Fam166a gene missense mutation molecular marker and application thereof in esophageal cancer risk assessment or prognosis

Abstract

This invention relates to the field of biotechnology, specifically to the FAM166A gene missense mutation molecular marker and its application in esophageal cancer risk assessment or prognosis. It was found that the expression level of FAM166A in esophageal cancer patient tissues was significantly higher than that in normal controls, and its expression level was positively correlated with the occurrence and development of esophageal cancer. In vitro cell function experiments confirmed that the FAM166A gene P84L missense mutant, compared with wild-type FAM166A, significantly enhanced the proliferation, invasion, and migration abilities of esophageal cancer KYSE150 cells. Upregulation of wild-type FAM166A significantly promoted the malignant phenotype of esophageal cancer cells, while upregulation of the P84L mutant enhanced this effect. An esophageal cancer cell model stably overexpressing wild-type FAM166A and the P84L mutant was constructed, which can serve as a drug screening platform for high-throughput screening of anti-tumor candidate drugs targeting FAM166A or its mutants.

Description

Technical Field

[0001] This invention relates to the field of biotechnology, specifically to the molecular marker of missense mutation in the FAM166A gene and its application in risk assessment or prognosis of esophageal cancer. Background Technology

[0002] Esophageal cancer (EC) is one of the most common malignant tumors of the digestive tract. Despite advancements in comprehensive treatments such as surgery, radiotherapy, chemotherapy, and immunotherapy, offering the possibility of radical cure for some patients, the highly insidious and nonspecific early symptoms of esophageal cancer mean that over 70% of patients are already in the middle or late stages at initial diagnosis, resulting in a low overall 5-year survival rate and poor prognosis. Currently, the clinical diagnosis of esophageal cancer mainly relies on endoscopy and pathological biopsy; however, these methods are invasive, expensive, and have poor adherence in asymptomatic individuals, making them unsuitable for large-scale screening. Therefore, identifying non-invasive biomarkers for early diagnosis and risk assessment has become a key breakthrough in improving the prognosis of esophageal cancer.

[0003] However, the molecular mechanisms underlying the development and progression of esophageal cancer are not yet fully understood, and there is a lack of highly sensitive and specific early diagnostic biomarkers. Current research largely focuses on gene mutations or abnormal protein expression within tumor tissue itself, while systematic exploration of the tumor microenvironment, epigenetic alterations, and circulating biomarkers (such as liquid biopsy) remains insufficient. This severely restricts early warning, personalized treatment, and efficacy monitoring of esophageal cancer. Therefore, elucidating the molecular mechanisms of esophageal cancer pathogenesis and identifying novel biomarkers for early diagnosis, disease assessment, and prognosis is of significant clinical value and an urgent research need for achieving early detection and treatment of esophageal cancer, reducing mortality, and improving patients' quality of life.

[0004] The FAM family of genes, a group of genes with similar protein sequences but not yet fully characterized, has been the subject of numerous studies demonstrating its crucial role in the malignant development of various human cancers, influencing tumor proliferation, invasion, migration, and drug resistance. Furthermore, specific members of the FAM family have been recognized as promising therapeutic targets and prognostic biomarkers for multiple cancer types. FAM166A, located at 9q34.3, has a 954bp CDS region encoding 318 amino acids. Previous research has shown that FAM166A is a conserved microtubule-inner protein (MIP), specifically highly expressed primarily in the testes and widely distributed in the cilia and flagella of eukaryotes. It binds to the tubulin interface, participating in the stability and functional regulation of microtubule structure and playing a major role in reproductive system diseases. However, the relationship between FAM166A and tumorigenesis and development has not yet been reported in the literature.

[0005] During their research, the inventors' team discovered that immunohistochemical staining of esophageal cancer paraffin sections showed significantly higher FAM166A expression levels in esophageal cancer patient tissue sections compared to normal patients, and that FAM166A expression levels in Kazakh esophageal cancer tissue sections were significantly higher than those in Han Chinese patients. Further investigation, using a wild-type FAM166A overexpression plasmid to upregulate FAM166A expression in KYSE150 cells after in vitro culture, significantly promoted cell proliferation, invasion, and migration. However, upregulation of the FAM166A P84L mutant further promoted cell proliferation, invasion, and migration on top of FAM166A overexpression. These results suggest that the FAM166A gene missense mutant and its potential role as a biomarker in promoting tumor progression could lead to the development of drugs that inhibit FAM166A and its mutant proteins for controlling the occurrence and progression of esophageal cancer. Summary of the Invention

[0006] The primary objective of this invention is to provide the application of a reagent for detecting a missense mutation molecular marker in the FAM166A gene in the preparation of an esophageal cancer risk assessment product. The molecular marker is characterized in that it is located at base position 140140111 in the coding region of the FAM166A gene and has a polymorphism of G or A.

[0007] A second objective of this invention is to provide the application of a reagent for detecting a missense mutation molecular marker in the FAM166A gene in the preparation of a kit for assessing the risk of esophageal cancer or determining its prognosis for non-disease diagnostic purposes. The molecular marker is located at base 140140111 in the coding region of the FAM166A gene and has a polymorphism of G or A.

[0008] A third objective of this invention is to provide the application of a reagent for detecting the P84L missense mutation in the FAM166A gene in the preparation of a kit for assessing the risk of progression or prognosis of esophageal cancer; wherein the P84L missense mutation refers to the mutation of amino acid 84 in the coding sequence of the FAM166A gene from proline to leucine.

[0009] A fourth object of the present invention is to provide a pharmaceutical composition for inhibiting the proliferation, invasion, and / or migration of esophageal cancer cells, comprising: (a) Inhibitors that suppress the expression of the FAM166A gene or its encoded protein; and / or (b) Mutation-specific inhibitors that suppress protein expression in the FAM166A P84L mutant; And pharmaceutically acceptable carriers or excipients.

[0010] Preferably, the inhibitor is selected from: siRNA, shRNA, antisense oligonucleotide, CRISPR-Cas9 gene editing system, small molecule inhibitor, or monoclonal antibody or antigen-binding fragment thereof that specifically binds to the FAM166A protein.

[0011] Preferably, the inhibitor is an antibody or small molecule compound that specifically inhibits the FAM166A P84L mutant protein without inhibiting the wild-type FAM166A protein.

[0012] A fifth objective of this invention is to provide the use of reagents for inhibiting the expression of the FAM166A gene or the FAM166A P84L mutant protein in the preparation of medicaments for treating esophageal cancer, wherein the treatment is achieved by inhibiting the proliferation, invasion, and / or migration of esophageal cancer cells.

[0013] Preferably, the esophageal cancer is an esophageal cancer with high expression of the FAM166A gene and / or carrying the FAM166A P84L missense mutation.

[0014] The sixth objective of this invention is to provide a kit for the diagnosis or prognosis of esophageal cancer, comprising: Primers or probes for specific detection of FAM166A gene mRNA; and / or Antibodies that specifically detect FAM166A protein; and / or Sequencing primers or mutation-specific probes for specific detection of the P84L mutation in the FAM166A gene.

[0015] A seventh objective of this invention is to provide a method for screening candidate drugs for treating esophageal cancer carrying the FAM166A missense mutation, comprising:

[0016] (1) The candidate drug was exposed to esophageal cancer cells that overexpressed the FAM166A P84L mutant protein;

[0017] (2) To detect the cell's ability to proliferate, invade, or migrate;

[0018] (3) Using untreated homologous cells as a control, select compounds that significantly inhibit the ability described in step (2) as candidate drugs.

[0019] The beneficial effects of this invention are: (1) Through analysis of clinical tissue samples, this invention has for the first time discovered that the expression level of FAM166A in esophageal cancer patient tissues is significantly higher than that in normal controls, and its expression level is positively correlated with the occurrence and development of esophageal cancer. This suggests that FAM166A can be used as a novel biomarker for the auxiliary diagnosis or risk warning of esophageal cancer. Furthermore, this invention has for the first time discovered that the expression level of FAM166A in esophageal cancer patient tissues of Kazakh ethnicity is significantly higher than that of Han ethnicity patients, suggesting that this biomarker can be used for risk stratification and individualized prognostic assessment of esophageal cancer patients of different ethnicities, filling the gap in the existing technology for the lack of molecular biomarkers for esophageal cancer in specific ethnic groups.

[0020] (2) Through in vitro cell function experiments, this invention has for the first time demonstrated that the P84L missense mutant of the FAM166A gene, compared with the wild-type FAM166A, can significantly enhance the proliferation, invasion, and migration of esophageal cancer KYSE150 cells. This discovery reveals that the P84L mutation is a gain-of-function mutation with stronger oncogenic activity than the wild-type.

[0021] (3) This invention demonstrates through overexpression experiments that upregulation of wild-type FAM166A significantly promotes the malignant phenotype of esophageal cancer cells, while upregulation of the P84L mutant further enhances this effect. This provides direct functional evidence for inhibiting FAM166A or its mutants as a therapeutic strategy for esophageal cancer.

[0022] (4) This invention constructs a stable esophageal cancer cell model (KYSE150) that overexpresses FAM166A wild-type and P84L mutant. This model can be used as a drug screening platform for high-throughput screening of anti-tumor candidate drugs targeting FAM166A or its mutants. Specific primers, probes, or antibodies for detecting FAM166A expression levels and P84L mutations are also provided, which can be used to prepare diagnostic kits or companion diagnostic kits to achieve patient stratification and precision treatment. Attached Figure Description

[0023] Figure 1 The expression level of FAM166A in the esophageal tissues of healthy Han and Kazakh individuals and esophageal cancer patients in Han and Kazakh individuals.

[0024] Figure 2Quantitative graph of immunohistochemistry results.

[0025] Note: FAM166A wild-type plasmid and FAM166A P84L mutant plasmid were used to treat KYSE250 cells for 24 hours.

[0026] Figure 3 The proliferation level of esophageal cancer cells in each group

[0027] Figure 4 : Area under the curve and quantization of the CCK8 results graph

[0028] Figure 5 The invasive and migratory abilities of esophageal cancer cells in each group.

[0029] Figure 6 Quantification of Transwell transfer results

[0030] Figure 7 Quantification of Transwell invasion results

[0031] Figure 8 Cell scratch assay was used to assess the migration ability of esophageal cancer cells in each group.

[0032] Figure 9 Quantification of cell scratch results. Detailed Implementation

[0033] The following description, in conjunction with the accompanying drawings, illustrates exemplary embodiments of the present invention, including various details to aid understanding. These details should be considered merely exemplary. Therefore, those skilled in the art will recognize that various changes and modifications can be made to the embodiments described herein without departing from the scope and spirit of the invention. Similarly, for clarity and brevity, descriptions of well-known functions and structures are omitted in the following description.

[0034] Unless otherwise defined, the technical terms used in the following embodiments have the same meanings as commonly understood by those skilled in the art. Unless otherwise specified, the experimental reagents used in the following embodiments are all conventional biochemical reagents; and the experimental methods described are all conventional methods.

[0035] In the following examples, the detection results are expressed as Mean ± SEM, and SPSS 25.0 software was used for statistical analysis. When the data conformed to a normal distribution, t-tests and one-way ANOVA (for comparisons among multiple groups) were used; when the data did not conform to a normal distribution, nonparametric rank-sum tests were used. GraphPad Prism 8.0.1 software was used for graphing. Data are expressed as mean ± SEM. * P<0.05, **P<0.01, *** P<0.001, the difference is statistically significant.

[0036] In the following embodiments, the gene at position 140140111 G>A (Phe→Leu) of Family With SequenceSimilarity 166 Member A (FAM166A) is located at 9q34.3.

[0037] Example 1: Screening Experiment

[0038] 1. Sampling

[0039] This study included 40 participants from Xinjiang, China (randomly selected individuals aged 40-60 years, with an average number of males and females within each group, and no distinction made based on BMI, etc.). Samples were sent to Shanghai Ouyi Biomedical Technology Co., Ltd. for whole-exome sequencing and divided into Han (n=20) and Kazakh (n=20) ethnic groups.

[0040] 2. Results

[0041] The sequencing results are shown in Table 1. After analysis and comparison, a group of missense mutation genes with a high mutation rate were screened out in Kazakh subjects. Among them, the missense mutation of Family With Sequence Similarity 166 Member A (FAM166A) at position 140140111 G>A (Phe→Leu) occurred only in Kazakhs, with a mutation rate of 25% in Kazakhs and 0% in Han Chinese, showing a significant difference.

[0042] Table 1 Sequencing results

[0043] Example 2: Clinical Sample Experiment

[0044] 1. Sampling

[0045] This study included paraffin-embedded tissue samples from 80 individuals in Xinjiang, China. Among them, 40 were diagnosed with esophageal cancer (20 Kazakhs and 20 Han Chinese), and the remaining 40 were diagnosed with other non-esophageal cancer conditions (20 Kazakhs and 20 Han Chinese). Exclusion criteria included other tumors and malignant wasting diseases.

[0046] 2. Detection Method

[0047] 2.1 Immunohistochemical staining:

[0048] ① Place paraffin sections in an oven at 60℃ for 30 minutes; ② Dewax with xylene for 3×5 minutes; ③ Dexylene with alcohol for 3×5 minutes; ④ Use sodium citrate retrieval solution, preheat in a microwave oven, then in a pressure cooker, bring to a boil, reduce to 800W, and cook for 8 minutes before turning off the heat; ⑤ Place the retrieval box in a water basin and allow it to reach room temperature; ⑥ Treat the sections with 3% hydrogen peroxide in the dark for 10 minutes to inactivate endogenous peroxidase activity; ⑦ Wash with PBS for 3×5 minutes; ⑧ Add an appropriate amount of diluted primary antibody and incubate overnight at 4℃. Day 2: ① Warm to 37℃ for 30 minutes; ② Wash away primary antibody with PBS for 3×5 minutes; ③ Add biotin-labeled secondary antibody (PV6000) and incubate at room temperature or 37℃ for 30~60 minutes; ④ Wash away secondary antibody with PBS for 3×5 minutes; prepare DAB chromogenic solution, 50~100μl per slide; ⑤ Immediately rinse away the chromogenic solution with tap water after chromogenic development; ⑥ Stain the nucleus with hematoxylin for 3~5 minutes, then with acid alcohol for 1~3 seconds, and rinse with water; ⑦ Dehydrate with alcohol and clear with xylene; ⑧ Mount the slides with an appropriate mounting medium (neutral resin).

[0049] 3. Results

[0050] Immunohistochemical staining results showed that the expression level of FAM166A in esophageal cancer tissue sections was significantly higher than that in normal patients, and the expression level of FAM166A in esophageal cancer tissue sections of Kazakhs was significantly higher than that of Han Chinese. Figure 1-2 ).

[0051] Example 2: Cell Experiment

[0052] 1. Materials and Methods

[0053] Human esophageal squamous cell carcinoma cell line KYSE150 was cultured in vitro using 1640 medium + 10% FBS (fetal bovine serum) + 1% penicillin (100 μg / mL) / streptomycin (100 μg / mL) and routinely cultured in a 37℃ incubator containing 5% CO2.

[0054] Wild-type and mutant overexpression plasmids and their corresponding negative control vectors were all purchased from Shanghai Jima Pharmaceutical Technology Co., Ltd.

[0055] Cell treatment method: FAM166A was overexpressed in the human esophageal squamous cell carcinoma cell line KYSE150 (the treatment method for wild-type and mutant FAM166A plasmids is the same; the wild-type FAM166A overexpression plasmid is used as an example here):

[0056] (1) Preparation of reagents before operation: Lipofectamine 2000 (Lipo2000) or Lipofectamine 3000 (Lipo3000) containing dry powder of FAM166A wild-type overexpression plasmid, centrifuge at 12000 rpm for 1 minute, avoid bumping or tilting, open the cap vertically, add DEPC water according to the ratio, gently aspirate and then cap the tube to allow the powder to dissolve fully, keep at 4℃ for later use, and can be stored for a long time at -20℃.

[0057] (2) The amount of transfection reagent used is taken as an example of KYSE150 human esophageal cancer cells in 12-well plate: Prepare 4 enzyme-free EP tubes. Add 300uL of RPMI1640 and an appropriate amount of P3000 to each of the 2 EP tubes. Incubate with a certain dose of FAM166A, NC plasmid or interference fragment for 5 minutes. Add 300uL of RPMI1640 and an appropriate amount of Lipo3000 to each of the 2 EP tubes. Add the two tubes containing the plasmid to the tubes containing Lipo3000 respectively, gently pipette and mix, and incubate at room temperature for 20 minutes (the entire transfection process should be completed within 40 minutes).

[0058] (3) The amount of transfection reagent used is taken as KYSE150 human esophageal cancer cells in 12-well plate: Prepare 4 enzyme-free EP tubes. Add 600uL of RPMI1640 and an appropriate amount of Lipo2000 to each of the 2 EP tubes and incubate for 5 minutes. Add a certain dose of FAM166A, NC plasmid or interference fragment and 600uL of RPMI1640 to the other 2 EP tubes and mix. Transfer the mixture containing Lipo2000 to the mixture containing FAM166A, NC plasmid or interference fragment, gently pipette and mix, and incubate at room temperature for 20 minutes (the entire transfection process should be completed within 40 minutes).

[0059] (4) During incubation, discard the cell supernatant in the 12-well plate, add 1 mL of 1×PBS buffer to each well to wash once, and then add 0.9 mL of RPMI 1640.

[0060] (5) Add the mixture that has been incubated for 20 minutes to the 12-well plate, 100 μL per well, and place it stably in a 37°C cell culture incubator for 4-6 hours.

[0061] (6) After 4-6 h, discard the supernatant, add 2 mL of culture medium to each well (culture medium preparation: 45 mL RPMI 1640 + 5 mL LFBS), and place it stably in a 37℃ cell culture incubator for 24-36 h.

[0062] 2. Detection Method

[0063] 2.1 Detection of tumor cell proliferation capacity:

[0064] (1) Cell seeding: Take cells in the logarithmic growth phase, digest with trypsin, centrifuge, add an appropriate amount of culture medium to prepare a single-cell suspension, and seed 1*10 cells per well. 4 One cell was seeded in a 96-well plate with 100 μl of culture medium per well.

[0065] (2) Cell culture: The cells are cultured in an incubator for 24 hours.

[0066] (3) Color development: Add 10 μl of CCK-8 solution to each well, gently shake to mix, and continue to incubate in the incubator for 2-3 hours.

[0067] (4) Colorimetric analysis: Place the 96-well plate into the microplate reader, select the wavelength of 450nm, measure the light absorption value of each well, save the results, and plot the cell growth curve based on the absorbance value.

[0068] 2.2 Detection of tumor cell invasiveness:

[0069] (1) Place the pipette tips, chambers, 24-well plates, and serum-free 1640 medium used in the experiment into a -20℃ pre-cooling chamber for 1 hour. Dilute the pre-cooled medium with the matrix gel at a ratio of 1:8, take 90 μl and spread it evenly into the upper chamber of the chamber, and incubate at 37℃ for at least 6 hours to promote matrix gel solidification.

[0070] (2) Wash the gel gently with 1640 medium to hydrate the basement membrane.

[0071] (3) Add 250 μl of cell suspension (containing approximately 12*10 cells) to each well in the upper chamber. 4 (Cells), ensuring even distribution of cells. Add 600 μl of culture medium containing 20% ​​fetal bovine serum to each of the upper and lower chambers, ensuring there are no air bubbles between the upper and lower chambers. Continue culturing at 37°C and 5% CO2 for 24 hours.

[0072] (4) Take out the chamber and rinse twice with PBS, pre-cool with methanol at 4°C for 15 minutes, invert and air dry, and stain with 1% crystal violet for 10 minutes.

[0073] (5) Remove uninvaded cells from the chamber with a cotton swab, rinse twice with PBS, observe under a microscope (make sure to keep the chamber membrane moist), take a picture, and count the cells.

[0074] 2.3 Detection of tumor cell migration ability:

[0075] (1) Take cells in the logarithmic growth phase, digest and centrifuge them, and add serum-free 1640 medium to prepare a single-cell suspension.

[0076] (2) Add 250 μl of cell suspension (containing approximately 6*10 cells) to each well in the upper chamber. 4 (100 cells), ensuring the cells are evenly spread. Add 600 μl of culture medium containing 20% ​​fetal bovine serum to each of the upper and lower chambers, ensuring there are no air bubbles between the upper and lower chambers. Continue culturing at 37°C and 5% CO2 for 24 hours.

[0077] (3) Rinse the top and bottom of the chamber twice with PBS, fix it in methanol pre-cooled at 4°C for 15 minutes, remove the chamber, invert it to air dry, and stain it with 1% crystal violet solution for 10 minutes.

[0078] (4) Remove the unmigrated cells on the chamber with a cotton swab, rinse twice with PBS, place the chamber under a microscope for observation (make sure to keep the chamber membrane moist), take a picture, and count the cells.

[0079] 2.4 Scratch Healing Experiment:

[0080] Log-grown EC cells were digested with trypsin and seeded into streaked six-well plates. When the cells were in good growth condition and the confluence reached more than 90%, a linear wound perpendicular to the bottom line was made in the middle of the six-well plate using a sterile medium-sized pipette tip, and the detached cells were washed with sterile PBS. Then, images were taken using a 40× microscope at 0h and 24h to compare the cell migration ability.

[0081] 3. Results:

[0082] In vitro cell results showed that after esophageal cancer cells were treated with wild-type FAM166A overexpression plasmid, the proliferation capacity of esophageal cancer cells overexpressing FAM166A was significantly higher than that of the NC group, as revealed by CCK8 assay and area under the curve analysis. Furthermore, the proliferation capacity of esophageal cancer cells was further enhanced after upregulation of the FAM166A P84L mutant. Figure 3-4 Esophageal cancer cells from the NC group, the group overexpressing FAM166A, and the group overexpressing the FAM166A P84L mutant were cultured in separate chambers. Transwell assays combined with quantitative data analysis revealed that esophageal cancer cells overexpressing FAM166A exhibited significantly higher migration and invasion abilities than those in the NC group. Furthermore, upregulation of the FAM166A P84L mutant further enhanced these migration and invasion abilities. Figure 5-7 Cell scratch assays combined with quantitative analysis further verified that the migration ability of esophageal cancer cells overexpressing FAM166A was significantly higher than that in the NC group, and that the migration ability of esophageal cancer cells was further enhanced after upregulation of the FAM166A P84L mutant. Figure 8-9These results demonstrate the missense mutant of the FAM166A gene and its potential role as a biomarker in promoting tumor progression. Furthermore, the development of drugs that inhibit FAM166A and its mutant proteins could be used to control the occurrence and progression of esophageal cancer.

[0083] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some or all of the technical features; and these modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. The application of a reagent for detecting missense mutation molecular markers in the FAM166A gene in the preparation of esophageal cancer risk assessment products, characterized in that, The molecular marker is located at base 140140111 in the coding region of the FAM166A gene, and its polymorphism is G or A.

2. The application of a reagent for detecting a missense mutation molecular marker in the FAM166A gene in the preparation of a kit for assessing the risk of esophageal cancer or determining its prognosis for non-disease diagnostic purposes, wherein the molecular marker is located at base 140140111 in the coding region of the FAM166A gene and has a polymorphism of G or A.

3. The application of a reagent for detecting the P84L missense mutation in the FAM166A gene in the preparation of a kit, characterized in that, The kit is used for risk assessment or prognosis of esophageal cancer progression; wherein the P84L missense mutation refers to the mutation of amino acid 84 in the coding sequence of the FAM166A gene from proline to leucine.

4. A pharmaceutical composition for inhibiting the proliferation, invasion, and / or migration of esophageal cancer cells, characterized in that, Include: (a) Inhibitors that suppress the expression of the FAM166A gene or its encoded protein; and / or (b) Mutation-specific inhibitors that suppress protein expression in the FAM166A P84L mutant; And pharmaceutically acceptable carriers or excipients.

5. The pharmaceutical composition according to claim 4, characterized in that, The inhibitor is selected from: siRNA, shRNA, antisense oligonucleotides, CRISPR-Cas9 gene editing system, small molecule inhibitors, or monoclonal antibodies or antigen-binding fragments that specifically bind to the FAM166A protein.

6. The pharmaceutical composition according to claim 4, characterized in that, The inhibitor is an antibody or small molecule compound that specifically inhibits the FAM166AP84L mutant protein without inhibiting the wild-type FAM166A protein.

7. The use of a reagent for inhibiting the expression of the FAM166A gene or the FAM166A P84L mutant protein in the preparation of a drug for treating esophageal cancer, characterized in that, The treatment is achieved by inhibiting the proliferation, invasion, and / or migration of esophageal cancer cells.

8. The application as described in claim 7, characterized in that, The esophageal cancer referred to is esophageal cancer with high expression of the FAM166A gene and / or carrying the FAM166A P84L missense mutation.

9. A reagent kit for the diagnosis or prognosis of esophageal cancer, characterized in that, Include: Primers or probes for specific detection of FAM166A gene mRNA; and / or Antibodies that specifically detect FAM166A protein; and / or Sequencing primers or mutation-specific probes for specific detection of the P84L mutation in the FAM166A gene.

10. A method for screening candidate drugs for treating esophageal cancer carrying the FAM166A missense mutation, characterized in that, include: (1) The candidate drug was exposed to esophageal cancer cells that overexpressed the FAM166A P84L mutant protein; (2) To detect the cell's ability to proliferate, invade, or migrate; (3) Using untreated homologous cells as a control, select compounds that significantly inhibit the ability described in step (2) as candidate drugs.

Images