Combined medicine for resisting gastric cancer pulmonary metastasis and application thereof

Abstract

The invention relates to a combined medicine for resisting gastric cancer pulmonary metastasis and application thereof, and relates to the technical field of biological medicine. According to the combined medicine for resisting gastric cancer pulmonary metastasis, the active ingredients comprise curcumin and quercetin, LSD1 and PD-L1 protein expression of lung tissue is inhibited, reduced or inhibited, and meanwhile, cancer suppressor protein PTEN expression in the lung tissue is up-regulated, so that tumor recurrence and metastasis can be prevented and / or treated, prognosis of patients is improved, and life quality is improved.

Description

Technical Field

[0001] The present invention relates to the field of biomedical technologies, and particularly relates to a combined drug for anti-gastric cancer lung metastasis and its application. Background Art

[0002] Gastric cancer (GC) occupies an important position in the global incidence of malignant tumors. In 2022, there were approximately 960,000 new cases globally, and the number of deaths exceeded 650,000, accounting for 6.8% of global cancer deaths, making it the fifth leading cause of cancer-related deaths globally. The metastasis and recurrence of gastric cancer are the main causes of tumor-related deaths in gastric cancer patients. Whether gastric cancer metastasizes directly to the peritoneum or hematogenously to distant organs such as the liver, lungs, and bones, the overall prognosis is poor. Clinically, gastric cancer lung metastasis patients are relatively common, and the 5-year survival rate of these patients is only 2% - 4%. Therefore, how to effectively prevent and control and reduce the occurrence and progression of gastric cancer lung metastasis is a major problem to be solved at present.

[0003] Tumors can evade the surveillance and attack of the body's immune system in various ways, thereby surviving, proliferating, and metastasizing in the body, that is, tumor immune escape. The abnormal upregulation of the programmed cell death ligand-1 (PD-L1) level in cancer cells and the interaction between PD-L1 and programmed death protein 1 (PD-1) inhibit T cell function by inducing T cell exhaustion in the tumor microenvironment, ultimately promoting immune escape. And reducing the expression of PD-L1 in tumor cells through artificial intervention has become one of the important means to promote the recovery of the immune system and contain tumors.

[0004] Lysine-specific demethylase 1 (LSD1) has become a hot molecule in tumor research in recent years. It is a highly conserved FAD-dependent amine oxidase that can oxidatively remove the methyl groups of methylated H3K4 and H3K9 of the substrate, and can also remove the methyl groups on structures other than histones, thereby affecting gene expression, and is known as the "gene switch deep in the cell"; it can regulate the genes related to epithelial-mesenchymal transition (EMT) of gastric cancer cells, resulting in the disappearance of cell polarity and the reduction of cell-cell adhesion, and then enhancing the invasion ability of cancer cells. Research has found that LSD1 can regulate the transcription of the epithelial cell marker protein (E-cadherin) through the Snail / Slug transcription factor, suggesting that LSD1 may become an important therapeutic target for inhibiting EMT-mediated tumor invasion and metastasis.

[0005] Numerous studies have confirmed that the overexpression of LSD1 is associated with the development, invasion, and poor prognosis of various tumors such as breast cancer and prostate cancer. In addition, small molecule inhibitors of LSD1 show good therapeutic prospects in the treatment of hematological malignancies and solid tumors. After inhibiting the expression of LSD1, the process of epithelial-mesenchymal transition (EMT) can be blocked, thereby delaying the progression of tumors. LSD1 inhibition can increase the immunogenicity of tumor cells in the tumor microenvironment and convert "cold tumors" into "hot tumors", making tumor cells sensitive to immune checkpoint inhibitors. LSD1 inhibition can reduce the expression level of exosomal programmed cell death 1 ligand 1 (PD-L1), thus restoring the anti-tumor immune response activity of T cells, further suggesting that LSD1 also plays an important role in anti-tumor immune regulation. The tumor suppressor gene PTEN protein is a key negative regulator in the AKT signaling pathway, and PD-L1 is often regulated by the AKT pathway. LSD1 can negatively regulate the expression of PTEN. These factors are all closely related to the control of tumor development and progression.

[0006] Curcumin (Cur) is a polyphenolic substance extracted from the rhizomes of plants such as turmeric and curcuma. Quercetin (Qu) is also a flavonoid polyphenolic compound widely present in vegetables, fruits, and tea. It has a wide range of pharmacological activities and has the characteristics of weak toxicity and side effects. In recent years, the application of a variety of natural plant drugs in combination to treat tumors and regulate the body's anti-tumor immune response has become a hot topic in anti-tumor research.

[0007] Whether curcumin combined with quercetin plays an important role in anti-gastric cancer lung invasion and metastasis and its related mechanism of action have not been reported in relevant studies. Therefore, it has important clinical research significance. Summary of the Invention

[0008] To solve the above technical problems, the purpose of the present invention is to provide a combined drug for anti-gastric cancer lung metastasis and its application. The combined drug for anti-gastric cancer lung metastasis of the present invention has active ingredients including curcumin and quercetin. By inhibiting or reducing the expression of LSD1 and PD-L1 proteins in the lung tissue, and simultaneously up-regulating the expression of the tumor suppressor protein PTEN in the lung tissue, it can prevent and / or treat tumor recurrence and metastasis, thereby improving the prognosis of patients and enhancing the quality of life.

[0009] The technical solution of the present invention to solve the above technical problems is as follows:

[0010] The first object of the present invention is to provide a combined drug for anti-gastric cancer lung metastasis, and the active ingredients of the combined drug include curcumin and quercetin.

[0011] On the basis of the above technical solutions, the present invention can also be improved as follows.

[0012] Further, the curcumin and the quercetin are respectively independent dosage units, or the curcumin and the quercetin jointly form a combined dosage unit.

[0013] Further, the curcumin and the quercetin are respectively independent dosage units, and the curcumin is an oral preparation or an injection preparation; the quercetin is an oral preparation or an injection preparation.

[0014] Further, when the curcumin is an oral preparation, it can be one of a suspension preparation, a granule preparation, a tablet preparation, a capsule preparation, can also be made into a drink, or made into an extract / health food together with other Chinese herbal medicines; when the quercetin is an oral preparation, it can be one of a suspension preparation, a granule preparation, a tablet preparation, a capsule preparation, can also be made into a drink, or made into an extract / health food together with other Chinese herbal medicines.

[0015] Further, the mass ratio of the curcumin to the quercetin is 37.5 - 75:30 - 180.

[0016] The beneficial effect of adopting the above further scheme is that: in the present invention, the specific human application dose can be calculated with reference to the animal - human body surface area conversion formula. The combined use of curcumin and high - dose quercetin may significantly reduce the tumor burden and relieve the inflammatory exudation or edema in the lungs, thereby playing a role in protecting the lungs.

[0017] Further, the drug is also prepared from a pharmaceutically acceptable carrier and / or excipient.

[0018] Further, the excipient includes one or more of a filler, a binder, a disintegrant, a lubricant, a glidant, a wetting agent, an effervescent agent, a colorant, a sweetener, an aromatic, a preservative, a dispersant, a film - forming agent, a plasticizer, a pore - forming agent, a light - shielding agent, a retarder, a solvent.

[0019] Further, the curcumin and the quercetin inhibit the invasion and metastasis of tumor cells by reducing or inhibiting the expression of LSD1 and PD - L1 proteins in lung tissue and up - regulating the expression of the tumor suppressor protein PTEN in lung tissue.

[0020] The second object of the present invention is to provide an application of the combined drug for anti - gastric cancer lung metastasis, and use the combined drug for anti - gastric cancer lung metastasis in the prevention and / or treatment of tumor recurrence and / or metastasis.

[0021] Further, the combined drug for anti - gastric cancer lung metastasis is prepared from curcumin and quercetin as active ingredients, plus a pharmaceutically acceptable carrier and / or excipient, to obtain curcumin preparations and quercetin preparations for simultaneous or separate administration.

[0022] Furthermore, the tumor cells include gastric cancer cells.

[0023] The present invention's research findings show that curcumin and quercetin can weaken the migration and invasion abilities of tumor cells, thereby inhibiting the occurrence of gastric cancer lung metastasis. At the same time, the present invention discovers that curcumin and quercetin can prevent and / or treat tumor recurrence and metastasis by inhibiting or reducing the expression of LSD1 and PD-L1 proteins in lung tissue, while upregulating the expression of the tumor suppressor protein PTEN in lung tissue, thereby improving the prognosis of patients and enhancing the quality of life. The present invention provides a basis for the clinical drug treatment of patients with gastric cancer lung metastasis and has good application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 It is a diagram of establishing a gastric cancer lung metastasis model by injecting MFC murine gastric cancer cells into the tail vein of mice in Example 1 of the present invention;

[0025] Figure 2 It is a picture of the lungs of mice with MFC lung metastases in each group in Example 1 of the present invention; among them, (a) is the Control group; (b) is the CUR group; (c) is the QUE Low dose group; (d) is the QUE High dose group; (e) is the C+Q Low dose group; (f) is the C+Q High dose group;

[0026] Figure 3 It is a HE staining diagram (10×) of the lung tissues of mice with MFC lung metastases in each group in Example 1 of the present invention; among them, (a) is the Control group; (b) is the CUR group; (c) is the QUE Low dose group; (d) is the QUE High dose group; (e) is the C+Q Lowdose group; (f) is the C+Q High dose group;

[0027] Figure 4 It is a picture of LSD1 protein (immunohistochemistry, 20×) in the lung tissues of mice with MFC lung metastases in each group in Example 1 of the present invention; among them, (a) is the Control group; (b) is the CUR group; (c) is the QUE Low dose group; (d) is the QUE High dose group; (e) is the C+Q Low dose; (f) is the C+Q High dose group;

[0028] Figure 5 It is the AOD value of LSD1 protein in the lung tissues of mice with MFC lung metastases in each group in Example 1 of the present invention (where * P < 0.05; ** P < 0.01; *** P < 0.001; nsP > 0.05);

[0029] Figure 6 This is the expression of PD-L1 protein in the lung tissue of mice with MFC lung metastases in each group in Example 1 of the present invention (immunohistochemistry, 20×); where (a) is the Control group; (b) is the CUR group; (c) is the QUE Low dose group; (d) is the QUE High dose group; (e) is the C+Q Low dose; (f) is the C+Q High dose group;

[0030] Figure 7 This is the AOD value of PD-L1 protein in the lung tissue of mice with MFC lung metastases in each group in Example 1 of the present invention (where * P < 0.05; ** P < 0.01; *** P < 0.001; ns P > 0.05);

[0031] Figure 8 This is the expression of PTEN protein in the lung tissue of mice with MFC lung metastases in each group in Example 1 of the present invention (immunohistochemistry, 20×); where (a) is the Control group; (b) is the CUR group; (c) is the QUE Low dose group; (d) is the QUE High dose group; (e) is the C+Q Low dose; (f) is the C+Q High dose group;

[0032] Figure 9 This is the AOD value of PTEN protein in the lung tissue of mice with MFC lung metastases in each group in Example 1 of the present invention (where * P < 0.05; ** P < 0.01; *** P < 0.001; ns P > 0.05). Detailed implementation method

[0033] The principles and features of the present invention are described below. The examples given are only used to explain the present invention and are not intended to limit the scope of the present invention. For those not specifying specific techniques or conditions in the examples, the techniques or conditions described in the literature in the field or according to the product specifications are followed. For reagents or instruments without indicating the manufacturer, they are all conventional products that can be purchased through regular channels.

[0034] Example 1: Administration of curcumin combined with quercetin to a mouse gastric cancer lung metastasis model

[0035] I. Establishment of a mouse gastric cancer lung metastasis model.

[0036] 1. Culture of experimental cells: Mouse forestomach carcinoma (MFC) cells (Pronocyte Biosciences Inc., Wuhan, China, catalog number: CL-0156) were cultured in a cell culture incubator (Thermo Scientific, USA) at 37 °C and 5% CO2 using RPMI-1640 complete medium (Gibco, USA). The passage cycle was 2 - 3 days.

[0037] 2. Experimental animals: SPF-grade KM mice (Jiangsu Huachuang Xinnuo Medical Technology Co., Ltd.).

[0038] 3. Establishment of a mouse model of MFC gastric cancer cell lung metastasis

[0039] (1) Collect MFC cells in the logarithmic growth phase. After centrifugation, discard the supernatant, and resuspend the cells in PBS to form a single-cell suspension. Adjust the cell density to 1×10 7 cells / mL.

[0040] (2) Fix SPF-grade KM mice (Jiangsu Huachuang Xinnuo Medical Technology Co., Ltd., production license number: SCXK (Su) 2020-000) on a mouse intravenous visible tail injection fixator (Beijing Chuangbo Global Biotechnology Co., Ltd., China). Disinfect the mouse tail with alcohol to fully expose the vein, and inject 0.2 ml of the single-cell suspension per mouse via the tail vein. Start the experiment on the 3rd day after modeling. Observe the general condition of the mice every day, measure and record the body weight change every 2 days. After 7 days, a mouse model of MFC gastric cancer cell lung metastasis is obtained. The construction process of the mouse model of MFC gastric cancer cell lung metastasis is as Figure 1 shown.

[0041] II. Animal grouping and drug intervention

[0042] 1. Grouping

[0043] The mice in the model constructed in step one were randomly divided into a model control group (Control), a curcumin group (CUR), a low-dose quercetin group (QUE Low dose), a high-dose quercetin group (QUE High dose), a curcumin combined with low-dose quercetin group (C+Q Low dose), and a curcumin combined with high-dose quercetin group (C+Q High dose), with 6 mice in each group.

[0044] 2. Drug administration intervention

[0045] Curcumin (Dalian Meilun Biotechnology Co., Ltd., China) and quercetin (MCE, USA) were stored in the dark. Before each administration, they were dissolved and diluted with DMSO to prepare curcumin drug solution with a concentration of 50 mM and quercetin drug solution with a concentration of 50 mM for standby.

[0046] (1) Curcumin group: Mice were intragastrically administered with curcumin solution at a dosage of 37.5 mg / kg once a day for 14 days of intervention.

[0047] (2) Low-dose quercetin group: Mice were intragastrically administered with quercetin solution at a dosage of 30 mg / kg once a day for 14 days of intervention.

[0048] (3) High-dose quercetin group: Mice were intragastrically administered with quercetin solution at a dosage of 90 mg / kg once a day for 14 days of intervention.

[0049] (4) Curcumin combined with low-dose quercetin group: Mice were intragastrically administered with curcumin solution at a dosage of 37.5 mg / kg once a day for 14 days of intervention; meanwhile, mice were intragastrically administered with quercetin solution at a dosage of 90 mg / kg once a day for 14 days of intervention.

[0050] (5) Curcumin combined with high-dose quercetin group: Mice were intragastrically administered with curcumin solution at a dosage of 37.5 mg / kg once a day for 14 days of intervention; meanwhile, mice were intragastrically administered with quercetin solution at a dosage of 90 mg / kg once a day for 14 days of intervention.

[0051] (6) Model control group: Mice were intragastrically administered with 0.9% sodium chloride solution at the same volume as that of the curcumin combined with high-dose quercetin group once a day for 14 days of intervention.

[0052] III. Detection

[0053] 1. Calculate the inhibition rate of lung metastases

[0054] After the last drug intervention on the 14th day for the mice in groups (1) - (6), the mice were anesthetized and sacrificed, and the lungs were completely removed. Observe and count the number of metastatic nodules on the lung surface of each group of mice, convert it into the total number of metastatic nodules on the lung surface using the formula, and then calculate the inhibition rate of lung metastases for each group of mice. The criteria for judging lung nodules and the calculation formula for the inhibition rate of lung metastases are as follows:

[0055] Criteria for judging lung nodules: According to the diameter of lung nodules, they are divided into 4 grades: Grade I < 0.5 mm, 0.5 mm ≤ Grade II < 1 mm, 1 mm ≤ Grade III < 2 mm, Grade IV ≥ 2 mm. The total number of metastatic nodules on the lung surface = the number of Grade I nodules × 1 + the number of Grade II nodules × 2 + the number of Grade III nodules × 3 + the number of Grade IV nodules × 4.

[0056] Calculation formula for the inhibition rate of lung metastases in each group: Inhibition rate of lung metastases (%) = (average number of metastatic nodules on the lung surface in the model control group - average number of nodules on the lung surface in the experimental group) / average number of metastatic nodules on the lung surface in the model control group × 100%.

[0057] The results are shown in Table 1.Figure 2 As shown below:

[0058] Table 1

[0059]

[0060]

[0061] From Table 1, Figure 2 it can be obtained that: in terms of the total number of metastatic nodules on the lung surface: compared with the control group, the number of lung nodules in the medium-dose C+Q group and the high-dose C+Q group was significantly reduced (P < 0.05); compared with the CUR group, the low-dose QUE group, and the high-dose QUE group, the high-dose C+Q group was significantly reduced (P < 0.05). In terms of the inhibition rate of lung metastases: the lung metastasis inhibition rate of each combination group was higher than that of the single-drug group, and the combination of the two drugs was better than the single-drug effect. In particular, the high-dose C+Q group had the maximum lung metastasis inhibition rate, reaching 94.8% ± 5.54% (see Table 1, P < 0.05 when compared with each group: compared with the control group, * P < 0.05; compared with the CUR group, # P < 0.05; compared with the low-dose QUE group, △ P < 0.05; compared with the high-dose QUE group, ○ P < 0.05).

[0062] 2. HE staining

[0063] (1) The lung tissue was fixed with 10% formaldehyde routinely, dehydrated, and embedded to obtain the fixed lung tissue. Subsequently, the fixed lung tissue was taken out from the 10% formalin solution and rinsed slowly in running water, and then placed successively in alcohol with concentrations of 75%, 85%, 95%, and 100% for 5 minutes of dehydration each time. Then it was soaked successively in xylene solutions I, II, and III (xylene solutions I, II, and III are exactly the same, with a concentration of 100% each) for 5 minutes of soaking each time. Finally, the lung tissue was embedded in paraffin and sectioned.

[0064] (2) The sections were baked in an oven at 60°C for 1 hour, then soaked successively in xylene solutions I, II, and III (xylene solutions I, II, and III are exactly the same, with a concentration of 100% each) for 5 minutes of soaking each time, and then placed successively in alcohol with concentrations of 100%, 95%, 85%, and 75% for 5 minutes of soaking each time. Finally, it was washed with double-distilled water for 2 minutes.

[0065] (3) An appropriate amount of hematoxylin was dropped on the sections, waited for 5 minutes and then rinsed slowly in running water, soaked in 1% hydrochloric acid alcohol, and finally rinsed slowly in running water for 10 minutes.

[0066] (4) Add an appropriate amount of eosin staining solution to the section. After waiting for 2 minutes, take out the section and shake off the staining solution, then rinse it slowly in running water for 10 minutes.

[0067] (5) Place the stained section successively in alcohol with concentrations of 75%, 85%, 95%, and 100%, soak for about 5 minutes each time, and then soak successively in xylene solutions I, II, and III (xylene solutions I, II, and III are exactly the same, with a concentration of 100% each), soak for 5 minutes each time. Finally, take out the section and place it in a cool and dry place to air dry and mount the slide.

[0068] (6) Observe the tissue section under a microscope, and use a slide scanning imaging system (Shenzhen Shengqiang Technology Co., Ltd., China) to collect images and analyze them.

[0069] The results are as Figure 3 shown:

[0070] It can be seen from Figure 3 that HE staining makes the cell nucleus blue and the cytoplasm red. The formation of tumors in each group was observed and compared under a microscope, and the results further confirmed that the combined drug group had a better effect on inhibiting the formation of gastric cancer lung metastases.

[0071] 3. Immunohistochemical staining

[0072] The expression of LSD1, PD-L1, and PTEN proteins in the lung tissues of mice in each group was detected by immunohistochemistry. Use DAB staining solution (Titan Super) (Fuzhou Maixin Co., Ltd.; TT0805), and detect according to the instructions of the kit. The specific steps are as follows:

[0073] (1) Fix the lung tissue routinely with 10% formaldehyde, dehydrate, and embed to obtain the fixed lung tissue; take out the fixed lung tissue from the 10% formalin solution, rinse it slowly in running water, and place it successively in alcohol with concentrations of 75%, 85%, 95%, and 100%, dehydrate for 5 minutes each time, and then soak successively in xylene solutions I, II, and III for 5 minutes each time. Finally, embed the lung tissue with paraffin and section it;

[0074] (2) After baking the section in an oven at 60°C for 1 hour, take it out and add the dewaxing solution, then continue to bake it in an oven at 60°C (Shanghai Hesheng Instrument Technology Co., Ltd., China) for 10 minutes to dewax, soak it in gradient alcohol, take it out, wash the section with double-distilled water, and then wash it with PBS solution;

[0075] (3) EDTA Antigen Retrieval Solution (Fuzhou Maixin Co., China, prepared by mixing antigen retrieval buffer and purified water at a ratio of 1:50). After boiling, place the sections into it, cover with a lid, and continue boiling at 100 °C for 20 min. Turn off the heat and let it stand and cool to room temperature. Open the lid, take out the sections, and wash the sections with double-distilled water for 10 min;

[0076] (4) Use an I.S.CIRCLE WRITER marker to draw a circle, then add 2 drops of endogenous peroxidase blocking solution (Fuzhou Maixin Co., China) onto the sections, let it stand at room temperature in a humid chamber for 10 min, and wash with PBS solution 3 times, 5 min each time;

[0077] (5) Take primary antibodies prepared at different dilution ratios (Anti-LSD1 antibody, Abcam, USA, clone number: EPR6825, diluted 1:100; Anti-PD-L1 antibody, CST, USA, catalog number: 64988T, diluted 1:200; recombinant Anti-PTEN antibody (Abcam, USA, clone number: EPR9941-2, diluted 1:200)), add them dropwise onto the sections, place them in a humid chamber and incubate overnight in a 4 °C refrigerator, then wash with PBS solution 3 times, 5 min each time;

[0078] (6) Add 30 μl of reaction enhancer onto the sections, ensuring complete coverage of the specimens; incubate at room temperature in a humid chamber for 10 min, wash with PBS solution 3 times, 5 min each time; use a pipette to add 30 μl of secondary antibody (high-sensitivity enzyme-labeled anti-mouse / rabbit IgG polymer, Fuzhou Maixin Biotechnology Development Co., China) onto the sections; then place the sections in a humid chamber and incubate at room temperature for 1 h, wash with PBS solution 3 times, 5 min each time;

[0079] (7) Prepare DAB chromogenic solution. After the last wash of the sections, add 40 μl of the mixed DAB chromogenic solution, stain for about 1 - 3 min, observe under a microscope, and then rinse with double-distilled water for 5 min;

[0080] (8) Place the sections into the hematoxylin staining jar, submerging the tissue sections. The counterstaining time should not exceed 10 s. Then wash with double-distilled water 3 times, 5 min each time, and then place in 1% hydrochloric acid alcohol differentiation solution for several seconds;

[0081] (9) Place the sections successively into alcohol with volume concentrations of 85%, 95%, and 100%, soak for about 5 min each time, mount with neutral resin, and air-dry at room temperature;

[0082] (10) Observe the tissue sections under a microscope, use a slide scanning imaging system to collect images and perform preliminary analysis;

[0083] (11) Immunohistochemical result analysis: Under the microscope, 3 fields of view were randomly selected in each group to observe the expression of PD-L1, PTEN, and LSD1 proteins in the tissue. At the same time, Image J software was used to measure the integrated optical density (IOD) and the area of the region where the target protein was distributed (Area), and the average optical density (AOD) was calculated, where AOD = IOD / Area.

[0084] The judgment criteria for positive protein expression are as follows:

[0085] a. Judgment criteria for positive LSD1 protein expression: Light yellow to brown-yellow granules appear in the cell nuclei or cytoplasm of tissue cells;

[0086] b. Judgment criteria for positive PD-L1 protein expression: Light yellow to brown-yellow granules appear in the cytoplasm or cell membrane of tissue cells;

[0087] c. Judgment criteria for positive PTEN protein expression: Light yellow to brown-yellow granules appear in the cytoplasm of tissue cells.

[0088] The results are as Figures 4 to 9 shown:

[0089] The immunohistochemical staining of LSD1 protein in the lung tissues of mice with MFC lung metastases in each group is as Figure 4 、 5 shown; the immunohistochemical staining of PD-L1 protein in the lung tissues of mice with MFC lung metastases in each group is as Figure 6 、 7 shown; the immunohistochemical staining of PTEN protein in the lung tissues of mice with MFC lung metastases in each group is as Figure 8 、 9 shown;

[0090] As Figures 4 to 9 shown: Compared with the control group, single drug treatment could down-regulate the expression of LSD1 and PD-L1 proteins and up-regulate the expression of PTEN. The change trend of the above proteins in the combined drug treatment group was more obvious. The differences in the expression of the above proteins between the high-dose C+Q group and the control group and the single drug treatment group were significant (P < 0.05), suggesting that the combination of curcumin and quercetin against lung metastases of gastric cancer may be related to down-regulating LSD1 and PD-L1 and up-regulating PTEN protein expression.

[0091] 4. Lung index

[0092] On the 14th day after the end of the last drug intervention, the body weights of the mice were measured. Subsequently, the mice were anesthetized and sacrificed, and each group of mice was dissected. The lung weights were measured using an electronic balance. The lung index (%) was calculated using the formula: lung index (%) = wet lung weight (mg) × 100 / body weight (g) × 100%.

[0093] The lung weight of the control group mice was 396.00 ± 162.20 mg, and the lung weight of the high-dose C+Q group was 206.30 ± 16.37 mg, which was significantly lower than that of the control group (P < 0.05). The lung index of the control group was 14.42 ± 8.59 mg / g, and the lung index of the high-dose C+Q group was 6.15 ± 0.59 mg / g, with a significant difference between the two groups (P < 0.05). This suggests that the combined use of curcumin and high-dose quercetin can significantly reduce the tumor burden in mice, alleviate inflammatory exudation or edema in the lungs, and thus produce a protective effect on the lungs.

[0094] 5. General status and body weight changes of mice in each experimental group

[0095] During the administration intervention, the general status (appetite, skin and hair color, movement sensitivity, defecation) of each group of mice was also observed. After the end of the last drug intervention on the 14th day, the body weights of each group of mice were measured, as shown in Table 2.

[0096] Table 2: Final body weights (g) of mice with MFC lung metastases in each group (n = 6, )

[0097]

[0098]

[0099] The results are as follows:

[0100] (1) Before the experimental intervention, the body weights of each group were between 20.25 - 25.63 g, and there was no significant difference between groups (P < 0.05). There were no obvious differences in the general conditions of each group of mice in the early stage of the experiment. However, in the later stage of the experiment (days 8 - 14), the growth of each group of mice was different. The mice in the model control group showed obvious listlessness, slow movement, poor vitality, and dull fur. The mice in the CUR group, low-dose QUE group, and high-dose QUE group had better spirits, normal eating and defecation, and basically normal activities. However, it was observed that one mouse in the low-dose QUE group and one mouse in the high-dose QUE group had poor general conditions, similar to those of the control group mice. The mice in the low-dose C+Q group and high-dose C+Q group had good spirits, shiny fur, sensitive movement, and normal eating and defecation, especially the mice in the high-dose C+Q group.

[0101] (2) The final body weight results are shown in Table 2. Among them, the body weight of the model control group was between (22.83 - 34.64) g, with the mean ± standard deviation being (29.73 ± 4.68) g; the mean ± standard deviation of the body weight of the CUR group was (28.43 ± 3.40) g; the mean ± standard deviation of the body weight of the low-dose QUE group was (30.42 ± 2.87) g; the mean ± standard deviation of the body weight of the high-dose QUE group was (32.11 ± 2.95) g; the mean ± standard deviation of the body weight of the low-dose C+Q group was (32.54 ± 1.97) g; the mean ± standard deviation of the body weight of the high-dose C+Q group was (33.59 ± 0.90) g. Compared with the CUR group, the body weight of the high-dose C+Q group increased significantly, and the difference was statistically significant (P < 0.05); there was no statistically significant difference in body weight compared with the other groups (P > 0.05). It is suggested that the mice in the high-dose C+Q group had the best general condition, and no obvious manifestations of drug toxicity and side effects were observed.

[0102] In summary, the anti-gastric cancer lung metastasis combination drug of the present invention has active ingredients including curcumin and quercetin. By inhibiting or reducing the expression of LSD1 and PD-L1 proteins in the lung tissue, and simultaneously upregulating the expression of the tumor suppressor protein PTEN in the lung tissue, it can prevent and / or treat the recurrence of gastric cancer and the invasion and metastasis of gastric cancer cells, especially lung metastasis, thereby improving the prognosis of patients and enhancing the quality of life.

[0103] Although the embodiments of the present invention have been shown and described above, it can be understood that the above embodiments are exemplary and should not be construed as limiting the present invention. Those of ordinary skill in the art can make changes, modifications, substitutions, and variations to the above embodiments within the scope of the present invention.

Claims

1. A combined drug for anti-gastric cancer lung metastasis, characterized in that, The active ingredients of the combined drug include curcumin and quercetin.

2. The combined drug for resisting gastric cancer lung metastasis according to claim 1, characterized in that, The curcumin and the quercetin are each an independent dosage unit, or the curcumin and the quercetin together form a combined dosage unit.

3. The combined drug for anti-gastric cancer lung metastasis according to claim 2, wherein The curcumin and the quercetin are each an independent dosage unit, and the curcumin is an oral preparation or an injection preparation, and the quercetin is an oral preparation or an injection preparation.

4. The combined drug for resisting gastric cancer lung metastasis according to claim 1, wherein The mass ratio of the curcumin to the quercetin is 37.5-75:30-180.

5. The combined drug for resisting gastric cancer lung metastasis according to claim 1, wherein, The drug is also prepared from pharmaceutically acceptable carriers and / or excipients.

6. The combined drug for resisting gastric cancer lung metastasis according to claim 5, wherein, The excipients include one or more of fillers, binders, disintegrants, lubricants, glidants, wetting agents, effervescent agents, colorants, sweeteners, fragrances, preservatives, dispersants, film formers, plasticizers, pore-forming agents, light blockers, blockers, solvents.

7. The combined drug for resisting gastric cancer lung metastasis according to claim 5, characterized in that, The curcumin and the quercetin inhibit the invasion and metastasis of tumor cells by reducing or inhibiting the protein expression of LSD1 and PD-L1 in lung tissue and upregulating the expression of the tumor suppressor protein PTEN in lung tissue.

8. Use of a combined drug for resisting gastric cancer lung metastasis, characterized in that, The combined drug for preventing and treating gastric cancer lung metastasis according to any one of claims 1 to 7 is used for preventing and / or treating tumor recurrence and / or metastasis.

9. Use of a combined drug for anti-gastric cancer lung metastasis according to claim 8, characterized in that, The combined drug for preventing and treating gastric cancer lung metastasis is prepared from curcumin and quercetin as active ingredients, plus pharmaceutically acceptable carriers and / or excipients, to prepare curcumin preparations and quercetin preparations for simultaneous or separate administration.

10. Use of a combined drug for resisting gastric cancer lung metastasis according to claim 8, characterized in that, The tumor cells include one of gastric cancer cells.

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