Use of reagent for detecting expression of hn1l in preparation of esophageal squamous cell carcinoma diagnosis kit

Abstract

The application discloses application of a reagent for detecting HN1L expression in preparation of an esophageal squamous carcinoma diagnosis kit. Through in-vitro and in-vivo function experiments, it is found that knocking down HN1L can inhibit esophageal squamous carcinoma cell metastasis and proliferation, and improve the sensitivity of tumor cells to chemotherapeutic drugs, so that HN1L can be used as a specific marker gene for diagnosing esophageal squamous carcinoma, and the diagnosis of esophageal squamous carcinoma is more accurate and rapid. HN1L can also be used as a drug target for preparing a drug for treating esophageal squamous carcinoma, and a new effective treatment scheme, i.e. a treatment scheme of targeting HN1L combined with a chemotherapeutic drug for treating esophageal squamous carcinoma, is provided, so that a new way for treating esophageal squamous carcinoma is provided.

Description

Technical Field

[0001] This invention relates to the field of biomedical technology, specifically to the application of a reagent for detecting HN1L expression in the preparation of a diagnostic kit for esophageal squamous cell carcinoma. Background Technology

[0002] Esophageal cancer is a common malignant tumor of the digestive system. The survival rate of patients with esophageal squamous cell carcinoma is low, mainly due to late onset of clinical symptoms and the lack of early diagnostic biomarkers. Therefore, the diagnosis and treatment of esophageal squamous cell carcinoma urgently require new, highly sensitive, and cost-effective biomarkers and therapeutic targets.

[0003] The HN1L gene, also known as JPT2, C16orf34, or L11, encodes a 190-amino acid protein (NCBI Reference Sequence: NP_653171.1). This protein is primarily located in the cell nucleus, cytoplasm, and cell membrane, and is specifically expressed in organs and tissues such as the liver, kidney, prostate, testes, and uterus. The HN1L gene was first identified in a mouse zygote cDNA library in 2000. Currently, existing technologies have revealed that HN1L plays a pro-cancer role in tumorigenesis and development: HN1L promotes the invasion and metastasis of adenocarcinoma at the esophagogastric junction (PMID:33471419), HN1L promotes the migration and invasion of breast cancer by upregulating HMGB1 expression (PMID:33191617), HN1L-mediated transcriptional axis AP-2γ / METTL13 / TCF3-ZEB1 drives tumor growth and metastasis of hepatocellular carcinoma (PMID:30778199), HN1L promotes triple-negative breast cancer stem cells through the LEPR-STAT3 pathway (PMID:29249663), and HN1L overexpression promotes the malignant proliferation of non-small cell lung cancer cells (PMID:29053395), etc. Summary of the Invention

[0004] The purpose of this invention is to overcome the aforementioned shortcomings of the prior art and provide an application of a reagent for detecting HN1L expression in the preparation of a diagnostic kit for esophageal squamous cell carcinoma. This invention, through in vitro and in vivo functional experiments, has discovered that knocking down HN1L can inhibit the metastasis and proliferation of esophageal squamous cell carcinoma cells and increase the sensitivity of tumor cells to docetaxel or cisplatin. Therefore, the HN1L gene and its expression products can serve as diagnostic biomarkers and drug targets for esophageal squamous cell carcinoma, and targeting HN1L in combination with chemotherapy drugs is a new and effective treatment option for esophageal squamous cell carcinoma.

[0005] The first objective of this invention is to provide the application of reagents for detecting HN1L gene expression, HN1L mRNA expression, and / or HN1L protein expression in the preparation of diagnostic kits for esophageal squamous cell carcinoma.

[0006] A second objective of this invention is to provide the application of reagents for detecting HN1L gene expression, HN1L mRNA expression, and / or HN1L protein expression in the preparation of a prognostic kit for esophageal squamous cell carcinoma.

[0007] A third objective of this invention is to provide the application of reagents for detecting HN1L gene expression, HN1L mRNA expression, and / or HN1L protein expression in the preparation of a chemosensitivity assay kit for patients with esophageal squamous cell carcinoma.

[0008] A fourth objective of this invention is to provide the use of inhibitors of HN1L gene expression, HN1L mRNA expression, and / or HN1L protein expression in the preparation of drugs for esophageal squamous cell carcinoma.

[0009] A fifth objective of this invention is to provide the use of inhibitors of HN1L gene expression, HN1L mRNA expression, and / or HN1L protein expression in the preparation of potentiators for chemotherapy drugs for esophageal squamous cell carcinoma.

[0010] The sixth objective of this invention is to provide a drug for esophageal squamous cell carcinoma.

[0011] To achieve the above objectives, the present invention is implemented through the following solution:

[0012] The application of reagents for detecting HN1L gene expression, HN1L mRNA expression and / or HN1L protein expression in the preparation of diagnostic kits for esophageal squamous cell carcinoma. The expression levels of HN1L gene and / or HN1L protein in patients with esophageal squamous cell carcinoma are higher than those in patients without esophageal squamous cell carcinoma.

[0013] Preferably, the esophageal squamous cell carcinoma diagnostic kit is a kit for diagnosing tumor tissue invasion. Patients with high expression of HN1L gene, HN1L mRNA and / or HN1L protein, and patients with relatively low expression, have severe tumor tissue invasion.

[0014] Preferably, the esophageal squamous cell carcinoma diagnostic kit is a kit for diagnosing lymph node metastasis. Patients with high expression of HN1L gene, HN1L mRNA, or / and HN1L protein, and patients with relatively low expression, have severe lymph node metastasis.

[0015] Preferably, the esophageal squamous cell carcinoma diagnostic kit is a kit for diagnosing staging. Patients with high expression of HN1L gene, HN1L mRNA, or / and HN1L protein, and patients with relatively low expression, are in a later stage.

[0016] This invention also claims protection for the application of reagents for detecting HN1L gene expression, HN1L mRNA expression, and / or HN1L protein expression in the preparation of a prognostic kit for esophageal squamous cell carcinoma, wherein patients with high expression of HN1L gene, HN1L mRNA, and / or HN1L protein, and patients with relatively low expression, have a poor prognosis.

[0017] The present invention also claims protection for the use of reagents for detecting HN1L gene expression, HN1L mRNA expression and / or HN1L protein expression in the preparation of a chemotherapy sensitivity test kit for patients with esophageal squamous cell carcinoma, wherein patients with high expression of HN1L gene, HN1L mRNA and / or HN1L protein, and relatively low expression, have poor chemotherapy sensitivity.

[0018] Preferably, the chemotherapy drugs used are docetaxel or cisplatin.

[0019] The present invention also claims protection for the use of inhibitors of HN1L gene expression, inhibitors of HN1L mRNA expression, and / or inhibitors of HN1L protein expression in the preparation of drugs for esophageal squamous cell carcinoma.

[0020] The present invention also claims protection for the use of inhibitors of HN1L gene expression, inhibitors of HN1L mRNA expression, and / or inhibitors of HN1L protein expression in the preparation of potentiators for chemotherapy drugs for esophageal squamous cell carcinoma.

[0021] Preferably, the chemotherapy drug is docetaxel or cisplatin.

[0022] The present invention also claims a drug for esophageal squamous cell carcinoma containing an inhibitor of HN1L gene expression, an inhibitor of HN1L mRNA expression, and / or an inhibitor of HN1L protein expression.

[0023] Preferably, it also contains a chemotherapy drug, said chemotherapy drug being docetaxel or cisplatin.

[0024] Compared with the prior art, the present invention has the following beneficial effects:

[0025] This invention, through in vitro and in vivo functional experiments, discovered that knocking down HN1L can inhibit the metastasis and proliferation of esophageal squamous cell carcinoma cells and increase the sensitivity of tumor cells to chemotherapy drugs. Therefore, it can serve as a specific marker gene for diagnosing esophageal squamous cell carcinoma, making the diagnosis of esophageal squamous cell carcinoma more accurate and faster. It can also serve as a drug target for preparing drugs to treat esophageal squamous cell carcinoma, providing a new and effective treatment option, namely, targeting HN1L in combination with chemotherapy drugs to treat esophageal squamous cell carcinoma, providing a new approach to the treatment of esophageal squamous cell carcinoma. Attached Figure Description

[0026] Figure 1To analyze the expression level of the HN1L gene in esophageal squamous cell carcinoma tissue using microarray analysis; (A) In the TCGA (The Cancer Genome Atlas) database, the expression level of HN1L at the mRNA level in esophageal squamous cell carcinoma (ESCC) was higher than that in esophageal adenocarcinoma (ADC) and normal esophageal epithelial tissue; (B) Immunohistochemical staining analysis of the expression level of HN1L in esophageal squamous cell carcinoma and adjacent adjacent esophageal epithelial cells; (C) Immunohistochemical staining analysis of the expression level of HN1L in esophageal squamous cell carcinoma; (D) Staining score of HN1L in normal tissue and esophageal squamous cell carcinoma tissue.

[0027] Figure 2 Correlation analysis of HN1L expression level and clinicopathological features in esophageal squamous cell carcinoma (ESCC) tissues; (A) High expression of HN1L is associated with tumor invasion, lymph node metastasis and later stage; (B) High expression of HN1L in ESCC tissues suggests poor prognosis in patients with esophageal squamous cell carcinoma.

[0028] Figure 3 The results of cell metastasis experiments after knocking down HN1L in esophageal squamous cell carcinoma cells are as follows: (A) HN1L was knocked down using lentiviral transfection, and the expression level of HN1L in KYSE150 esophageal squamous cell carcinoma cells was detected by Western blot; (B) In vitro Transwell assay showed that knocking down HN1L reduced the migration ability of tumor cells; (C) Nude mouse tail vein lung metastasis assay showed that knocking down HN1L significantly inhibited the metastatic ability of tumor cells.

[0029] Figure 4 Results of cell growth experiments after knocking down HN1L in esophageal squamous cell carcinoma cells; (A) In vitro CCK8 cell proliferation assay showed that knocking down HN1L inhibited tumor cell proliferation; (B) Subcutaneous xenograft experiment in nude mice showed that the xenografts formed after knocking down HN1L were smaller in weight (3 mice per group); (C) Immunohistochemical staining analysis showed that the proportion of proliferating cells (Ki67 positive) was lower after knocking down HN1L.

[0030] Figure 5 The results of HN1L knockdown in esophageal squamous cell carcinoma cells were used to assess cell sensitivity to docetaxel. (A) In vitro CCK8 cell proliferation assays showed that HN1L knockdown increased tumor cell sensitivity to docetaxel. (B) Subcutaneous xenograft and drug assays in nude mice showed that silencing HN1L reversed tumor cell resistance to docetaxel.

[0031] Figure 6 To assess the sensitivity of esophageal squamous cell carcinoma cells to cisplatin after HN1L knockdown: Subcutaneous xenograft and drug experiments in nude mice showed that silencing HN1L reversed tumor cell resistance to cisplatin (3 mice per group). Detailed Implementation

[0032] The present invention will be further described in detail below with reference to the accompanying drawings and specific embodiments. These embodiments are for illustrative purposes only and are not intended to limit the scope of the invention. Unless otherwise specified, the experimental methods used in the following embodiments are conventional methods; the materials and reagents used, unless otherwise specified, are commercially available.

[0033] Example 1: High expression of HN1L gene in esophageal squamous cell carcinoma tissue

[0034] I. Experimental Methods

[0035] The mRNA expression levels of the HN1L gene in normal esophageal tissue, esophageal squamous cell carcinoma, and esophageal adenocarcinoma were obtained using the online analysis tool UALCAN (http: / / ualcan.path.uab.edu / analysis.html). The data source for this analysis tool was the TCGA database.

[0036] II. Experimental Results

[0037] The results are as follows Figure 1 As shown in Figure A, the expression levels of the HN1L gene in esophageal squamous cell carcinoma (n=95) and esophageal adenocarcinoma (n=89) were higher than those in normal esophageal tissue (n=11, P<0.001). Furthermore, the expression level of the HN1L gene in esophageal squamous cell carcinoma was higher than that in esophageal adenocarcinoma tissue (P<0.001).

[0038] Example 2: High expression of HN1L protein in esophageal squamous cell carcinoma tissue

[0039] I. Experimental Methods

[0040] Immunohistochemical staining was performed on esophageal squamous cell carcinoma tumor tissues and adjacent normal tissues to compare the expression levels of the HN1L gene in these tissues. Esophageal squamous cell carcinoma tumor tissues and paired adjacent normal esophageal epithelial tissue samples were collected from Linzhou Cancer Hospital, Henan Province.

[0041] The specific method for immunohistochemical staining is as follows:

[0042] (1) Dewaxing and hydration of tissue sections: Place the tissue sections in a 65℃ oven for 2 hours; soak in xylene three times for 10 minutes each time; soak in gradient alcohol (100%, 95%, 75%, 50%) four times for 5 minutes each time; then inactivate endogenous peroxidase with 3% H2O2, soak for 10 minutes, and soak in deionized water for 3 minutes.

[0043] (2) Remove the slide along with the beaker and let it cool naturally (do not remove the slide directly). Wash with PBS 3 times, 5 min each time.

[0044] (3) Spin the slide and wipe the liquid around the tissue with lint-free paper (be careful to avoid drying the tissue). Place it flat in a humidified box, circle the tissue on the slide with an oil-based marker, and add 5% BSA blocking solution to the circled tissue. Then place it in a humidified box and incubate at 37°C for 30 min.

[0045] (4) Discard the blocking solution, add HN1L specific primary antibody working solution (purchased from Sigma, USA, catalog number: #HPA041908, working concentration: 1:4000 dilution), place in a humidified chamber, and incubate overnight at 4°C. The next day, remove the humidified chamber and place at room temperature for 10 min, then wash three times with PBS for 5 min each time.

[0046] (5) Split the slide, add secondary antibody working solution (purchased from Dako, Denmark, product number: #K5007, ready-to-use) to the circled tissue, place it in a humidified chamber, and incubate at 37°C for 30 min.

[0047] (6) Wash three times with PBS, 5 minutes each time. Shake off and wipe the liquid around the tissue section, being careful not to let the tissue dry out. Place the slide flat in the humidified chamber, add the prepared DAB working solution, and control the color development under a light microscope. After complete color development, gently rinse with distilled water to stop the color development.

[0048] (7) Counterstain with hematoxylin for 30 seconds, rinse with running water, then soak in differentiation solution for 1-3 seconds, rinse again with running water, and the tissue will turn blue. Finally, dehydrate with a gradient of 50%, 75%, 95%, and 100% alcohol for 3 minutes each.

[0049] (8) Clear the slide with xylene, repeat twice, 5 min each time. Mount the slide with neutral resin and photograph it under a light microscope.

[0050] HN1L expression was assessed using immunohistochemical staining scores: staining intensity score: negative, 0; weakly positive, 1; moderately positive, 2; strongly positive, 3. The proportion of HN1L-positive cells was: <25%, 1; 25%-50%, 2; 50%-75%, 3; >75%, 4. The total staining score was calculated as staining intensity × positive proportion. A paired t-test (two-tailed) was used to statistically analyze the difference in HN1L staining scores between esophageal squamous cell carcinoma tissue and normal esophageal epithelial tissue. P < 0.05 was considered statistically significant.

[0051] II. Experimental Results

[0052] The results are as follows Figure 1As shown in B, the expression level of HN1L protein is low in adjacent esophageal epithelial cells, and it is mainly expressed in the nucleus and cytoplasm of esophageal squamous cell carcinoma cells.

[0053] The results are as follows Figure 1 As shown in C and 1D, immunohistochemical staining using an esophageal squamous cell carcinoma tissue microarray (n=229) revealed that the expression level of HN1L protein in the tumor tissue was significantly higher than that in the adjacent normal esophageal epithelial tissue (P<0.001).

[0054] Example 3: High expression of HN1L in esophageal squamous cell carcinoma tissue is associated with poor patient prognosis.

[0055] I. Experimental Methods

[0056] Based on the HN1L staining score in esophageal squamous cell carcinoma tissue as described in Example 2, samples were divided into a high HN1L expression group and a low HN1L expression group using the ROC curve method. The chi-square test was used to analyze the correlation between HN1L expression level and local tumor invasion, lymph node metastasis, and later tumor stage. The Kaplan-Meier method was used to analyze the correlation between HN1L expression level and overall survival. P < 0.05 was considered statistically significant.

[0057] II. Experimental Results

[0058] The results are as follows Figure 2 As shown, correlation analysis of clinicopathological data revealed that upregulation of HN1L in esophageal squamous cell carcinoma tissue was associated with local tumor invasion (P<0.001), lymph node metastasis (P<0.05), and later tumor stage (P<0.001). Figure 2 As shown in A), it was significantly associated with poor patient prognosis (P = 0.038) (as shown in A). Figure 2 (As shown in B), the difference was statistically significant. The results indicate that high expression of HN1L protein in esophageal squamous cell carcinoma tissue suggests a poor prognosis for patients.

[0059] Example 4: Establishment of an esophageal squamous cell carcinoma cell line with stable HN1L knockdown using lentivirus-mediated gene interference technology.

[0060] I. Experimental Methods

[0061] Using Lipo3000 transfection reagent (purchased from Dojindo, Japan, catalog number: #H357), the lentiviral interference vector psi-LVRU6P containing shRNA (vector purchased from GeneCopoeia, USA, interference sequence: 5'-GGCGTAAGCAGAAACACTAAC-3') and three lentiviral packaging vectors pLP1, pLP2, and pLP-VSVG (purchased from Invitrogen, USA) were transduced into 293FT tool cells. After 48 hours, the viral supernatant was collected and transfected into esophageal squamous cell carcinoma cells. The cells were further screened for two weeks using the drug Puromycin (2 μg / ml) to establish a stable HN1L knockdown cell line, resulting in K150-shHN1L cells.

[0062] Interference negative control group: K150-Scramble cells were obtained by performing the same operation on scramble shRNA vector (purchased from GeneCopoeia, USA).

[0063] Western blot was used to verify the protein expression level of HN1L in K150-shHN1L cells and K150-Scramble cells.

[0064] II. Experimental Results

[0065] The results are as follows Figure 3 As shown in Figure A, the esophageal squamous cell carcinoma cells with stable HN1L knockdown (K150-shHN1L) were analyzed by Western blot. The results showed that HN1L was knocked down at the protein level compared to the scramble interference negative control group (K150-Scramble).

[0066] Example 5: Knockdown of HN1L inhibits esophageal squamous cell carcinoma metastasis

[0067] I. Experimental Methods

[0068] (1) Transwell migration experiment

[0069] K150-Scramble cells and K150-shHN1L cells prepared in Example 4 during the logarithmic growth phase were added to a medium containing 0.1% FBS and starved for 24 hours. Cells were digested with trypsin, centrifuged at 1000 rpm for 5 minutes, resuspended in medium, counted, and diluted to 1×10⁶ cells with serum-free medium. 5Add 0.5 ml of medium to the upper chamber, and add 0.75 ml of medium containing 10% FBS to the lower well. Incubate at 37°C and 5% CO2 for 24 hours. Remove non-invasive cells with a small cotton swab, fix with alcohol for 10 minutes, and stain with crystal violet for 30 minutes. Aspirate the crystal violet, rinse with tap water, air dry, and mount on a glass slide. Observe and photograph under a microscope. Randomly select 10 fields of view and calculate the average number of migrating cells per field.

[0070] (2) Nude mouse tail vein lung transfer experiment

[0071] Female BALB / c nude mice, 4–5 weeks old and weighing 18–20 g, were purchased from the Guangdong Provincial Medical Laboratory Animal Center. The mice were housed in an SPF-grade environment for one week to acclimatize. K150-Scramble and K150-shHN1L cells prepared in Example 4 during the logarithmic growth phase were collected, trypsinized, centrifuged at 1000 rpm for 5 minutes, resuspended in culture medium, and counted. The injection volume was 5 × 10⁻⁶ cells / year. 5 Cells / 100 μl. First, nude mice were anesthetized with isoflurane gas and their limbs were fixed with tape. A 1 ml insulin needle was inserted into the tail vein of the mouse, and the cell suspension was slowly injected. Two months later, the mice were euthanized by cervical dislocation, and lung tissue was surgically harvested and fixed in 10% neutral formalin solution. The tissue was then embedded in paraffin, sectioned, and stained with hematoxylin and eosin. Finally, the presence of tumor cells was confirmed by microscopic observation.

[0072] II. Experimental Results

[0073] In vitro Transwell migration assays (e.g.) Figure 3 The results (shown in B) showed that fewer tumor cells (K150-shHN1L) migrated through the Transwell chambers after HN1L knockdown compared to control cells (K150-Scramble).

[0074] In vivo nude mouse tail vein lung transfer experiment (e.g.) Figure 3 The results (shown in C) showed that esophageal squamous cell carcinoma cells knocked down by HN1L (K150-shHN1L) formed fewer nodules in the lungs compared to control cells (K150-Scramble).

[0075] The results of in vivo and in vitro experiments indicate that knocking down HN1L significantly inhibits the metastatic ability of esophageal squamous cell carcinoma cells.

[0076] Example 6: Knockdown of HN1L inhibits the growth of esophageal squamous cell carcinoma cells

[0077] I. Experimental Methods

[0078] (1) In vitro CCK-8 cell proliferation experiment

[0079] K150-Scramble and K150-shHN1L cells prepared in Example 4 during the logarithmic growth phase were taken, trypsinized, centrifuged at 1000 rpm for 5 minutes, resuspended in DMEM complete medium, counted, and the cell suspensions were appropriately diluted and added to 96-well plates (1000 cells / well), 100 μl per well. Four parallel wells were set up for each group, and the cells were continuously cultured at 37℃ and 5% CO2 for 3 days. At the same time each day, 10 μl of CCK-8 solution (purchased from Dojindo, Japan, catalog number: #CK04) was added, and the cells were cultured for another 2 hours. After complete color development, the OD value of each well was measured at 450 nm using an ELISA reader. Cell growth curves were plotted based on the OD values ​​of each well.

[0080] (2) Subcutaneous tumor formation experiment in nude mice

[0081] Female BALB / c nude mice, 4–5 weeks old and weighing 18–20 grams, were purchased from the Guangdong Provincial Medical Laboratory Animal Center. The mice were housed in an SPF-grade environment for one week to acclimatize. K150-Scramble and K150-shHN1L cell suspensions in logarithmic growth phase, prepared in Example 4, were injected into the left and right sides of the mice's backs, respectively. The injection volume was 4 × 10⁻⁶ cells / mL. 6 One cell / 100 μl. After four weeks, the mice were sacrificed, the tumor tissue was removed, and the tumor weight was measured using an electronic balance.

[0082] Immunohistochemical staining: The specific procedure is the same as in Example 2. Ki67 specific antibody was purchased from Abcam, USA, catalog number: #ab16667, working concentration: 1:400.

[0083] II. Experimental Results

[0084] In vitro CCK8 cell proliferation assay (e.g.) Figure 4 As shown in Figure A, the proliferation rate of cells (K150-shHN1L) after knocking down HN1L was lower than that of control cells (K150-Scramble).

[0085] Nude mouse subcutaneous xenograft tumor experiment (e.g.) Figure 4 As shown in B, the xenografts formed by knocking down HN1L in esophageal squamous cell carcinoma cells (K150-shHN1L) were smaller in weight than those formed by control cells (K150-Scramble) (P<0.05).

[0086] Immunohistochemical staining analysis (e.g.) Figure 4 (As shown in C) Further verification confirmed that the proportion of proliferating cells (Ki67 positive) in the xenograft was low after knocking down HN1L.

[0087] The results of in vitro and in vivo experiments showed that knocking down HN1L significantly reduced the proliferative capacity of esophageal squamous cell carcinoma cells.

[0088] Example 7: Knocking down HN1L enhances the sensitivity of esophageal squamous cell carcinoma cells to docetaxel.

[0089] I. Experimental Methods

[0090] (1) In vitro CCK8 cell proliferation experiment

[0091] The K150-Scramble and K150-shHN1L cells prepared in Example 4 were treated with different concentration gradients of docetaxel. After 24 hours, the cell proliferation activity was detected using the CCK-8 reagent. The specific experimental procedures were the same as in Example 6.

[0092] (2) Subcutaneous xenografts and drug experiments in nude mice

[0093] K150-Scramble and K150-shHN1L cells prepared in Example 4 were subcutaneously implanted into nude mice, with an injection volume of 4 × 10⁴ cells. 6 One cell / 100 μl. Two weeks after tumor cell injection, docetaxel (purchased from Chia Tai Tianqing Pharmaceutical Group Co., Ltd., China, dosage: 3 mg / kg mouse body weight) was administered intraperitoneally using a sterile syringe, once every three days for a total of four doses. One week after the end of treatment, the mice were euthanized by cervical dislocation. The tumor tissue was surgically removed, and the tumor weight was measured using an electronic balance.

[0094] II. Experimental Results

[0095] The results are as follows Figure 5 As shown, HN1L knockdown stable cell lines and control cells were treated with docetaxel, a first-line chemotherapy drug for esophageal squamous cell carcinoma. In vitro CCK8 cell proliferation assays revealed that HN1L knockdown increased the sensitivity of tumor cells to docetaxel (e.g., Figure 5 As shown in Figure A). Subcutaneous xenograft tumors in nude mice and drug experiments further confirmed that silencing HN1L can reverse tumor cell resistance to docetaxel (e.g., Figure 5 (as shown in B).

[0096] Example 8: Knockdown of HN1L enhances the efficacy of cisplatin against esophageal squamous cell carcinoma cells.

[0097] I. Experimental Methods

[0098] K150-Scramble and K150-shHN1L prepared in Example 4 were subcutaneously implanted in nude mice, with an injection volume of 4 × 10⁻⁶. 6One cell / 100 μl. Two weeks after tumor cell injection, cisplatin (purchased from Jiangsu Hansoh Pharmaceutical Group Co., Ltd., China, dosage: 5 mg / kg mouse body weight) was administered intraperitoneally using a sterile syringe, once every three days for a total of four doses. One week after the end of treatment, the mice were euthanized by cervical dislocation. The tumor tissue was surgically removed, and the tumor weight was measured using an electronic balance.

[0099] II. Experimental Results

[0100] The results are as follows Figure 6 As shown, the results of subcutaneous xenograft and drug experiments in nude mice showed that the xenografts derived from K150-shHN1L treated with cisplatin (DDP) were lighter in weight, indicating that silencing HN1L can enhance the killing effect of cisplatin on esophageal squamous cell carcinoma cells.

[0101] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit the scope of protection of the present invention. For those skilled in the art, other variations or modifications can be made based on the above description and ideas, and it is neither necessary nor possible to exhaustively describe all implementation methods here. Any modifications, equivalent substitutions, and improvements made within the spirit and principles of the present invention should be included within the scope of protection of the claims of the present invention.

Claims

1. The application of inhibitors of HN1L gene expression, HN1L mRNA expression, and / or HN1L protein expression in the preparation of potentiators for esophageal squamous cell carcinoma chemotherapy drugs, wherein the chemotherapy drug is docetaxel or cisplatin. The inhibitors of HN1L gene expression, HN1L mRNA expression, and / or HN1L protein expression are shRNAs with the interfering sequence GGCGTAAGCAGAAACACTAAC.

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