Use of a GLYR1 protein for non-therapeutic purposes in regulating the proliferation and / or migration of renal clear cell carcinoma cells

Abstract

The present invention discloses the application of a GLYR1 protein for non-therapeutic purposes in regulating the proliferation and / or migration of renal clear cell carcinoma cells. The feature is to knock down the expression of GLYR1, thereby promoting the proliferation and / or migration of renal clear cell carcinoma cells. The reagent used to knock down the expression of GLYR1 is si-GLYR1#1. The sense strand of si-GLYR1#1 is CGGUAGAUGCUGUCGAAGA, and the antisense strand of si-GLYR1#1 is UCUUCGACAGCAUCUACCG. The advantages are high sensitivity, strong specificity, short cycle, etc., which are beneficial to the early diagnosis and treatment of renal clear cell carcinoma, and have the potential to become a novel biomarker and an effective therapeutic target in renal clear cell carcinoma.

Description

Technical Field

[0001] The present invention belongs to the field of molecular biology, and particularly relates to the application of a GLYR1 protein for non-therapeutic purposes in regulating the proliferation and / or migration of clear cell renal carcinoma cells. Background Art

[0002] Renal cell carcinoma is a malignant tumor originating from renal tubular epithelial cells. According to histological classification, about 85% of renal cell carcinomas are clear cell renal cell carcinomas (ccRCC for short). ccRCC mainly originates from renal tubular epithelium and occurs in the renal parenchyma. Its incidence rate is increasing year by year, and it is the main cause of renal cancer-related deaths. The 5-year survival rate of early ccRCC patients after surgery is over 90%, that of mid-stage patients is about 80%, but for late-stage patients, the survival rate is only about 20%. Secondly, ccRCC is insensitive to chemotherapy. At present, surgical resection is mostly performed clinically, but up to 1 / 3 of the patients will have metastases after surgery. In addition, surgery with or without postoperative adjuvant therapy, chemotherapy, immunotherapy, targeted therapy, etc. still have a poor overall prognosis for ccRCC patients, especially for late-stage ccRCC patients. Therefore, it is urgent to deeply explore the occurrence and development mechanism of ccRCC, and finding accurate and effective diagnostic markers is very important for improving its treatment effect.

[0003] ccRCC is a highly vascularized tumor, and one of its characteristics is that the inactivation rate of the Von Hippel-Lindau (abbreviated as VHL ) gene is as high as 50-75%. VHL The product pVHL of the gene plays an important role in downregulating the expression of the hypoxia-inducible factor 1 (abbreviated as HIF1) transcription factor, which can lead to a reduction in angiogenesis. VHL The inactivation of the gene provides a theoretical basis for the development of anti-angiogenic drugs for the treatment of ccRCC (such as sunitinib, pazopanib, sorafenib, and axitinib). At present, local ccRCC can be treated by partial nephrectomy or radical nephrectomy. However, about 30% of ccRCC patients are unable to receive surgical treatment due to tumor metastasis, and the treatment effect of chemotherapy on metastatic ccRCC is poor. Therefore, the use of systemic treatments such as targeted therapy and immunotherapy is very important. Targeted therapy drugs include tyrosine kinase inhibitors, which can inhibit the kinase activities of the vascular endothelial growth factor receptor family (such as VEGFR-1 and VEGFR-2) and platelet-derived growth factor receptors (such as PDGFRα and PDGFRβ), as well as c-Kit. At present, sunitinib is still the first-line treatment drug for RCC, and in the phase III clinical trial of RCC, sunitinib is also used as a comparator drug. However, the phenomenon of sunitinib resistance has become a clinical challenge, and there is an urgent need to find new treatment strategies.

[0004] The glyoxylate reductase 1 homolog (GLYR1) protein is an epigenetic reader involved in chromatin modification and gene expression regulation, regulating gene expression by demethylating nucleosomes; the role of GLYR1 in cells includes promoting the transcription of RNA polymerase II (Pol II) in chromatin. It interacts with the region near the centromere of the nucleosome and dissociates the nucleosome in an ATP-independent manner, thereby reducing the stability of the nucleosome and promoting transcription. GLYR1 is highly expressed in almost all tissues and is essential in stem cells. In different diseases, the expression of GLYR1 is different: it is highly expressed in breast cancer, and this high expression is associated with poor prognosis; its expression is downregulated in colorectal cancer, and this downregulation reduces the sensitivity of colorectal cancer cells to 5-fluorouracil; other studies have shown that GLYR1 deficiency weakens spermatogenesis in mice. At present, there is no publicly reported research on using the GLYR1 protein as a diagnosis and treatment marker in clear cell renal cell carcinoma. Summary of the Invention

[0005] The technical problem to be solved by the present invention is to provide an application of a non-therapeutic GLYR1 protein with high sensitivity and specificity and negatively correlated with the prevalence of clear cell renal cell carcinoma in regulating the proliferation and / or migration of clear cell renal cell carcinoma cells.

[0006] The technical solution adopted by the present invention to solve the above technical problem is: an application of a non-therapeutic GLYR1 protein in regulating the proliferation and / or migration of clear cell renal cell carcinoma cells.

[0007] Furthermore, knocking down the expression of GLYR1 promotes the proliferation and / or migration of clear cell renal cell carcinoma cells. The reagent used for knocking down the expression of GLYR1 is si-GLYR1#1. The nucleotide sequence of the sense strand of si-GLYR1#1 is as shown in SQINO: 1: CGGUAGAUGCUGUCGAAGA, and the nucleotide sequence of the antisense strand of si-GLYR1#1 is as shown in SQI NO: 2: UCUUCGACAGCAUCUACCG.

[0008] The protein GLYR1 consists of 553 amino acids and the protein size is approximately 60 kDa.

[0009] Compared with the prior art, the advantages of the present invention are as follows: The present invention firstly discloses the protein GLYR1 that can be used to detect renal clear cell carcinoma and its application in the preparation of reagents for detecting renal clear cell carcinoma or reagents for assisting in the diagnosis of renal clear cell carcinoma. The detection is mainly completed through immunohistochemistry technology. By analyzing immunohistochemical staining pictures and calculating and comparing the expression of GLYR1 protein in samples of patients with renal clear cell carcinoma and normal samples through Image J software, it is used to assist in the diagnosis of renal clear cell carcinoma. It has the characteristics of high sensitivity, strong specificity, and short cycle compared with traditional renal clear cell carcinoma detection technologies, which is beneficial to the early diagnosis and treatment of renal clear cell carcinoma. And GLYR1 has the potential to become a new biomarker and an effective therapeutic target in renal clear cell carcinoma. BRIEF DESCRIPTION OF THE DRAWINGS

[0010] Figure 1 To verify the expression of GLYR1 in human renal clear cell carcinoma tissues and paired adjacent cancer tissues by immunohistochemistry, where A are two representative images of GLYR1 immunohistochemical staining in 60 pairs of renal clear cell carcinoma tissues and paired adjacent cancer tissues (n = 60); B is a stacked bar chart of the immunohistochemical staining results of 60 pairs of human renal clear cell carcinoma tissues and paired adjacent cancer tissues (n = 60, ****: p <0.0001). In the figure, negative indicates no expression or low expression of GLYR1, and positive indicates high expression of GLYR1;

[0011] Figure 2 To screen siRNA for knocking out GLYR1 by protein immunoblotting technology, where A is screening ACHN cells with knocked-out GLYR1 by protein immunoblotting technology, and B is screening 786O cells with knocked-out GLYR1 by protein immunoblotting technology; in the figure, si-NC is si-Negative Control, which is the negative control of si; GAPDH is glyceraldehyde-3-phosphate dehydrogenase, which is an internal reference gene; si-GLYR1#1-4 indicates no expression or low expression, α-GLYR1 is the GLYR1 antibody, and α-GAPDH is the GAPDH antibody;

[0012] Figure 3 To verify the effect of GLYR1 expression level on the growth of renal clear cell carcinoma cells by CCK8 experiment, where A is the CCK8 experiment to detect the effect of GLYR1 expression level on the growth of ACHN cells; B is the CCK8 experiment to detect the effect of GLYR1 expression level on the growth of 786O cells (*: p <0.05, **: p <0.01, ***: p <0.001, ****: p<0.0001); In the figure, EV represents the transfection of an empty vector plasmid without the target gene, serving as a negative control; oe-GLYR1 represents the transfection of the SFB-GLYR1 plasmid for overexpression of the GLYR1 gene; si-NC represents the transfection of si-Negative control, serving as a negative control; si-GLYR1 represents the transfection of si-GLYR1#1 for knockdown of the GLYR1 gene.

[0013] Figure 4 For the Transwell migration experiment to verify the effect of GLYR1 expression level on the migration of renal clear cell carcinoma cells, where A is the migration experiment of ACHN and 786O cells after overexpression or knockout of GLYR1; B is the quantification of the cell migration experiment results (*: p <0.05, **: p <0.01, ***: p <0.001, ****: p <0.0001); EV represents the transfection of an empty vector plasmid without the target gene, serving as a negative control; oe-GLYR1 represents the transfection of the SFB-GLYR1 plasmid for overexpression of the GLYR1 gene; si-NC represents the transfection of si-Negative control, serving as a negative control; si-GLYR1 represents the transfection of si-GLYR1#1 for knockdown of the GLYR1 gene.

[0014] Figure 5 For the colony formation assay to verify the effect of GLYR1 expression level on the growth of renal clear cell carcinoma cells, where A is the colony formation assay to detect the colony formation ability of ACHN and 786O after overexpression or knockout of GLYR1; B is the quantification of the colony formation assay results (*: p <0.05, **: p <0.01, ***: p <0.001, ****: p <0.0001); EV represents the transfection of an empty vector plasmid without the target gene, serving as a negative control; oe-GLYR1 represents the transfection of the SFB-GLYR1 plasmid for overexpression of the GLYR1 gene; si-NC represents the transfection of si-Negative control, serving as a negative control; si-GLYR1 represents the transfection of si-GLYR1#1 for knockdown of the GLYR1 gene. Detailed implementation methods

[0015] The following further describes the present invention in detail with reference to the accompanying drawings and embodiments.

[0016] I. Experimental methods.

[0017] 1. Collection of renal clear cell carcinoma tissue specimens: The 60 pairs of white slices of renal clear cell carcinoma tissues included in the study were sourced from Shanghai Core Biotech Co., Ltd. All human tissue specimens involved in this experiment were only used for laboratory research under the condition of being approved by the Human Ethics Committee of Ningbo University School of Medicine upon the application of the research group.

[0018] 2. Immunohistochemical staining of specimens was performed as follows:

[0019] (1) Baking the slides: Place the tissue microarray on a preheated oven at 63 °C for baking the wax for 1 h, or place it in an oven at 37 °C overnight.

[0020] (2) Deparaffinization and rehydration: Take out the tissue microarray from the oven and place it in an automatic staining machine for deparaffinization. The specific deparaffinization process is as follows: Immerse the tissue microarray in xylene twice for 15 min each; immerse it in absolute ethanol twice for 7 min each; immerse it in 90% ethanol once for 5 min; immerse it in 80% ethanol once for 5 min; immerse it in 70% ethanol once for 5 min. Finally, place the slide rack in deionized water and wash for 5 min.

[0021] (3) Antigen retrieval: Use a DAKO automatic immunohistochemistry pretreatment system instrument for retrieval.

[0022] (4) Primary antibody incubation: After retrieval, take out the slides and rinse them 3 times with PBST buffer for 1 min each. Take out the anti-GLYR1 rabbit primary antibody (brand: proteintech, model: 14833-1-AP) from the refrigerator, centrifuge it at 7200 rpm for 1 min in a centrifuge. Take out the anti-GLYR1 rabbit primary antibody and dilute it with antibody diluent at a dilution ratio of 1:500. Drop the anti-GLYR1 rabbit primary antibody and incubate it overnight in a 4 °C refrigerator.

[0023] (5) Secondary antibody incubation: Let the slides warm up at room temperature for more than 30 min, then rinse them 3 times with PBST buffer for 1 min each. Dry the liquid around the tissue, drop an appropriate concentration of HRP-labeled rabbit secondary antibody on the tissue to completely cover the tissue, close the lid of the wet box, and incubate at 37 °C for 1 h.

[0024] (6) DAB color development: Rinse the slides 3 times with PBST buffer for 1 min each. Drop the DAB color development solution on the position of the tissue on the glass slide, place the glass slide on an inverted microscope to observe the tissue staining situation. When the staining intensity reaches the best, discard the color development solution and rinse with deionized water for 5 min.

[0025] (7) Hematoxylin counterstaining: Drop Harris hematoxylin (abbreviated as SIGMA) on the slides for 1 min, immerse it in 0.25% hydrochloric acid alcohol for no less than 2 s, and rinse with tap water for more than 2 min.

[0026] (8)Cover slip mounting: Mount the slides after air-drying at room temperature;

[0027] (9)Scoring: All samples were reviewed by two independent pathologists with extensive experience in IHC assessment who were blinded to the clinical outcomes of these patients. We evaluated the percentage of positively stained cells and the staining intensity to semi-quantitatively determine GLYR1 expression. The percentage score of positively stained cells was as follows: 0, none; 1, <=50%; 2, >=50%. The staining intensity was graded as follows: 0 (no or weak staining = light yellow), 1 (moderate staining = tan), and 2 (strong staining = brown). The total score of GLYR1 expression was the sum of the percentage score of positively stained cells and the staining intensity score, with a total score ranging from 0 to 4. For statistical analysis, the final score was a combination of the independent scores assigned by the two pathologists reported in this study. Any differences in scores were resolved through discussion between the two pathologists.

[0028] 3. Screening of si-GLYR1 by Western blotting, the steps are as follows:

[0029] (1)si-GLYR1 is a small interfering RNA of GLYR1, responsible for silencing the expression of the GLYR1 gene. Design the small interfering RNA primers as follows: The nucleotide sequence of the sense strand of si-GLYR1#1 is shown in SQI NO: 1: 5'-CGGUAGAUGCUGUCGAAGA-3', and the nucleotide sequence of the antisense strand of si-GLYR1#1 is shown in SQI NO: 2: 5'-UCUUCGACAGCAUCUACCG-3';

[0030] The nucleotide sequence of the sense strand of si-GLYR1#2 is shown in SQI NO: 3: 5'-GACCAGUCUGACAACGAUA-3', and the nucleotide sequence of the antisense strand of si-GLYR1#2 is shown in SQI NO: 4: 5'-UAUCGUUGUCAGACUGGUC-3';

[0031] The nucleotide sequence of the sense strand of si-GLYR1#3 is shown in SQI NO: 5: 5'-CGGAGUCUAGUACCGUGAA -3', and the nucleotide sequence of the antisense strand of si-GLYR1#3 is shown in SQI NO: 6: 5'-UUCACGGUACUAGACUCCG-3';

[0032] The nucleotide sequence of the sense strand of si-GLYR1#4 is shown in SQI NO: 7: 5'-GCAAUCACGAAGAAGUUGA -3', and the nucleotide sequence of the antisense strand of si-GLYR1#4 is shown in SQI NO: 8: 5'-UCAACUUCUUCGUGAUUGC-3';

[0033] (2)Cells in 12-well plates were transiently transfected with Negative Control (control group), si-GLYR1#1-4 respectively. After 48 h, the culture medium of adherent cells of ACHN and 7860 renal clear cell carcinoma with good growth was aspirated, and 100 μL of RIPA lysis buffer (weak) was added to each well. After shaking on a shaker at 4°C for 20 minutes, it was transferred to -80°C and frozen overnight.

[0034] (3)The samples were taken out from -80°C, melted at 4°C and transferred to 1.5 mL centrifuge tubes. After centrifugation at 4°C and 12000 rpm for 20 min, the supernatant was taken and sodium dodecyl sulfate (SDS) was added, and then boiled in a metal bath at 95°C for 5 min.

[0035] (4)Prepare 10% WB gel in advance for standby. Run electrophoresis at a constant voltage of 80V, transfer membrane at a constant current of 220 mA for 120 min, wash the membrane with TBST for 5 min, block with 5% milk powder prepared with TBST for 40 min and then wash the membrane, and incubate with corresponding antibodies overnight.

[0036] (5)Prepare developing solution, water and fixing solution in the darkroom in advance. After drying the membrane dropped with ECL luminescent solution, it was laid flat on the plastic wrap, then fixed in the dark box, and exposed by pressing the film in the darkroom. After the exposure was completed, the dark box was opened, the film was taken out, immersed in the developing solution, observed under the red light until the bands appeared, washed in water and then put into the fixing solution, and then observed the exposure result with the light on. Finally, the film was scanned to analyze the result.

[0037] 4. CCK8 cell proliferation assay

[0038] (1)ACHN and 786O renal clear cell carcinoma adherent cells with good growth were digested into a uniform cell suspension after transient transfection with PCIN4 plasmid, SFB-GLYR1 plasmid, Negative Control (control group), and si-GLYR1 for 24 h respectively.

[0039] (2)20 μL of the above uniform cell suspension was aspirated and added to a cell counting plate, and its concentration was measured using a cell counter. The cell suspension was diluted according to the corresponding ratio and pipetted evenly.

[0040] (3)Take 3 96-well cell plates, add 1.5×10 3 cells to each well, with a volume of 0.1 mL. Set 6 replicates for each treatment group, shake well using the "8" method, and place in a constant temperature incubator.

[0041] (4)When three 96-well cell culture plates are cultured for 0, 1, 2, 3, 4, and 5 days respectively, add 10 µL of CCK8 reagent to each well under light-proof conditions, place it in a constant temperature incubator for 2 h, and observe that the well plate turns orange-yellow. Set the wavelength of the microplate reader to 450 nm, measure the absorbance of each well, repeat the measurement 3 times, record and organize the corresponding data.

[0042] 5. Transwell migration experiment, the steps are as follows:

[0043] (1)After transiently transfecting the PCIN4 plasmid, SFB-GLYR1 plasmid, Negative Control (control group), and si-GLYR1 for 24 h respectively, digest the adherent cells of well-grown ACHN and 786O renal clear cell carcinoma into a uniform cell suspension.

[0044] (2)Collect the cell suspension into a 1.5 mL centrifuge tube and centrifuge at 1,000 rpm for 4 min.

[0045] (3)Discard the supernatant, resuspend the cell pellet with 1 mL of PBS solution, centrifuge again according to the above method, resuspend the cells with 1 mL of DMEM medium, take a cell counting plate to count them, and dilute them as needed with DMEM.

[0046] (4)Use sterile forceps to pick up 4 Transwell chambers, put them into a new 24-well cell plate, add 200 µL of the above cell suspension to the upper chamber, quantify 2×10 4 cells, add 700 µL of complete medium to the lower chamber, let it stand for 30 min, and then place the well plate in a constant temperature incubator.

[0047] (5)After 24 h, discard the culture medium inside and outside the chamber, rinse the chamber and the bottom of the well plate with PBS buffer twice.

[0048] (6)Add 500 µL of pre-cooled ice methanol to the chamber and place it on ice for 5 min.

[0049] (7)Discard the fixing solution, rinse with PBS solution twice, add 500 µL of 0.1% crystal violet dye, and place the well plate on a shaker for staining for 30 min.

[0050] (8)Recover the staining agent, rinse with PBS solution three times, air-dry the chamber naturally, take pictures and count the data.

[0051] 6. Colony formation, the steps are as follows:

[0052] (1)After transiently transfecting PCIN4 plasmid, SFB-GLYR1 plasmid, Negative Control (control group), and si-GLYR1 respectively for 24 h, the adherent cells of ACHN and 786O renal clear cell carcinoma with good growth were digested into a uniform cell suspension.

[0053] (2)Aspirate 20 µL of the above uniform cell suspension and add it to a cell counting chamber. Measure its concentration using a cell counter, dilute the cell suspension according to the corresponding ratio, and pipette it evenly.

[0054] (3)Take several 6-well cell culture plates, add 1×10 3 cells to each well, quantify to 2 mL, set 4 replicates for each treatment group, shake well using the "8" method, place in a constant temperature incubator, and change the medium every 5 days for a total of 10 - 15 days.

[0055] (4)When cell colonies are visible to the naked eye in the 6-well plate, discard the medium in the wells, rinse twice with PBS buffer, add an appropriate amount of ice-cold methanol to each well to fix the cells, and let stand for 30 min.

[0056] (5)Discard the fixing solution, rinse twice with PBS buffer, add 1 ml of 0.1% crystal violet dye to each well, place on a shaker for 30 min, recover the dye, rinse three times with PBS buffer, place in a 37℃ oven overnight, and take pictures and count data after the 6-well plate is dried.

[0057] 7. The result analysis method is as follows:

[0058] In this experiment, the experimental data and the collected original clinical data were comprehensively examined, proofread, and sorted to ensure that the data was as complete, accurate, and error-free as possible. Then, tissue and cell databases were established using Excel software respectively, and the original clinical data involved was grouped, summarized, and entered into a table. Finally, the final experimental data was sorted and analyzed using SPSS 26.0 statistical software. All data statistical tests were two-sided probabilities, and statistical tests were performed according to the significance level of α = 0.05. If p <0.05, it was considered that the difference between the two groups was statistically significant; otherwise, the difference was considered not statistically significant. The experimental result graphs such as bar graphs and line graphs were jointly completed using GraphPad Prism 10.0 and SPSS 26.0 software.

[0059] II. Experimental Results.

[0060] To examine the expression of GLYR1 in renal clear cell carcinoma tissues, we collected tissue microarrays of 60 pairs of renal clear cell carcinoma tissues and adjacent para-carcinoma tissues, and detected the expression level of GLYR1 in clinical samples using immunohistochemical staining.

[0061] As shown in Figure 1 Figure A, immunohistochemical staining results showed that GLYR1 staining presented as brownish yellow with different shades and was mainly localized in the cytoplasm of adjacent tissues to cancer and a small amount of renal clear cell carcinoma tissues. Compared with renal clear cell carcinoma tissues, GLYR1 was more strongly expressed in the corresponding adjacent tissues to cancer. Using Image J software to analyze the immunohistochemical staining pictures, the results showed that the expression levels of GLYR1 in renal clear cell carcinoma tissues and adjacent tissues to cancer were concentrated in the two grades of Negative (0 - 1 point) and (2 - 4 points). At the same time, as shown in Figure 1 Figure B, in renal clear cell carcinoma tissues, the positive expression rate of GLYR1 was 25% (15 / 60), while that in adjacent tissues to cancer was as high as 92% (55 / 60), and the difference was statistically significant ( p <0.0001). The expression level of GLYR1 in renal clear cell carcinoma tissues was significantly lower than that in the corresponding adjacent tissues to cancer ( p <0.0001).

[0062] For the subsequent cell function experiments to further verify the function of GLYR1 protein, we first screened si - GLYR1, used si - GLYR1 to knockdown GLYR1 in renal clear cell carcinoma cell lines ACHN and 786O, selected effective si - GLYR1, and further confirmed it using Western blotting. The results are as shown in Figure 2 Figure A and Figure 2 Figure B. The final results showed that si - GLYR1#1 was an effective knockdown siRNA, and si - GLYR1#1 was used as the si - GLYR1 described in the subsequent experiments.

[0063] In the experiments to further verify the effects of GLYR1 on the proliferation and migration abilities of renal clear cell carcinoma cells, we adopted CCK8 assay, Transwell migration assay, and colony formation assay. As shown in Figure 3 Figure A and Figure 3 Figure B, the CCK8 assay showed that compared with renal clear cell carcinoma cells in the NC group, the proliferation rate of cells in the GLYR1 high - expression group was significantly slower, while the si - GLYR1 group showed accelerated proliferation compared with the NC group. It indicated that GLYR1 had the ability to inhibit the proliferation of renal clear cell carcinoma cells.

[0064] As shown in Figure 4 Figure A and Figure 4As shown in Figure B, the Transwell migration assay showed that in renal clear cell carcinoma cells, the number of cells penetrating the Transwell chamber in the GLYR1 high-expression group was significantly reduced compared to the NC group, while the si-GLYR1 group showed a significantly increased number of cells penetrating the Transwell chamber compared to the NC group. This indicates that GLYR1 has the ability to inhibit the migration of renal clear cell carcinoma cells.

[0065] As Figure 5 shown in Figure A and Figure 5 Figure B, the colony formation assay showed that compared with renal clear cell carcinoma in the NC group, the number of cell colonies in the GLYR1 high-expression group was significantly reduced, while the si-GLYR1 group showed a significantly increased number of cell colonies compared to the NC group. This indicates that GLYR1 has the ability to inhibit the proliferation and colony formation of renal clear cell carcinoma cells.

[0066] These results suggest that the detection of GLYR1 protein has high value for diagnosis, especially immunohistochemistry. The expression level of GLYR1 in renal clear cell carcinoma tissues was significantly lower than that in the corresponding adjacent tissues (p < 0.0001). Therefore, detecting the expression of GLYR1 protein in renal clear cell carcinoma tissues can achieve a diagnosis of renal clear cell carcinoma with high sensitivity and specificity.

[0067] The present invention provides a detection method with high sensitivity, strong specificity, short cycle, and stable results by detecting the expression of GLYR1 in renal clear cell carcinoma tissues and the effects of GLYR1 high-expression and low-expression on the proliferation and migration of renal clear cell carcinoma cells, combined with statistical principles and modern biological techniques, providing a scientific basis for the diagnosis and treatment of patients with renal clear cell carcinoma and the possibility of molecular targeted therapy.

[0068] The above description is not a limitation of the present invention, and the present invention is not limited to the above examples. Changes, modifications, additions, or substitutions made by those of ordinary skill in the art within the scope of the essence of the present invention should also fall within the protection scope of the present invention.

Claims

1. Use of a reagent for specifically detecting the expression of GLYR1 protein in the preparation of a reagent for assisting in the diagnosis of clear cell renal carcinoma, characterized in that, The GLYR1 protein described above is a glyoxylate reductase 1 homolog protein, and the reagent for specifically detecting the expression of the GLYR1 protein is an antibody against the GLYR1 protein.

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