effects of the present invention are as follows:

[0018] The present invention provides a new therapeutic target for colorectal cancer, provides a new idea for the treatment of colorectal cancer, and reveals that liquiritin inhibits miR-23b to target and promote the expression of PTPRG, thereby playing a role in preventing and treating colorectal cancer, and has broad application prospects and economic value. Description of the Drawings

[0019] Figure 1 It is a diagram of the prediction and verification results of miR-23b target protein in Example 1 of the present invention. Among them, A is a prediction diagram of the direct interaction between miR-23b and the 3'UTR of PTPRG mRNA, and B-C are diagrams of relative luciferase activity analysis;

[0020] Figure 2 It is a diagram of the effect of CCK-8 detection of PTPRG downregulation on the proliferation of SW480 cells in Example 2 of the present invention. Among them, A is a diagram of the PTPRG mRNA expression level, and B is a diagram of the change in the proliferation level of SW480 cells;

[0021] Figure 3 This is the figure showing the effect of detecting the down - regulation of PTPRG on the proliferation of SW480 cells in Example 2 of the present invention. Among them, A is the figure of detecting cell proliferation by EdU, and B is the statistical analysis figure of cell proliferation;

[0022] Figure 4 This is the figure showing the effect of the down - regulation of PTPRG on the migration of SW480 cells in Example 2 of the present invention. Among them, A is the cell migration figure, and B is the statistical analysis figure of cell migration;

[0023] Figure 5 This is the figure showing that overexpression of PTPRG reverses the down - regulation of PTPRG mediated by miR - 23b in Example 2 of the present invention. Among them, A is the protein band figure of PTPRG, and B is the statistical analysis figure of the relative expression of PTPRG protein;

[0024] Figure 6 This is the figure showing the migration ability of SW480 cells in which overexpression of PTPRG reverses the miR - 23b - mediated effect in Example 2 of the present invention;

[0025] Figure 7 This is the figure showing the proliferation ability of SW480 cells in which overexpression of PTPRG reverses the miR - 23b - mediated effect in Example 2 of the present invention. On the left of the figure is the figure of detecting cell proliferation by EdU, and on the right is the statistical analysis figure of cell proliferation;

[0026] Figure 8 This is the figure showing that liquiritin dose - dependently increases the expression level of PTPRG in SW480 cells in Example 3 of the present invention. Among them, A is the figure of the expression level of PTPRG mRNA, B is the figure of the expression level of PTPRG protein, and C is the figure of detecting the expression level of PTPRG by immunofluorescence;

[0027] Figure 9 This is the figure showing that liquiritin inhibits the expression level of miR - 23b in SW480 cells in Example 3 of the present invention. Among them, A is the figure of the effect of liquiritin on the expression level of miR - 23b under the condition of overexpression of miR - 23b, and B is the figure of the effect of liquiritin on the expression level of miR - 23b under the condition of inhibition of miR - 23b;

[0028] Figure 10This is the figure showing that liquiritin inhibits the expression of miR-23b and increases the level of PTPRG in SW480 cells in Example 3 of the present invention. Among them, A is the figure showing the effect of liquiritin on the expression level of PTPRG mRNA under the condition of overexpression of miR-23b, B is the figure showing the effect of liquiritin on the expression level of PTPRG mRNA under the condition of inhibition of miR-23b, C is the figure showing the effect of liquiritin on the expression level of PTPRG protein under the condition of overexpression of miR-23b, and D is the figure showing the effect of liquiritin on the expression level of PTPRG protein under the condition of inhibition of miR-23b;

[0029] Figure 11 This is the figure showing the analysis of in vivo imaging of the inhibition of the expression of miR-23b in the tumor tissues of nude mouse xenografts by liquiritin in Example 4 of the present invention;

[0030] Figure 12 This is the figure showing the comparison of the sizes of the tumor tissues of nude mouse xenografts in each group in Example 4 of the present invention;

[0031] Figure 13 This is the figure showing the effect of liquiritin on the growth of nude mouse xenografts mediated by miR-23b in Example 4 of the present invention. Among them, A is the figure showing the change in body weight of nude mice in each group, and B is the figure showing the change in tumor mass of nude mice in each group;

[0032] Figure 14 This is the figure showing the in vivo reversal effect of liquiritin on the down-regulation of PTPRG mediated by miR-23b in Example 4 of the present invention. The upper figure is the PTPRG protein band figure, and the lower figure is the statistical analysis figure of the relative expression of PTPRG protein. Detailed implementation manners

[0033] To make the objectives, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below in conjunction with the embodiments of the present invention. Obviously, the described embodiments are part of the embodiments of the present invention, rather than all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative efforts shall fall within the protection scope of the present invention.

[0034] The test materials and reagents used in the following embodiments can be obtained from commercial channels without special instructions.

[0035] For those not specifying specific technologies or conditions in the embodiments, they can all be carried out according to the technologies or conditions described in the literature in the field or according to the product specifications. Without special instructions, the quantitative tests in the following embodiments are all set with more than three repeated experiments, and the results are averaged.

[0036] Experimental materials

[0037] 1. Animals

[0038] Twelve BALB / c nude mice, weighing approximately 24 - 26 g, were purchased from Zhejiang Vital River Laboratory Animal Technology Co., Ltd. All mice were housed in an SPF - level animal room, maintained with a 12 - hour light - dark cycle, the indoor temperature was kept at 18 - 22 °C, and the relative humidity was 40 - 60%. They had free access to food and water. All animal experiments were preceded by acclimating the animals in the animal room for 7 days. All animal experimental procedures were approved by the Animal Protection and Utilization Committee of Suzhou University before implementation. All research protocols were designed in accordance with the "Guidelines for the Principles of Laboratory Animal Protection and Utilization" (No. SZULL - 01 - 002) of Suzhou University.

[0039] 2. Cells

[0040] The SW480 cell line was obtained from the Shanghai Institute of Life Sciences, Chinese Academy of Sciences. The cells were cultured in DMEM medium containing 10% FBS and 1% penicillin / streptomycin, and placed in a 37 °C incubator containing 5% CO 2 ₂.

[0041] Example 1:

[0042] Prediction and verification of miR - 23b target proteins

[0043] (1) Bioinformatics analysis

[0044] A multi - database joint analysis strategy was used to explore the regulatory mechanism of miR - 23b in colorectal cancer. First, gene datasets related to colorectal cancer were obtained from the DisgeNet database (https: / / www.disgenet.org / search); second, the mRNA - omics analysis results were integrated with genes related to colorectal cancer; subsequently, potential target genes of miR - 23b were screened through the STARBASE database (https: / / starbase.sysu.edu.cn / ), and miR - 23b - regulated genes related to colorectal cancer were determined. To verify the regulatory relationship between miR - 23b and the candidate target gene PTPRG, the potential binding sites of miR - 23b in the 3'UTR region of PTPRG mRNA were further predicted using the STARBASE and TargetScan databases (https: / / www.targetscan.org / vert_72).

[0045] (2) Luciferase reporter gene

[0046] To verify the targeted regulatory relationship between miR-23b and PTPRG, a luciferase reporter gene vector was constructed. First, the 800bp fragment of the PTPRG 3'-UTR region containing the miR-23b binding site was cloned into the pMIR-REPORT luciferase reporter vector. At the same time, a wild-type (WT) luciferase reporter vector was designed and constructed as a control. To further verify the specificity of the binding site, the miR-23b interaction core sequence AUGUGA was site-directed mutated to UACACU to construct a mutant (MUT) luciferase reporter vector.

[0047] After SW480 cells were inoculated in 24-well plates and cultured to an appropriate density, 0.2 μg of luciferase reporter plasmid (WT or MUT) and 0.2 μg of β-galactosidase (β-gal) internal reference plasmid and 50 pmol of miR-23b mimic (miR-23bmimic) or negative control (NC) were co-transfected into the cells, and Lipofectamine 2000 transfection reagent was used in the transfection process. To evaluate the transfection efficiency, a single transfection of β-gal plasmid was set as a control. After 24 hours of transfection, luciferase activity was detected using a luciferase reporter gene assay kit, and β-gal activity was used as an internal reference for standardization.

[0048] The results showed that miR-23b had a direct interaction with the 3'UTR of PTPRG mRNA ( Figure 1 A).

[0049] For validation, we constructed luciferase reporter plasmids containing the PTPRG 3'-UTR fragment and the mutated PTPRG 3'-UTR fragment, and then introduced the corresponding miR-23b-mimics into SW480 cells.

[0050] The results showed that miR-23b overexpression significantly reduced luciferase activity, while after mutation of the miR-23b binding site in the PTPRG 3'-UTR fragment, the luciferase activity remained unchanged even after miR-23b overexpression compared with the negative control group ( Figure 1 BC). These results suggest that there is a connection between the 3' non-coding region of the PTPRG gene and miR-23b.

[0051] Embodiment 2:

[0052] In vitro confirmation that PTPRG is a key target protein for inhibiting the occurrence and development of colorectal cancer

[0053] 1. PTPRG downregulation promotes the proliferation of SW480 colorectal cancer cells

[0054] (1) CCK-8 method

[0055] To investigate the functional role of PTPRG in SW480 colorectal cancer cells, siRNA (siPTPRG) was used to interfere with the expression of PTPRG, and qRT-PCR and CCK-8 assays were performed to detect the PTPRG mRNA level and cell proliferation ability, respectively.

[0056] The results are as Figure 2 shown in A. Compared with the control group (siRNA-NC), the PTPRG mRNA level in the siPTPRG group was significantly decreased (P<0.01), indicating that siPTPRG effectively inhibited the expression of PTPRG. The CCK-8 cell proliferation assay showed ( Figure 2 B) that the proliferation ability of SW480 cells was significantly enhanced after siPTPRG treatment. At the 48 h and 72 h time points, the OD values of the siPTPRG group were higher than those of the siRNA-NC group, suggesting that PTPRG may play an inhibitory role in cell proliferation in SW480 cells.

[0057] (2) EdU assay

[0058] To further investigate the role of PTPRG in the proliferation of SW480 colorectal cancer cells, an EdU staining assay was used to evaluate the effect of siRNA-mediated downregulation of PTPRG (siPTPRG) on the proliferation of SW480 cells. The results are as Figure 3 shown in A. Compared with the control group (siRNA-NC), the number of EdU-positive cells (red) in the siPTPRG group was significantly increased, indicating that the proliferation ability of SW480 cells was enhanced after PTPRG downregulation; the results of statistical analysis showed ( Figure 3 B) that the cell proliferation rate of the siPTPRG group was significantly higher than that of the siRNA-NC group (P<0.05). The experimental results indicated that PTPRG, as a tumor suppressor gene, plays a key role in the proliferation process of colorectal cancer cells. Combining the previous experimental results that miR-23b promotes the proliferation of SW480 cells by downregulating PTPRG, it is speculated that miR-23b may promote the proliferation of SW480 cells by negatively regulating PTPRG.

[0059] 2. Downregulation of PTPRG promotes the migration ability of SW480 colorectal cancer cells

[0060] To investigate the role of PTPRG in the migration of SW480 colorectal cancer cells, a Transwell migration assay was used to evaluate the effect of siRNA-mediated downregulation of PTPRG (siPTPRG) on the migration ability of SW480 cells.

[0061] The results are as Figure 4As shown in Figure A. The Transwell results showed that compared with the control group (siRNA-NC), the number of migrating cells in the siPTPRG group was significantly increased, and the number of cells penetrating the membrane was significantly increased. The results of statistical analysis further confirmed ( Figure 4 B) that the number of migrating cells in the siPTPRG group was significantly higher than that in the siRNA-NC group (P<0.01), indicating that down-regulation of PTPRG could promote the migration ability of SW480 cells. The experimental results showed that PTPRG, as a tumor suppressor gene, played an important role in the migration process of colorectal cancer cells. Combining the above experimental results of miR-23b negatively regulating PTPRG, it was speculated that miR-23b might promote the migration ability of SW480 cells by inhibiting PTPRG.

[0062] 3. Overexpression of PTPRG reverses miR-23b-mediated down-regulation of PTPRG

[0063] To explore whether PTPRG was a direct target of miR-23b and to evaluate whether overexpression of PTPRG could reverse the inhibitory effect of miR-23b-mediated PTPRG, Western blot was used to detect the protein level of PTPRG.

[0064] The results were as Figure 5 shown in Figure A. Compared with the control group (NC), overexpression of miR-23b (miR-23b group) significantly decreased the protein level of PTPRG, while after co-transfection with the PTPRG overexpression vector (miR-23b+PTPRG vector group), the expression level of PTPRG was partially restored. The results of statistical analysis ( Figure 5 B) further confirmed that overexpression of miR-23b significantly decreased the protein expression of PTPRG (P<0.05), while overexpression of PTPRG could partially reverse this inhibitory effect. The research results showed that miR-23b promoted the progression of colorectal cancer by negatively regulating PTPRG, and overexpression of PTPRG could effectively restore its expression level, further supporting that PTPRG might be a direct target of miR-23b and play a tumor suppressor role in colorectal cancer.

[0065] 4. Overexpression of PTPRG reverses miR-23b-mediated migration ability of SW480 cells

[0066] To explore whether overexpression of PTPRG could reverse the enhanced migration effect of miR-23b-mediated SW480 cells, Transwell migration assay was used to evaluate the effects of overexpression of miR-23b (miR-23b) and co-transfection with the PTPRG overexpression vector (miR-23b+PTPRG vector) on the migration ability of SW480 cells. The results were as Figure 6As shown, Transwell results showed that the number of migratory cells in the miR-23b overexpression group increased significantly, while overexpression of PTPRG could partially reverse the pro-migratory effect mediated by miR-23b, reducing the number of migratory cells. Statistical analysis results further indicated that overexpression of miR-23b significantly promoted the migration of SW480 cells (P<0.05), while overexpression of PTPRG could significantly inhibit the migration-enhancing effect mediated by miR-23b. These results suggest that PTPRG may serve as a direct target of miR-23b and play a tumor-suppressive role in colorectal cancer by negatively regulating cell migration, further verifying the functional role of miR-23b in promoting the migration of colorectal cancer cells.

[0067] 5. Overexpression of PTPRG reverses the proliferative ability of miR-23b-mediated SW480 cells

[0068] To investigate whether PTPRG can reverse the proliferative effect of miR-23b-mediated SW480 cells, an EdU cell proliferation assay was used to evaluate the effects of miR-23b overexpression (miR-23b) and co-transfection of the PTPRG overexpression vector (miR-23b + PTPRG vector) on the proliferation of SW480 cells.

[0069] The results were as Figure 7 shown. Compared with the control group (NC), the number of EdU-positive cells (red-labeled) in the miR-23b overexpression group increased significantly, suggesting that miR-23b could promote the proliferation of SW480 cells. In the PTPRG overexpression group (miR-23b + PTPRG vector), the number of EdU-positive cells decreased, indicating that overexpression of PTPRG could partially reverse the pro-proliferative effect mediated by miR-23b. Statistical analysis results further confirmed that overexpression of miR-23b significantly increased the proliferation rate of SW480 cells (P<0.05), while overexpression of PTPRG could significantly inhibit the cell proliferation-enhancing effect mediated by miR-23b. The research results showed that miR-23b promoted the proliferation of SW480 cells by downregulating PTPRG, while overexpression of PTPRG could effectively weaken the pro-proliferative effect mediated by miR-23b.

[0070] Example 3:

[0071] Liquiritin inhibits the tumor growth of SW480 cells through the miR-23b-PTPRG pathway

[0072] 1. Liquiritin can regulate the expression level of PTPRG in SW480 cells

[0073] To investigate whether liquiritin can regulate the expression level of PTPRG in SW480 cells and participate in the progression of colorectal cancer, qRT-PCR technology, Western blot and immunofluorescence experiments were used to examine the expression level of PTPRG in SW480 cells. The results are as Figure 8 shown that liquiritin can dose-dependently increase the expression levels of PTPRG mRNA and protein (P < 0.01). The positive drug oxaliplatin group also significantly increased the expression levels of PTPRG mRNA and protein (P < 0.01), and the high-dose liquiritin group was significantly better than the positive drug oxaliplatin group in increasing the expression levels of PTPRG mRNA and protein. It is shown that liquiritin dose-dependently increases the gene and protein expression levels of PTPRG.

[0074] 2. Liquiritin can regulate the expression level of miR-23b in SW480 cells

[0075] To investigate whether liquiritin (Liq) can regulate the expression level of miR-23b in SW480 cells and participate in the progression of colorectal cancer, the effects of liquiritin on the expression level of miR-23b in colorectal cancer cells were investigated under the conditions of miR-23b overexpression (mimics) and inhibition (inhibitor). The results are as Figure 9 shown that compared with the control group, the level of miR-23b in the miR-23b mimics group was significantly increased (P < 0.01); compared with the miR-23b mimics group, liquiritin treatment (miR-23b mimics + Liq) could significantly reduce the level of miR-23b (P < 0.05). At the same time, compared with the control group, the level of miR-23b in the miR-23b inhibitor group was significantly reduced (P < 0.01); compared with the miR-23b inhibitor group, liquiritin treatment (miR-23b inhibitor + Liq) had no obvious effect on the level of miR-23b. It is shown that liquiritin may play a role by inhibiting the expression level of miR-23b.

[0076] 3. Liquiritin inhibits the expression of miR-23b in SW480 cells and increases the level of PTPRG

[0077] To investigate whether liquiritin can reverse the miR-23b-mediated down-regulation effect of PTPRG, the expression levels of PTPRG gene and protein were detected under the conditions of miR-23b overexpression (mimic) and inhibition (inhibitor), respectively.

[0078] The results are as Figure 10As shown, compared with the control group (miR-NC), overexpression of miR-23b (miR-23b mimic) significantly decreased the gene and protein levels of PTPRG (P<0.01), while liquiritin treatment (miR-23b mimic+Liq) partially restored the gene and protein expression of PTPRG (P<0.05), indicating that liquiritin may restore PTPRG expression by inhibiting the action of miR-23b. In addition, compared with the control group (miR-NC), inhibition of miR-23b (miR-23b inhibitor) significantly upregulated the gene and protein expression of PTPRG (P<0.05), while liquiritin treatment (miR-23b inhibitor+Liq) further enhanced the gene and protein expression levels of PTPRG (P<0.05), indicating that liquiritin may cooperate with the inhibitory effect of miR-23b to increase the gene and protein levels of PTPRG, suggesting that liquiritin can reverse the miR-23b-mediated inhibitory effect of PTPRG by restoring PTPRG expression. These results indicate that miR-23b promotes the progression of colorectal cancer by inhibiting PTPRG, while liquiritin can play its anti-cancer role by downregulating the level of miR-23b and increasing PTPRG expression.

[0079] Example 4:

[0080] Liquiritin inhibits the growth of colorectal cancer xenografts in nude mice through the miR-23b-PTPRG pathway

[0081] 1. Liquiritin reverses the pro-cancer effect of miR-23b in vivo

[0082] (1) In vivo imaging analysis of miR-23b in tumors of nude mice in each group

[0083] To evaluate the effect of liquiritin on the growth of miR-23b-mediated colorectal cancer tumors in vivo, in vivo bioluminescence imaging (IVIS) of small animals was used to monitor the expression of miR-23b in tumor tissues of nude mice in the miR-23b-NC SW480 cell line stable transfection group (Model group), the miR-23b overexpression SW480 cell line stable transfection group (miR-23b group), and the miR-23b overexpression SW480 cell line + liquiritin (40 mg / kg) treatment group (miR-23b+Liq group).

[0084] The results are as Figure 11As shown, the luminescence signal intensity represents the expression level of miR-23b in tumor tissues. The color ranges from blue (low signal) to red (high signal), corresponding to the expression level of miR-23b in tumor tissues. In the Model group, the luminescence signal of the tumor tissues in nude mice was weak and the range was small, indicating a low expression level of miR-23b; in the miR-23b group, the luminescence signal of the tumor tissues in nude mice was significantly enhanced and the range was wide, and the signal was red in some areas, indicating overexpression of miR-23b; in the miR-23b+Liq group, the luminescence signal of the tumor tissues in nude mice was significantly weaker than that in the miR-23b group and the range was small, approaching the level of the Model group. In vivo imaging experiments showed that liquiritin could effectively inhibit the expression of miR-23b in the tumor tissues of nude mice. It is suggested that liquiritin can inhibit the growth of colorectal cancer by regulating the miR-23b-related pathway, further supporting its potential as an anti-cancer drug.

[0085] (2) Comparison of tumor sizes in nude mice of each group

[0086] To further explore whether liquiritin (40 mg / kg) can inhibit the growth of miR-23b-mediated colorectal cancer tumors, a subcutaneous tumorigenesis experiment in nude mice was used to evaluate the effects of the stable transfection miR-23b-NC SW480 cell line group (Model group), the stable transfection miR-23b overexpression SW480 cell line group (miR-23b group), and the stable transfection miR-23b overexpression SW480 cell line + liquiritin treatment group (miR-23b+Liq group) on tumor growth.

[0087] The results are as Figure 12 shown. Compared with the Model group, larger tumors were visible on the body surface of nude mice in the miR-23b group, indicating that miR-23b significantly promoted the growth of tumors in nude mice; while in the miR-23b+Liq group, the tumor volume on the body surface of nude mice was significantly reduced, indicating that liquiritin could significantly inhibit the pro-cancer effect mediated by miR-23b. After the tumor tissues on the body surface of nude mice in each group were dissected, compared with the Model group, the tumor tissue volume of nude mice in the miR-23b group was significantly increased, while after treatment with liquiritin (40 mg / kg), the tumor tissue volume was significantly reduced. The above results prove that liquiritin can inhibit the growth of colorectal cancer by inhibiting the expression of miR-23b.

[0088] (3) Changes in body weight and tumor mass of nude mice in each group

[0089] To explore the effect of liquiritin (40 mg / kg) on the tumor growth promotion effect mediated by miR-23b and evaluate its effects on the body weight and tumor weight of experimental animals, the changes in body weight and tumor weight of mice were recorded respectively.

[0090] The results are as Figure 13As shown, the body weights of the mice in the Model group and the miR-23b + Liq group increased steadily over time, and the changing trends were similar, suggesting that liquiritin had little effect on the overall health status of the mice and did not affect the normal growth of the animals. Low expression of miR-23b also had no significant effect on the overall health status of the mice. The body weight gain of the mice in the miR-23b group was slightly higher than that of the other two groups, indicating that overexpression of miR-23b promoted tumor growth and body weight gain, but there was no significant difference. In addition, the tumor weight in the miR-23b group was significantly higher than that in the Model group, indicating that overexpression of miR-23b significantly promoted tumor growth; the miR-23b + Liq group significantly reduced the tumor weight, approaching the level of the Model group, indicating that liquiritin could effectively inhibit the pro-cancer effect mediated by miR-23b.

[0091] 2. Liquiritin can reverse the low expression of PTPRG caused by miR-23b in vivo

[0092] To further explore in vivo whether liquiritin can reverse the down-regulation effect of miR-23b-mediated PTPRG, Western blot was used to evaluate the protein expression of PTPRG in different tissue samples.

[0093] The results are as Figure 14 shown. Compared with the Model group, the expression level of PTPRG in the miR-23b group was significantly decreased (P < 0.01). The miR-23b + Liq group could significantly restore the expression level of PTPRG (P < 0.01), but it was still lower than that in the Model group. The results showed that overexpression of miR-23b in vivo could significantly down-regulate the expression of PTPRG, while liquiritin could effectively restore its level, suggesting that liquiritin up-regulated the expression of PTPRG by inhibiting the level of miR-23b.

[0094] The above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those of ordinary skill in the art should understand that they can still modify the technical solutions recorded in the foregoing embodiments, or perform equivalent replacements for some of the technical features; and these modifications or replacements do not make the essence of the corresponding technical solutions deviate from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. Use of a reagent for detecting the expression level of PTPRG in the preparation of a product for diagnosing colorectal cancer.

2. The use according to claim 1, characterized in that The reagent for detecting the expression level of PTPRG includes a reagent for detecting the expression level of PTPRG gene mRNA and / or a reagent for detecting the expression level of PTPRG protein.

3. The use according to claim 1, characterized in that The reagent for detecting the expression level of the PTPRG protein includes an antibody against the PTPRG protein.

4. The use according to claim 1, characterized in that The product includes a test kit, a chip or a test paper.

5. Application of PTPRG as a regulatory target in the preparation of drugs for the treatment of colorectal cancer.

6. Use of reagents that promote PTPRG expression in the preparation of drugs for the treatment of colorectal cancer.

7. Application of glycyrrhizin in the preparation of drugs for preventing and / or treating colorectal cancer by inhibiting miR-23b and targeting and promoting the expression of PTPGR.

8. The use according to claim 7, characterized in that: The drug also contains a pharmaceutically acceptable carrier.

9. The use according to claim 8, characterized in that: The pharmaceutically acceptable carrier is selected from one or more of a diluent, an excipient, a filler, a binder, a wetting agent, a disintegrant, an absorption promoter, a surfactant, an adsorption carrier and a lubricant.

10. The use according to claim 7, characterized in that: The drug is prepared into a pharmaceutically acceptable dosage form.

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