Construction and identification method of melanoma liver metastasis model

Abstract

The present application relates to the field of animal model construction, and in particular to a melanoma liver metastasis model construction method, which uses a spleen injection method to construct a human melanoma A375-M1 cell line liver metastasis model, and is identified in vivo by PET-CT and MRI, and is identified ex vivo by Bouin's and H&E.The human melanoma liver metastasis tumor model constructed by the present application has the advantages of short construction time, simple method and 100% modeling success rate.This metastasis model simulates the development process and pathological characteristics of melanoma liver metastasis, solves the problem of the lack of an ideal animal model for human melanoma during liver metastasis, and helps to explore and screen the development of anti-liver metastasis drugs for human melanoma, and plays a positive promoting role in new strategies for anti-melanoma liver metastasis.

Description

Technical Field

[0001] This invention relates to the field of animal model construction, specifically, to a method for constructing and identifying liver metastasis models using melanoma cells. Background Technology

[0002] Malignant melanoma is a highly aggressive and easily metastatic malignant tumor. In recent years, it has become one of the fastest-growing malignant tumors in terms of incidence, and its mortality rate ranks first among skin malignant tumors. Melanoma is highly malignant and prone to metastasis; approximately 50-80% of patients with advanced melanoma will develop liver metastases, and 8-46% will develop brain or liver metastases.

[0003] In tumor research, model construction is crucial. Among them, metastatic tumor models involve artificially inoculating experimental mice with human or mouse cells. The tumor cells spread to various parts of the body through multiple pathways within the mice, and the tumor cells colonize non-primary organs or tissues to form metastatic lesions, similar to the process of tumor metastasis in humans.

[0004] Currently, preclinical studies of advanced metastatic melanoma primarily utilize murine B16F10 cell models, but these models suffer from differences in the expression of related proteins and antibodies between murine and human cells. Furthermore, the spontaneous metastasis of subcutaneous tumors using the human A375 cell line has also been used in preclinical studies of advanced metastatic melanoma, but this cell line exhibits low success rates in constructing melanoma metastatic models, long modeling times, and highly randomized metastatic sites, making it unsuitable for studying specific organs affected by melanoma metastasis. Therefore, finding a simple and stable human melanoma metastasis model is crucial for melanoma research. Summary of the Invention

[0005] The purpose of this invention is to provide a method for constructing a liver metastasis model using melanoma cells and a method for identification.

[0006] To achieve the above objectives, the present invention adopts the following solution:

[0007] (1) Select 3-4 week old male BALB / c nude mice, each weighing 17-20g. Trypsin digest A375-M1 cells in the logarithmic growth phase, resuspend in pre-cooled PBS, and adjust the concentration to 1.5×10⁻⁶. 7Prepare on ice at a concentration of 10,000 mg / mL. Sterilize surgical instruments by autoclaving. Anesthetize nude mice that have been fasted for 12 hours with 1% pentobarbital at 50 mg / kg. After anesthesia, make a 0.5-1 cm longitudinal incision along the left side of the mouse, slightly below the rib and aligned with the ear. Gently lift the spleen with forceps, grasp the lower end of the spleen with forceps, and insert the needle along the lowest point of the spleen to a depth of one-third of a No. 5 needle. Slowly inject 50 μL of cell suspension over 1 minute. After slowly withdrawing the needle, apply pressure to the injection site with a dry alcohol swab for 2 minutes to stop bleeding. After hemostasis, suture and disinfect the incision, and place the mouse on a 37°C water-warming bag to await recovery. Disinfect the incision with povidone-iodine daily for one week.

[0008] (2) Weekly statistics were collected on changes in liver weight in the control group and the A375-M1 melanoma liver metastasis model.

[0009] (3) The liver slices were tested according to the detection method in Example 7.

[0010] (4) MRI and PET-CT experiments were performed according to the detection methods in Examples 4 and 5.

[0011] (5) Detection of weight changes and survival period of blank control group and model group.

[0012] Based on the above technical solution, in a first aspect, the present invention provides a method for constructing a melanoma liver metastasis model, comprising the following steps:

[0013] A375-M1 cells in logarithmic growth phase were suspended in PBS and the concentration was adjusted to 1.5 × 10⁻⁶. 7 A melanoma liver metastasis model was obtained by injecting 50 μL into the spleen of 3-4 week old male BALB / c nude mice at a concentration of 10,000 μL / mL.

[0014] The present invention uses the A375-M1 cell line, which is derived from the low metastatic melanoma cell population A375 (female, 54 years old, ATCC CRL-1619) cells. The A375-M1 cell line (ATCC CRL-3222) was obtained by intravenously injecting A375 cells into nude mice.

[0015] Furthermore, after culturing A375-M1 melanoma cells to the logarithmic growth phase, they were digested with trypsin, and the cell suspension was centrifuged and resuspended in PBS.

[0016] Furthermore, the liver metastasis of nude mice was examined weekly using MRI, PET-CT, and H&E slides. After 1-2 weeks, a human melanoma nude mouse liver metastasis model was obtained.

[0017] Further, male BALB / c nude mice aged 3-4 weeks with a weight of (16±4) g were selected. A375-M1 melanoma cells were cultured to the logarithmic growth phase, digested with trypsin, and the cell suspension was centrifuged and resuspended in PBS. The cells were injected into the spleen of each nude mouse at a concentration of 1.5 × 10⁻⁶ cells / mL. 7 The A375-M1 cell suspension, with a density of 10,000 cells / mL, was 50 μL. Nude mice were examined weekly using MRI, PET-CT, and H&E sections to assess liver metastasis. A human melanoma liver metastasis model in nude mice was obtained after 1-2 weeks.

[0018] Tumor cells are implanted into the spleen, and as they grow, infiltrate, and invade the splenic vein, they then colonize the liver via the portal vein, thus constructing a liver metastasis model. This method is relatively simple to perform and has a high success rate in locating liver metastases.

[0019] Magnetic Resonance Imaging (MRI) is a medical imaging technique that generates images by utilizing the signals produced by atomic nuclei in living tissue under the influence of strong magnetic fields and radio frequency pulses. MRI uses the interaction between strong magnetic fields and radio frequency pulses and the atomic nuclei in living tissue, generating detailed living images through signal detection, spatial coding, and image reconstruction. This imaging technique is widely used in medical diagnostics, providing non-invasive, high-contrast, and high-resolution structural and functional information.

[0020] Positron emission tomography-computed tomography (PET-CT) is an imaging technique that combines PET and CT. PET provides functional and metabolic information about the tumor, while CT precisely locates the anatomical position of the tumor, offering advantages such as sensitivity, accuracy, specificity, and precise localization. PET-CT can facilitate early diagnosis and differential diagnosis of tumors, stage and restate tumors, aiding in the development of treatment plans and evaluation of treatment effectiveness, and timely identification of tumor recurrence. 2-Fluoro-2-deoxy-D-glucose ( 18 F-FDG is a glucose analog that can be actively taken up and aggregated by highly metabolically active solid tumor tissues. It is a radiopharmaceutical commonly used for tumor detection, staging, assessment, and monitoring of metabolism, and is also the most commonly used imaging agent for PET-CT imaging.

[0021] A second aspect of the present invention provides a method for identifying melanoma liver metastasis models constructed using the method described above, comprising the following steps:

[0022] (A) Take control BALB / C nude mice and BALB / C nude mice that have been modeled for one, two, three, and four weeks. After sacrifice, weigh the liver and perform H&E sections. Analyze and identify the results.

[0023] (B) Take control BALB / C nude mice and BALB / C nude mice that have been modeled for three weeks, and inject 100±10 μCi of fluorine

[18] deoxyglucose injection into the tail vein. 18 F-FDG), PET-CT imaging analysis of the abdomen;

[0024] (C) Take control BALB / C nude mice and BALB / C nude mice that have been modeled for three weeks, and inject 100±10 μCi of fluorine

[18] deoxyglucose injection into the tail vein. 18 F-FDG), MRI imaging analysis of the abdomen.

[0025] The advantages of this invention are:

[0026] 1. This invention provides a novel method for constructing animal models of melanoma liver metastases, different from existing human metastatic animal models. Furthermore, the method is simple and achieves a 100% success rate in constructing metastatic tumors.

[0027] 2. The melanoma liver metastasis animal model constructed by the method described in this invention uses multiple examination methods, including PET-CT, MRI, H&E sections, and Bouin's fixation staining, to detect and identify the formation and development process of the model from multiple aspects and angles.

[0028] 3. This invention simulates the biological characteristics of human melanoma liver metastasis, solving the problem of the lack of ideal animal models in the study of the occurrence, development, invasion and metastasis of human melanoma. It provides a good platform for screening drugs and preparations for melanoma liver metastasis and plays a positive role in exploring new strategies for treating melanoma liver metastasis. Attached Figure Description

[0029] Figure 1 H&E sections of the liver of nude mice injected via the tail vein of A375 were obtained, scale bar 5 mm.

[0030] Figure 2 H&E sections of the liver of nude mice injected via the tail vein of A375-M1 were obtained, scale bar 5 mm.

[0031] Figure 3 H&E sections of nude mouse liver were injected into the spleen of A375 cells. Scale bar: 5 mm.

[0032] Figure 4 H&E sections of nude mouse liver were injected into the spleen of A375-M1. Scale bar: 5 mm.

[0033] Figure 5These are MRI images of the liver metastasis model of this invention. A is a cross-sectional MRI image of a normal nude mouse, and B is a cross-sectional MRI image of a nude mouse with the liver metastasis model. In Figure B, the lesion boundaries are still clear, and a large area of ​​abnormal signal lesions is visible. Scale bar: 800 μm.

[0034] Figure 6 These are PET-CT images of the liver metastasis animal model of the present invention. From left to right in Figure A, they are the transverse, coronal, and sagittal sections of the abdomen of the control group; from left to right in Figure B, they are the transverse, coronal, and sagittal sections of the abdomen of the liver metastasis model group. The area marked in B is a bright signal of obvious abnormal metastatic lesions.

[0035] Figure 7 This is a liver section stained with H&E from the nude mouse liver metastasis model of this invention. Red marks indicate intrahepatic blood vessels, and black marks indicate tumor cell metastases. Group A scale bar: 500 μm; Group B scale bar: 100 μm.

[0036] Figure 8 Four weeks after nude mouse modeling, H&E sections of major organs such as heart, spleen, lung, and kidney were collected. Group A was the normal control group; Group B was the liver metastasis model group. The black marks indicate the tumor areas. Scale bar: 200μm.

[0037] Figure 9 This is a line graph showing the weight changes of the model group and the control group after liver transfer modeling in nude mice according to the present invention, n=6.

[0038] Figure 10 To investigate the survival curves of the model group and the control group after liver metastasis modeling in this invention, n=6.

[0039] Figure 11 This is a graph showing the change in liver weight over time in the model group and the control group after liver metastasis modeling according to the present invention, n=5. Detailed Implementation

[0040] The specific implementation methods provided by the present invention will be described in detail below with reference to the embodiments.

[0041] Example 1: Investigation of Tail Vein Injection Modeling Method

[0042] (1) Select 3-4 week old male BALB / c nude mice, each weighing (17±3)g. After culturing A375 melanoma cells to the logarithmic growth phase, digest them with trypsin. After centrifugation, resuspend the cell suspension in PBS and inoculate each mouse via the tail vein at a concentration of 1.5×10⁻⁶. 7 100 μL of A375 cell suspension (10,000 / mL) was added. One month later, liver sections were harvested for Hematologic and Escherichia coli (H&E) section examination to assess liver metastasis (n=3). Results showed that the liver sections contained tightly packed cells with clear structure, and no metastatic lesions were observed. Figure 1A375 melanoma cells, via tail vein approach, failed to form tumor metastases in the liver of BALB / C nude mice within one month.

[0043] (2) Following the method used in (1), A375 cells were replaced with A375-M1 cells to construct a liver metastasis model. One month later, liver sections were harvested for H&E analysis to examine liver metastasis (n=3). The results showed that the liver sections had tightly packed cells with clear structure and no metastatic lesions were observed. Figure 2 A375-M1 melanoma cells, via tail vein translocation, failed to form tumor metastases in the livers of BALB / C nude mice within one month.

[0044] Example 2: Investigation of spleen injection modeling method

[0045] (1) Take 3-4 week old male BALB / c nude mice, each weighing (17±3)g, digest A375 cells in the logarithmic growth phase with trypsin, resuspend in pre-cooled PBS, and adjust the concentration to 1.5×10⁻⁶. 7 Prepare on ice at 10,000 / mL. Sterilize surgical instruments by autoclaving. Anesthetize nude mice that have been fasted for 12 hours with 1% pentobarbital at 50 mg / kg. After anesthesia, make a 0.5-1 cm longitudinal incision along the left side of the mouse, slightly below the rib and aligned with the ear. Gently lift the spleen with forceps, grasp the lower end of the spleen with forceps, and insert the needle along the lowest point of the spleen to a depth of one-third of a No. 5 needle. Slowly inject 50 μL of cell suspension over 1 minute. After slowly withdrawing the needle, apply pressure to the injection site with a dry alcohol swab for 2 minutes to stop bleeding. After hemostasis, suture and disinfect the incision, and place the mouse on a 37°C water-warming bag for recovery. Disinfect the incision daily with povidone-iodine for one week. Test results showed ( Figure 3 Necrotic cysts appeared inside and on the surface of the liver, but no obvious metastatic lesions were observed. The A375 melanoma splenic injection method failed to form tumor metastases in the liver of BALB / C nude mice within one month, and inflammatory necrotic areas were observed in liver sections (n=3).

[0046] (2) Following the method used in (1), A375 cells were replaced with A375-M1 cells to construct a liver metastasis model. One month later, liver sections were harvested for H&E analysis to examine liver metastasis. The results showed slight congestion of liver cells, with scattered metastatic foci appearing at the locations indicated by arrows. Figure 4 A375-M1 melanoma cells injected into the spleen resulted in tumor metastases in the liver of BALB / c nude mice within one month (n=3).

[0047] In summary, it was decided to use A375-M1 cells at a concentration of 1.5 × 10⁻⁶. 7A melanoma liver metastasis model was constructed by injecting 50 μL of the BALB / c nude mouse spleen with a concentration of 10,000 μL / mL, and the modeling time was further determined.

[0048] Example 3: Construction of A375-M1 melanoma liver metastasis model

[0049] Male BALB / c nude mice aged 3-4 weeks, each weighing 17-20g, were selected. Logarithmic-phase A375-M1 cells were trypsinized, resuspended in pre-chilled PBS, and the concentration was adjusted to 1.5 × 10⁻⁶. 7 Prepare on ice at a concentration of 10,000 mg / mL. Sterilize surgical instruments by autoclaving. Anesthetize nude mice that have been fasted for 12 hours with 1% pentobarbital at 50 mg / kg. After anesthesia, make a 0.5-1 cm longitudinal incision along the left side of the mouse, slightly below the rib and aligned with the ear. Gently lift the spleen with forceps, grasp the lower end of the spleen with forceps, and insert the needle along the lowest point of the spleen to a depth of one-third of a No. 5 needle. Slowly inject 50 μL of cell suspension over 1 minute. After slowly withdrawing the needle, apply pressure to the injection site with a dry alcohol swab for 2 minutes to stop bleeding. After hemostasis, suture and disinfect the incision, and place the mouse on a 37°C water-warming bag to await recovery. Disinfect the incision with povidone-iodine daily for one week.

[0050] The liver metastasis of nude mice was examined weekly using MRI, PET-CT, and H&E slides. A human melanoma nude mouse liver metastasis model was obtained after 1-2 weeks.

[0051] Example 4:

[0052] 1. Laboratory animals:

[0053] Blank control BALB / C nude mice and BALB / C nude mice with A375-M1 melanoma cell liver metastasis for 3 weeks.

[0054] 2. Experimental materials and equipment

[0055] Isoflurane: Hebei Jindafu Pharmaceutical Co., Ltd.; Fluoride

[18] deoxyglucose injection ( 18 F-FDG: Self-made by the Department of Nuclear Medicine, Fudan University Cancer Hospital;

[0056] Experimental equipment: BioSpec 70 / 30USR: Bruker, small animal anesthesia machine: Shenzhen Ruiwode.

[0057] 3. Experimental Methods

[0058] The liver MRI scan of the 3-week BALB / C nude model of melanoma liver metastasis constructed in this invention was performed as follows:

[0059] (1) Before the experiment, the mice were fasted for 12 hours. The blank control BALB / C nude mice and the BALB / C nude model mice with liver metastasis for 3 weeks were injected with 18F-FDG 110±10μCi, 100μL each via the tail vein.

[0060] (2) Before imaging, mice were anesthetized with oxygen mixed with 3% (volume fraction) isoflurane on a small animal anesthesia machine. During imaging, anesthesia with oxygen containing 1% (volume fraction) isoflurane was maintained.

[0061] (3) The abdomen of the mice was scanned in transverse and coronal sections using an MRI scanner. After the scan, the MRI images were transferred to the workstation for measurement.

[0062] 4. Experimental Results

[0063] MRI scans showed multiple abnormal signals in the liver of the model group, with relatively clear lesion boundaries. The liver exhibited diffuse nodular and punctate abnormal signals, with high signal intensity on T1-weighted images and low signal intensity on T2-weighted images. The signal intensity of the liver in the model group was higher than that in the control group (see...). Figure 5 ).

[0064] Example 5:

[0065] 1. Laboratory animals, materials and equipment

[0066] Experimental animals: blank control BALB / C nude mice and BALB / C nude mice with A375-M1 melanoma cell liver metastasis for 3 weeks;

[0067] Isoflurane: Hebei Jindafu Pharmaceutical Co., Ltd.; Fluoride

[18] deoxyglucose injection ( 18 F-FDG): Self-made by the Department of Nuclear Medicine, Fudan University Cancer Hospital

[0068] Experimental equipment: Siemens Inveon PET / CT, small animal anesthesia machine: Shenzhen Ruiward

[0069] 2. Experimental Methods

[0070] The liver of the A375-M1 melanoma cell liver metastasis mouse model constructed in this invention was subjected to PET-CT scan as follows:

[0071] (1) Before the experiment, the mice were fasted for 12 hours. The blank control BALB / C nude nude mice and the BALB / C nude model mice with liver metastasis for 3 weeks were injected with the tail vein. 18 F-FDG 110±10μCi, 100μL.

[0072] (2) Before imaging, mice were anesthetized with oxygen mixed with 3% (volume fraction) isoflurane on a small animal anesthesia machine. During imaging, anesthesia with oxygen containing 1% (volume fraction) isoflurane was maintained.

[0073] (3) One hour later, the Siemens Inveon PET / CT system was used to perform transverse, coronal and sagittal scans of the mouse abdomen. The data were analyzed and processed using ASIPro VM software.

[0074] 3. Experimental Results

[0075] PET-CT scans showed high-brightness signals in the transverse abdominal sections of the model group, with multiple diffusely distributed high-uptake areas in the liver (see...). Figure 6 ).

[0076] Example 6:

[0077] Experimental procedure: Liver samples were taken from control nude mice and A375-M1 melanoma liver metastasis model at 7, 14, 21, and 28 days of age, washed with PBS, weighed, and statistically analyzed.

[0078] In the control group, liver weight increased over time; in the case of metastatic tumors, liver weight increased due to tumor growth within the liver (see...). Figure 11 ).

[0079] Example 7:

[0080] Reagents: Hematoxylin staining solution: Google Bio; Eosin staining solution: Google Bio

[0081] (1) Material collection: The heart, liver, spleen, lungs and kidneys of the 4-week model group were fixed in 4% paraformaldehyde for 24 hours. The heart, liver, spleen, lungs and kidneys were removed from the fixative and the target organs were trimmed with a scalpel in a fume hood. The trimmed tissues and corresponding labels were placed in a dehydration box.

[0082] (2) Dehydration: Place the dehydration box into the basket and dehydrate it in the dehydrator by sequentially applying alcohol. 75% alcohol 4h - 85% alcohol 2h - 90% alcohol 2h - 95% alcohol 1h - anhydrous ethanol I 30min - anhydrous ethanol II 30min - benzene 5-10min - xylene I 5-10min - xylene II 5-10min - wax I 1h - wax II 1h - wax III 1h.

[0083] (3) Embedding: The paraffin-soaked organs are embedded in an embedding machine. First, the molten paraffin is placed into the embedding frame. Before the paraffin solidifies, the tissue is removed from the dehydration box and placed into the embedding frame according to the embedding surface requirements, and the corresponding label is attached. The tissue is cooled in a -20°C refrigerator. After the paraffin solidifies, the paraffin block is removed from the embedding frame and the paraffin block is trimmed.

[0084] (4) Sectioning: Place the trimmed wax block on a paraffin microtome and section it to a thickness of 4μm. Float the sections on 40℃ warm water in a slide spreader to flatten the tissue. Use a glass slide to lift the tissue and place it in a 60℃ oven to bake.

[0085] (5) Dewaxing paraffin sections to water: Place the sections in xylene I for 20 min, xylene II for 20 min, anhydrous ethanol I for 10 min, anhydrous ethanol II for 10 min, 95% ethanol for 5 min, 90% ethanol for 5 min, 80% ethanol for 5 min, 70% ethanol for 5 min, and then wash with distilled water.

[0086] (6) Hematoxylin staining of cell nuclei: Slices are stained with Harris hematoxylin for 3-8 min, washed with tap water, differentiated with 1% hydrochloric acid alcohol for a few seconds, rinsed with tap water, blued with 0.6% ammonia water, and rinsed with running water.

[0087] (7) Eosin staining of cytoplasm: Immerse the section in eosin staining solution for 1-3 min.

[0088] (8) Dehydration and mounting: Immerse the sections sequentially in 95% ethanol I for 5 min, 95% ethanol II for 5 min, anhydrous ethanol I for 5 min, anhydrous ethanol II for 5 min, xylene I for 5 min, and xylene II for 5 min to dehydrate and clear the sections. Remove the sections from the xylene and allow them to dry slightly. Mount with neutral resin (e.g., Figure 7 , 8 ).

[0089] H&E results showed that almost no tumor metastases were found in the liver of nude mice at week 1. A small number of metastases appeared at week 2, mostly concentrated near the edge of the liver lobe and intrahepatic blood vessels. After week 3, the tumor nodules gradually spread from the edge of the liver lobe and near intrahepatic blood vessels into the liver, and the area of ​​metastatic lesions gradually increased. At week 4, the number and area of ​​tumor metastases further increased, and the tumor area in the H&E sections of the nude mouse liver reached more than 70%.

[0090] Two weeks after the surgery, metastatic lesions appeared on the liver of nude mice, increasing with each week. They were mostly concentrated around the edge of the liver and around intrahepatic blood vessels. It is speculated that this is because the capillaries at the end of the liver lobe are relatively narrow, making it easier to intercept tumor cells in the blood. Furthermore, the formation of metastatic lesions around intrahepatic blood vessels is due to the fact that tumor cells spread with the blood and pass through the inner wall of the blood vessels, first accumulating around the blood vessels. This causes the metastatic lesions to spread into the liver centered on the intrahepatic blood vessels. At the same time, in situ tumors appeared on the spleen.

[0091] Example 8:

[0092] After modeling, the body weight of blank control BALB / C nude mice and A375-M1 melanoma cell liver metastasis BALB / C nude mice was measured every 2 days (e.g. Figure 9 ).

[0093] Significant differences in body weight were observed between the control group and the model group two weeks after their establishment. Liver tumor metastases severely affected the metabolism of nude mice, leading to a decrease in their body weight.

[0094] Example 9:

[0095] BALB / C nude mice (blank control) and BALB / C nude mice with A375-M1 melanoma cell liver metastasis were housed under SPF conditions, and mortality was recorded (e.g., Figure 10 ).

[0096] The model mice began to die on day 32 after modeling, and all the nude mice in the model group died by day 40.

[0097] In summary, we successfully constructed a nude mouse liver metastasis model of melanoma and systematically investigated its characteristics. We determined the method for constructing the BALB / C nude mouse liver metastasis model by injecting A375-M1 melanoma cell suspension into the spleen, and established its identification methods using in vivo MRI, PET-CT, and in vitro H&E and Bouin's assays.

[0098] The preferred embodiments of the present invention have been described in detail above, but the present invention is not limited to the embodiments described. Those skilled in the art can make various equivalent modifications or substitutions without departing from the spirit of the present invention, and these equivalent modifications or substitutions are all included within the scope defined by the claims of this application.

Claims

1. A method for constructing a melanoma liver metastasis model, characterized in that, Includes the following steps: Male BALB / c nude mice aged 3-4 weeks with a body weight of (16±4) g were selected. A375-M1 melanoma cells were cultured to the logarithmic growth phase, digested with trypsin, and the cell suspension was centrifuged and resuspended in PBS. The cells were injected into the spleen of each mouse at a concentration of 1.5 × 10⁻⁶. 7 The A375-M1 cell suspension volume was 50 μL; the A375-M1 cell line was derived from the low-metastatic melanoma cell population A375 cells, and was obtained by intravenously injecting A375 cells into nude mice to obtain the highly metastatic cell line A375-M1.

2. The method for constructing a melanoma liver metastasis model according to claim 1, characterized in that, The liver metastasis of nude mice was examined weekly using MRI, PET-CT, and H&E slides. A human melanoma nude mouse liver metastasis model was obtained after 1-2 weeks.

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