Application of simvastatin in preparing medicine for preventing or treating liver cancer

Abstract

The present invention belongs to the field of medicine, and specifically relates to the use of simvastatin in the preparation of a drug for preventing or treating liver cancer. The results of pharmacological experiments show that simvastatin can inhibit the proliferation, migration and invasion of liver cancer cells HepG2, and promote apoptosis of liver cancer cells. In addition, the drug also contains a second active ingredient, which is an immune checkpoint inhibitor. The results of pharmacological experiments show that the drug containing the second active ingredient significantly inhibits the growth of human liver cancer Hep3B nude mouse transplanted tumors, and the effect is significantly better than the same dose of simvastatin and immune checkpoint inhibitors alone, which shows that the drug has a significant synergistic effect after combined use, and has good prospects for clinical application.

Description

Technical Field

[0001] The present invention belongs to the field of medicine, and specifically relates to the use of simvastatin in the preparation of a drug for preventing or treating liver cancer. The present invention also relates to the use of a combination of simvastatin and an immune checkpoint inhibitor in the preparation of a drug for preventing or treating liver cancer. Background Art

[0002] Hepatocellular carcinoma (HCC) ranks sixth in the incidence of malignant tumors worldwide, and more than half of the new cases and deaths of liver cancer each year occur in China. The latest statistics show that the incidence of liver cancer in my country ranks fourth among malignant tumors, and the mortality rate ranks second among malignant tumors. At present, traditional treatments for liver cancer mainly rely on surgical resection, liver transplantation, radiofrequency thermal therapy, and hepatic artery embolization chemotherapy. However, the vast majority of HCC patients have lost the opportunity for surgical resection when diagnosed, and the severe shortage of liver donors makes liver transplantation difficult to implement. Retrospective clinical investigations and studies have shown that various adjuvant treatments have not shown the effect of significantly prolonging the survival of HCC patients. Therefore, it is urgent to seek new ways and drugs to treat liver cancer, which is of great significance for achieving a healthy China.

[0003] Liver cancer patients are not sensitive to traditional chemotherapy drugs, but studies have shown that abnormal lipid metabolism is closely related to the occurrence and development of liver cancer. Compared with normal liver cells, liver cancer cells have significantly increased fatty acid and cholesterol synthesis, indicating that there is abnormal lipid metabolism in liver cancer. Therefore, it is believed that finding drugs that can inhibit abnormal lipid metabolism in liver cancer cells is expected to become a new strategy for liver cancer treatment.

[0004] Simvastatin (SVT) is a widely used lipid-lowering drug, belonging to the class of hydroxymethylglutaryl coenzyme A (HMG-CoA) reductase inhibitors, and is commonly used in the prevention and treatment of cardiovascular diseases in clinical practice. A large number of retrospective clinical investigations have found that statins can reduce the risk of liver cancer in high-risk groups (diabetes and liver cirrhosis, etc.) after adjusting other potential risk factors, significantly reduce the risk of chronic hepatitis B and C patients progressing to liver cancer, and reduce the recurrence of liver cancer in liver transplant patients. Based on the above literature support, the inventors speculate that simvastatin may reduce the occurrence of liver cancer, slow down the progression of liver cancer, and reduce the mortality rate of liver cancer through a series of mechanisms in addition to the known lipid-lowering effect.

[0005] In recent years, immune checkpoint inhibitors have achieved great breakthroughs in cancer immunotherapy and are quietly changing the overall layout of traditional cancer treatment methods. However, the use of cancer immunotherapy for the treatment of liver cancer is still in its infancy. The main reason is that many immunotherapies have poor therapeutic effects on liver cancer, which may be related to the fact that the immunosuppressive microenvironment of liver cancer hinders the effect of existing immunotherapy. In addition, clinical studies and clinical applications have found that despite the powerful efficacy, immune checkpoint inhibitors are prone to more serious adverse reactions in the liver, and even develop drug resistance in some patients. The above-mentioned shortcomings of immune checkpoint inhibitors seem to be overcome by combining drugs, such as overcoming tumor resistance, producing synergistic effects, and reducing toxic side effects. However, how to select a combination from a large number of drugs that can be used in combination with immune checkpoint inhibitors to produce synergistic effects and reduce their toxicity in the treatment of liver cancer has become a new challenge faced by scientists in the field of drug research and development.

[0006] There are currently no studies on the use of simvastatin for the prevention or treatment of liver cancer or on the combination of simvastatin and immune checkpoint inhibitors for the prevention or treatment of liver cancer. Summary of the invention

[0007] In order to solve the problem of insufficient clinically available drugs that can effectively treat liver cancer, the inventors have discovered that simvastatin can be used to prevent or treat liver cancer. In addition, the inventors have also discovered that the combination of simvastatin and immune checkpoint inhibitors has a synergistic effect in preventing or treating liver cancer. The above findings will provide a new idea for the safe, effective and economical clinical treatment of liver cancer.

[0008] Specifically, the technical solution to achieve the purpose of the present invention is as follows:

[0009] The present invention provides application of simvastatin in preparing medicine for preventing or treating liver cancer.

[0010] As an alternative, in the above use, the liver cancer is hepatocellular carcinoma (HCC).

[0011] As an optional mode, in the above-mentioned use, the hepatocellular carcinoma is caused by one or more diseases selected from the following: chronic hepatitis virus infection (such as hepatitis B virus infection, hepatitis C virus infection or hepatitis E virus infection), long-term exposure to carcinogens (such as aflatoxin, etc.), excessive drinking, non-alcoholic fatty liver disease, hemochromatosis or alpha-1 antitrypsin deficiency.

[0012] As an optional mode, in the above use, the simvastatin inhibits the proliferation, migration and invasion of liver cancer cells, and promotes the apoptosis of liver cancer cells.

[0013] As an option, in the above use, the medicament further comprises a second active ingredient.

[0014] Preferably, the second active ingredient is an immune checkpoint inhibitor.

[0015] As an optional method, in the above-mentioned use, the immune checkpoint inhibitor is selected from one or more of the following: PD-1 inhibitor, CTLA-4 inhibitor, LAG-3 inhibitor, TIM-3 inhibitor, BTLA inhibitor, PD-L1 inhibitor, PD-L2 inhibitor, B7-1 inhibitor, B7-2 inhibitor, galectin 9 inhibitor or HVEM inhibitor.

[0016] The immune checkpoint inhibitor is a small molecule compound or a biological product, preferably an antibody.

[0017] As an optional mode, in the above-mentioned use, the immune checkpoint inhibitor is selected from PD-1 inhibitor or PD-L1 inhibitor.

[0018] Preferably, the immune checkpoint inhibitor is selected from one or more of ipilimumab, nivolumab, pembrolizumab, atezolizumab, avelumab, and durvalumab.

[0019] As an optional mode, in the above-mentioned use, in the drug, the simvastatin and the immune checkpoint inhibitor show a synergistic effect in preventing or treating liver cancer.

[0020] As an optional manner, in the above-mentioned use, in the drug, the weight ratio of the simvastatin to the immune checkpoint inhibitor is 1:0.1-1.

[0021] Preferably, the weight ratio of the simvastatin to the immune checkpoint inhibitor is 1:0.5.

[0022] As an optional mode, in the above-mentioned use, in the medicine, the simvastatin and the immune checkpoint are administered simultaneously or separately in sequence.

[0023] Compared with the prior art, the present invention has the following beneficial effects:

[0024] The inventors have discovered for the first time that simvastatin can be used to prevent or treat liver cancer, which is a secondary development and utilization of a drug that has been marketed. Pharmacological experimental results show that simvastatin can inhibit the proliferation, migration and invasion of liver cancer cells HepG2, and promote apoptosis of liver cancer cells. Simvastatin is a widely used lipid-lowering drug in clinical practice. This drug has been on the market for many years and has accumulated a large amount of safety data, so that the safety of using simvastatin to prevent or treat liver cancer can be fully guaranteed.

[0025] In addition, the inventors have also found that the combination of simvastatin and immune checkpoint inhibitors has a synergistic effect in preventing or treating liver cancer, thereby making it possible to increase the effectiveness and safety of such drugs by reducing the dosage of immune checkpoint inhibitors.

[0026] The above findings will provide a new idea for the safe, effective and economical clinical treatment of liver cancer. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] The accompanying drawings are used to provide a further understanding of the present invention and constitute a part of the specification. Together with the embodiments of the present invention, they are used to explain the present invention and do not constitute a limitation of the present invention. In the accompanying drawings:

[0028] Figure 1 : Effects of simvastatin on the growth activity of HepG2 hepatoma cells.

[0029] Figure 2 :The effect of simvastatin on the migration ability of liver cancer cells HepG2. Figure A is a representative microscope photo of the cell scratch test, with a magnification of 40x, and Figure B is the quantitative analysis result of the effect of simvastatin on the migration ability of HepG2 cells.

[0030] Figure 3 :The effect of simvastatin on the invasion ability of liver cancer cells HepG2. Figure A is a representative microscope photo of the Transwell chamber experiment, with a magnification of 200x, and Figure B is the quantitative analysis result of the effect of simvastatin on the invasion ability of HepG2.

[0031] Figure 4 :Effects of simvastatin on apoptosis of liver cancer cells HepG2. Figure A is a representative Western blot, and Figure B is a quantitative analysis of the effects of simvastatin on the expression of anti-apoptotic protein Bcl-2 and pro-apoptotic protein Bax in liver cancer cells HepG2. DETAILED DESCRIPTION

[0032] The inventors, in their in-depth study of the relationship between lipid metabolism mechanisms and liver cancer, have discovered for the first time through extensive screening that simvastatin can be used to prevent or treat liver cancer. In addition, the inventors unexpectedly discovered during the research that the combination of simvastatin and immune checkpoint inhibitors has a significant synergistic effect in preventing or treating liver cancer. The present invention was completed on this basis.

[0033] In order to facilitate the understanding of those skilled in the art, the main active ingredients involved in the present invention are described below.

[0034] As used herein, the term "immune checkpoint inhibitor" refers to a type of drug that blocks certain proteins produced by certain types of immune cells, such as T cells and some cancer cells. These proteins help to keep the immune response in check and keep T cells from killing cancer cells. When these proteins are blocked, the brake signal on the immune system is released, and T cells are better able to kill cancer cells. Examples of immune checkpoint inhibitors include, but are not limited to, PD-1 inhibitors, CTLA-4 inhibitors, LAG-3 inhibitors, TIM-3 inhibitors, BTLA inhibitors, PD-L1 inhibitors, PD-L2 inhibitors, B7-1 inhibitors, B7-2 inhibitors, galectin 9 inhibitors, and HVEM inhibitors. Immune checkpoint inhibitors can be small molecules or biologics, preferably antibodies.

[0035] As used herein, the term "PD-1 inhibitor" refers to an antibody or other molecule that inhibits the function of programmed death protein 1 (PD1). Exemplary inhibitors / antibodies include, but are not limited to, antibodies listed in U.S. Pat. Nos. 7,029,674, 7,488,802, 7,521,051, 8,008,449, 8,354,509, 8,617,546, and 8,709,417.

[0036] As used herein, the term "PD-L1 inhibitor" refers to an antibody or other molecule that inhibits the function of programmed death ligand 1 (PDL1). Exemplary antibodies include, but are not limited to, those listed in U.S. Patent Nos. 8,217,149, 8,383,796, 8,552,154, and 8,617,546.

[0037] Preferably, the immune checkpoint inhibitor is selected from one or more of ipilimumab, nivolumab, pembrolizumab, atezolizumab, avelumab, and durvalumab.

[0038] In the medical uses described above, the administration time, number of administrations and frequency of administration of the drug of the present invention need to be determined according to the specific diagnosis results of the disease, which is within the technical scope mastered by those skilled in the art.

[0039] The treatment plan for mice can be applied to humans, and the effective doses of all drugs for humans can be converted by the effective doses of the drugs for mice, which is also easy to achieve for ordinary technicians in this field.

[0040] The present invention will be further described with reference to specific embodiments. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not intended to limit the scope of the present invention.

[0041] If no specific techniques or conditions are specified in the examples, the techniques or conditions described in the literature in the field or the product instructions are used. If no manufacturer is specified for the reagents or instruments used, they are all conventional products that can be purchased through regular channels.

[0042] The experimental methods in the following examples are conventional methods unless otherwise specified. The experimental materials used in the following examples are commercially available products unless otherwise specified.

[0043] Example 1: Effect of simvastatin on human hepatoma cells HepG2 cultured in vitro

[0044] 1. Main experimental materials

[0045] Human hepatocellular carcinoma cell line HepG2 was purchased from Shanghai Cell Bank, Chinese Academy of Sciences; simvastatin standard was purchased from MCE (purity above 99.5%); CCK-8 staining solution was purchased from Beijing Bio-Technology Co., Ltd.; Transwell chamber was purchased from Corning Incorporated, USA; Bcl-2, Bax and β-Actin monoclonal antibodies and HRP-labeled secondary antibodies were purchased from Cell signaling Technology, USA.

[0046] 2. Experimental Methods

[0047] 2.1 Culture of human hepatoma cell line HepG2

[0048] The human liver cancer cell line HepG2 was cultured in DMEM medium containing 10% fetal bovine serum, 1% penicillin and streptomycin in a 37°C, 5% CO2 incubator. When the cells grew to a confluence of 90%, they were passaged for subsequent experiments.

[0049] 2.2 CCK-8 assay to detect changes in cell viability

[0050] (1) Group by concentration (e.g. Figure 1 As shown, HepG2 cells were inoculated in 96-well plates with simvastatin (0, 1, 5, 10, 20, 40 and 80 μM, respectively). Each group had 5-6 replicate wells, 100 μl per well, and 1x10 4 cells;

[0051] (2) Place the 96-well plate seeded with cells in a 37°C, 5% CO2 incubator overnight;

[0052] (3) After the cells adhered to the wall, the old culture medium was aspirated, and 100 μl of simvastatin of different concentrations prepared in culture medium containing 1% serum was added to each well according to the grouping;

[0053] (4) Continue culturing in the incubator for 24 h;

[0054] (5) Add 10 μl of CCK8 solution to each well;

[0055] (6) Cultivate in an incubator for 4 h;

[0056] (7) Measure the absorbance (OD) value at 450 nm using an enzyme-labeled instrument.

[0057] 2.3 Cell scratch assay to detect changes in migration ability

[0058] (1) Place all necessary instruments in a clean bench for UV sterilization for 30 minutes;

[0059] (2) Use a marker pen to draw even horizontal lines on the back of the 6-well plate, using a ruler to draw lines approximately every 0.5-1 cm. Make at least 5 lines on each well.

[0060] (3) Digest the cells and inoculate them into 6-well plates, with approximately 5 x 10 cells per well. 5 Place the cells in an incubator overnight to allow them to adhere to the wall;

[0061] (4) The next day, after the cells have attached to the wall, use the tip of the gun to measure against the ruler and try to make a scratch perpendicular to the horizontal line on the back. The gun tip should be vertical, not tilted.

[0062] (5) Wash with PBS three times, remove the scratched cells, and add serum-free culture medium and simvastatin (20 μM) according to the group;

[0063] (6) Place the cells in an incubator and continue culturing. Take samples at 0, 12, 24, 48, and 96 hours and take photos. 2.4 Transwell chamber assay to detect changes in cell invasion ability

[0064] (1) Starve the HepG2 cells for 12-24 h in advance to eliminate the effect of serum;

[0065] (2) Coat the upper chamber surface of the bottom membrane of the Transwell chamber with 50 mg / L Matrigel diluted 1:8 and place in an incubator to air dry;

[0066] (3) Digest the cells, centrifuge to remove the culture medium after digestion, resuspend the cells in serum-free culture medium, and adjust the cell concentration to 5x10 5 / ml;

[0067] (4) 100 μl of the cell suspension was added to the upper chamber of the Transwell and divided into groups according to the experimental requirements. Simvastatin (20 μM) was added to the experimental group.

[0068] (5) Add 600 μl of culture medium containing 10% serum to the lower chamber;

[0069] (6) Cultivate in an incubator for 24 hours;

[0070] (7) After the incubation time is over, the chamber is removed, the upper chamber liquid is aspirated, and the chamber is transferred to a well containing approximately 800 μl of pre-cooled methanol and fixed at room temperature for 30 min;

[0071] (8) After the fixation time is over, aspirate the fixative solution in the upper chamber and transfer it to the well pre-added with about 800 μl of crystal violet, and stain at room temperature for 15-30 min;

[0072] (9) Gently soak and wash several times with PBS, remove the chamber, aspirate the liquid in the upper chamber, and carefully wipe the unpenetrated cells on the bottom membrane surface of the upper chamber with a wet cotton swab;

[0073] (10) Place the chamber on a glass slide and observe nine random fields of view under a microscope, and then count the results. 2.5 Western blot detection of changes in apoptosis-related proteins in HepG2 cells

[0074] (1) Aspirate the culture medium, wash the cells three times with pre-cooled PBS, scrape the cells off, collect them into a centrifuge tube, centrifuge at 2000 rpm for 5 min, and transfer the washed cells into a clean EP tube;

[0075] (2) Prepare cell lysis buffer: add 10 μl of phosphatase inhibitor, 1 μl of protease inhibitor and 10 μl of PMSF to 1 ml of cell lysis buffer, mix well and store on ice;

[0076] (3) Count the washed cells and calculate the number of cells per 10 6 Add cell lysis buffer at a ratio of 100 μl lysis buffer per cell;

[0077] (4) After adding lysis buffer, place on ice for 30 min;

[0078] (5) After the lysis time is over, add loading buffer at a ratio of lysis solution: loading buffer = 4:1;

[0079] (6) Cook in a water bath at 100°C for 10 min;

[0080] (7) Clean the glass plate and prepare glue with a concentration of 10%;

[0081] (8) Load the extracted proteins in order for electrophoresis, with an electrophoresis voltage of 80 V for about 2-3 h;

[0082] (9) Cut the PVDF membrane as required, activate it with methanol for more than 10 seconds, and transfer it in the order of black-filter paper-gel-membrane-filter paper-white. The transfer current is 300 mA and the time is about 90 minutes.

[0083] (10) After transfer, block with 5% skim milk powder at room temperature for 1-2 h;

[0084] (11) After the blocking time is over, prepare the required primary antibody at a ratio of 1:1000, cut the membrane according to the molecular weight, and incubate the primary antibody at 4°C overnight;

[0085] (12) The next day, the primary antibody was collected and washed three times with 1% TBST, each time for 10 min;

[0086] (13) Prepare the secondary antibody at a ratio of 1:10000 and prepare the corresponding secondary antibody according to the requirements of the primary antibody, and incubate the secondary antibody at room temperature for 1 hour;

[0087] (14) After the secondary antibody incubation time was completed, the cells were washed three times with 1% TBST, each time for 10 min;

[0088] (15) Finally, prepare the developer in a 1:1 ratio, expose on an exposure machine, and save the result.

[0089] 2.6 Statistical analysis methods

[0090] The results were analyzed using SPSS 18.0 statistical software, and the data were expressed as mean ± standard deviation. One-way analysis of variance was used for comparison among multiple groups, and t-test was used for comparison between two groups. P < 0.05 indicated statistically significant differences.

[0091] 3. Experimental results

[0092] 3.1 Effect of simvastatin on the growth activity of HepG2 liver cancer cells

[0093] like Figure 1 As shown, simvastatin has little inhibitory effect on the activity of liver cancer cells when the concentration is 1-10 μM. When the concentration of simvastatin is 20 μM, the cell survival rate is about 75%. As the concentration increases, the growth activity of liver cancer cells HepG2 gradually weakens. When the concentration of simvastatin is 40 μM, the activity rate of liver cancer cells is about 20%.

[0094] 3.2 Effect of simvastatin on the migration ability of HepG2 liver cancer cells

[0095] like Figure 2 As shown, Figure 2 The magnification of photo A is 40x. After stimulating liver cancer cells HepG2 with 20μM simvastatin, the migration speed of cells toward the middle of the scratch was significantly reduced compared with the control group, indicating that simvastatin can inhibit the migration ability of liver cancer cells. Figure 2 As shown in B, statistical analysis showed that there was statistical significance (P<0.05).

[0096] 3.3 Effect of simvastatin on the invasion ability of HepG2 liver cancer cells

[0097] As shown in Figure 3, Figure 3 The magnification of photo A is 200x. After stimulating liver cancer cells HepG2 with 20μM simvastatin, the number of cells penetrating to the bottom of the chamber was significantly reduced compared with the control group, proving that simvastatin can inhibit the invasive ability of liver cancer cells. Figure 3 As shown in B, the statistical analysis showed statistical significance.

[0098] 3.4 Effect of simvastatin on apoptosis of HepG2 liver cancer cells

[0099] like Figure 4 As shown in A, after stimulating liver cancer cells HepG2 with 20 μM simvastatin, the expression of anti-apoptotic protein Bcl-2 was reduced and the expression of pro-apoptotic protein Bax was upregulated compared with the control group, proving that simvastatin can promote apoptosis of liver cancer cells. Figure 4 As shown in B, statistical analysis showed statistical significance.

[0100] 4. Experimental Conclusion

[0101] The results of pharmacological experiments showed that simvastatin could inhibit the proliferation, migration and invasion of liver cancer cells HepG2 in vitro, and promote the apoptosis of liver cancer cells.

[0102] Example 2: Effect of the drug of the present invention on the human hepatocellular carcinoma Hep3B mouse transplant tumor model

[0103] 1. Experimental Materials

[0104] 1.1 Animals

[0105] SPF BALB / c female nude mice, 6-8 weeks old, weighing 18-22 g, were purchased from Shanghai Experimental Animal Research Center.

[0106] 1.2 Main drugs and reagents

[0107] Human hepatocellular carcinoma cell line Hep3B was purchased from Shanghai Cell Bank, Chinese Academy of Sciences; simvastatin standard and fenofibrate standard were purchased from MCE (purity above 99.5%); atezolizumab injection (60 mg / mL) was purchased from Roche.

[0108] 2. Experimental Methods

[0109] 2.1 Establishment of transplanted tumor model and drug administration

[0110] BALB / c female nude mice were raised at room temperature and inoculated with Hep3B cells after they adapted to the environment. Each nude mouse was injected with 6×10 7One week after inoculation, the nude mice were observed under the skin of the left armpit and rice-sized nodules were found at the inoculation site. The mice were kept until the tumor diameter was about 5 mm, and the nude mice with large differences in tumor size and weight were removed, and the remaining nude mice were randomly divided into groups.

[0111] The experiment was divided into the following groups, with 10 tumor-bearing mice in each group: (1) model group; (2) atezolizumab (5 mg / kg) group; (3) simvastatin (10 mg / kg) group; (4) fenofibrate (10 mg / kg) group; (5) atezolizumab (5 mg / kg) + simvastatin (10 mg / kg) group; (6) atezolizumab (5 mg / kg) + fenofibrate (10 mg / kg) group.

[0112] The tumor-bearing mice in the model group were given 0.9% saline by intraperitoneal injection and oral administration. The second, fifth and sixth groups were given atezolizumab by intraperitoneal injection according to the weight of the tumor-bearing mice, once a week for 4 consecutive weeks. The third, fourth, fifth and sixth groups were given simvastatin or fenofibrate (prepared into a suspension with 0.9% saline before use) by oral administration according to the weight of the tumor-bearing mice, once a week for 4 consecutive weeks.

[0113] After 4 weeks, the mice in each group were killed and the tumors were removed.

[0114] 2.2 Evaluation indicators

[0115] Tumor weight and tumor inhibition rate

[0116] After 4 weeks of administration, the mice in each group were killed, and the tumors were removed and weighed.

[0117] Tumor inhibition rate (%) = (average tumor weight of model group - average tumor weight of drug administration group) / average tumor weight of model group × 100%

[0118] 2.3 Statistical methods

[0119] The results were analyzed using SPSS 18.0 statistical software, and the data were expressed as mean ± standard deviation. One-way analysis of variance was used for comparison among multiple groups, and t-test was used for comparison between two groups. P < 0.05 indicated statistically significant differences.

[0120] 3. Experimental results

[0121] In the experiment, after nude mice were inoculated with Hep3B cells, the tumors grew rapidly after a 6-day incubation period. As shown in Table 1 below, the results showed that atezolizumab (5 mg / kg) alone could effectively inhibit tumor growth (p<0.05), with a tumor inhibition rate of 32.93%.

[0122] Simvastatin and fenofibrate, drugs that affect lipid metabolism, were administered once a week for four consecutive weeks and showed low tumor inhibition, with tumor inhibition rates of only 12.20% and 8.54%, respectively, and the tumor weights of these two groups of experimental animals were not statistically different from those of the model group. This may be related to the low dosage of these two drugs in this study and the fact that they were only administered once a week.

[0123] After 4 weeks of weekly administration of atezolizumab (5 mg / kg) + simvastatin (10 mg / kg) group, the tumor weight of nude mice was significantly reduced (p<0.01), and the tumor inhibition rate reached 64.63%. This result shows that simvastatin significantly enhances the anti-liver cancer effect of atezolizumab. However, after 4 weeks of weekly administration of atezolizumab (5 mg / kg) + fenofibrate (10 mg / kg) group, the anti-tumor effect of atezolizumab was not further enhanced. The tumor inhibition rate of the drug combination group was 35.37%, which is similar to the tumor inhibition rate of the atezolizumab monotherapy group (32.93%), suggesting that the anti-liver cancer effect of atezolizumab combined with fenofibrate may mainly reflect the effect of atezolizumab.

[0124] Table 1: Inhibitory effect of the drug of the present invention on tumors in tumor-bearing mice 4 weeks after administration

[0125]

[0126] Note: *p<0.05, **p<0.01, compared with the model group.

[0127] 4. Experimental Conclusion

[0128] Experiments have shown that when atezolizumab and simvastatin are used in combination at a certain ratio, their anti-liver cancer effect is significantly better than the effect of atezolizumab and simvastatin used alone, that is, atezolizumab and simvastatin have a synergistic effect in fighting liver cancer. In addition, although simvastatin and fenofibrate are both drugs that affect lipid metabolism, when these two drugs are used in combination with atezolizumab, they show significantly different combined results. This result suggests that the anti-liver cancer effect of atezolizumab and simvastatin combined may be related to other mechanisms of action that are currently unknown in addition to lipid metabolism.

[0129] Obviously, those skilled in the art can make various changes and modifications to the present invention without departing from the spirit and scope of the present invention. Thus, if these modifications and variations of the present invention fall within the scope of the claims of the present invention and their equivalents, the present invention is also intended to include these modifications and variations.

Claims

1. Use of the composition in the preparation of a drug for treating hepatocellular carcinoma, characterized in that: The active ingredients of the composition are composed of simvastatin and atezolizumab, wherein the weight ratio of simvastatin to atezolizumab is 1:0.

5. In the composition, the simvastatin and atezolizumab show a synergistic effect in treating hepatocellular carcinoma. The simvastatin inhibits the proliferation, migration and invasion of liver cancer cells and promotes the apoptosis of liver cancer cells. In the medicine, the simvastatin and atezolizumab are administered simultaneously or separately in sequence.

2. The use according to claim 1, characterized in that: The hepatocellular carcinoma is caused by one or more diseases selected from the following: chronic hepatitis virus infection, long-term exposure to carcinogens, excessive drinking, non-alcoholic fatty liver disease, hemochromatosis or alpha-1 antitrypsin deficiency.

3. The use according to claim 2, characterized in that: The chronic hepatitis virus infection is hepatitis B virus infection, hepatitis C virus infection or hepatitis E virus infection; the carcinogen is aflatoxin.

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