Prophylactic and therapeutic drug for nonalcoholic fatty liver disease

Abstract

To provide a combination of a medicinal composition and a drug by which nonalcoholic fatty liver disease and nonalcoholic steatohepatitis can be prevented and / or treated.SOLUTION: The present invention provides a combination of a peroxisome proliferator-activated receptor (PPAR)α agonist with a sodium glucose cotransporter 2 (SGLT2) inhibitor, which is to be used for preventing and / or treating nonalcoholic fatty liver disease and nonalcoholic steatohepatitis.SELECTED DRAWING: None

Description

[Technical field]

[0001] The present invention relates to an agent for the prevention and / or treatment of non-alcoholic fatty liver disease. [Background technology]

[0002] Nonalcoholic fatty liver disease (NAFLD) is a fatty liver disorder not caused by alcoholic hepatitis or viral hepatitis, and is estimated to occur in about 30% of the general population. NAFLD is a general term that includes symptoms ranging from simple steatosis, which has a relatively good prognosis due to only fatty deposition in hepatocytes, to nonalcoholic steatohepatitis (NASH), which can lead to relatively severe fibrosis of liver tissue, cirrhosis, and hepatocellular carcinoma. In recent years, the treatment of viral hepatitis, represented by hepatitis C, has rapidly advanced, and while the number of patients with viral liver cancer is expected to decrease in the future, there is concern about an increase in the number of patients with liver cancer based on NASH (Non-Patent Documents 1, 2, 3).

[0003] The mechanism of onset of NAFLD is widely known as the "two hit theory," which separates the stage of fat accumulation in hepatocytes from the stage of progression of inflammation and fibrosis (Non-Patent Document 4), but in recent years, the "multiple-parallel hit theory," in which other factors are simultaneously involved in the progression of the disease, has been proposed (Non-Patent Document 5). When diagnosing NASH, ballooning of hepatocytes, Mallory-Denk bodies, and fibrosis are important factors, and the Japanese Society of Gastroenterology's NAFLD / NASH treatment guidelines define NASH as a pathological diagnostic criterion that "in addition to large droplet fatty changes in hepatocytes, ballooning of hepatocytes accompanied by inflammation is observed." Matteoni et al. have divided NAFLD into four stages based on pathological findings with an emphasis on prognosis, and have defined types 3 and 4 as NASH (Non-Patent Documents 3, 6). Hepatocyte hypertrophy and balloon-like degeneration are findings of degeneration of the cytoskeleton resulting from fat accumulation in hepatocytes, and these indicators are important findings in distinguishing whether or not a patient has NASH.

[0004] The principle of NAFLD treatment is to correct the underlying obesity, diabetes, dyslipidemia, and hypertension by improving lifestyle habits such as dietary therapy and exercise therapy. In clinical practice, in addition to improving lifestyle habits, drug treatments targeting insulin resistance, lipid metabolism disorders, hypertension, oxidative stress, and the like are being performed. For insulin resistance, insulin sensitizers such as thiazolidine derivatives (pioglitazone, rosiglitazone, etc.), which are ligands of the nuclear receptor PPARγ involved in the enhancement of insulin sensitivity, or biguanide drugs (metformin, etc.) are used. For lipid metabolism disorders, fibrate drugs (fenofibrate, bezafibrate, etc.), which are PPARα agonists, statin preparations, small intestinal cholesterol reabsorption inhibitors (ezetimibe, etc.), and angiotensin II type 1 receptor antagonists (ARBs) and the like are used as hypertension treatment agents (Non-Patent Documents 1 and 3). In addition, vitamin E and the like are used as antioxidants.

[0005] These drug therapies are selected appropriately depending on the patient's underlying disease, etc. However, further verification is required for each drug therapy, and none has yet been established as a drug therapy for NAFLD.

[0006] For fibrate drugs, clinical trials on the effect of fenofibrate on NAFLD have been reported (Non-Patent Document 7). In addition, (R)-2-[3-[[N-(benzoxazol-2-yl)-N-3-(4-methoxyphenoxy)propyl]aminomethyl]phenoxy]butyric acid, or its salt or solvate thereof, which has selective PPARα activation activity as disclosed in Patent Document 1, has been shown to be useful for the prevention and treatment of non-alcoholic fatty liver disease (Patent Document 2). On the other hand, the action of remogliflozin, a sodium-glucose cotransporter 2 (SGLT2) inhibitor, has been investigated in clinical trials, and its improving effect on NASH has been reported (Non-Patent Document 8). With the increase in metabolic syndrome patients worldwide, the number of NASH patients is also expected to increase. Since NASH is thought to be the cause of non-viral hepatocellular carcinoma, which accounts for a large factor in cancer-related deaths (Non-Patent Document 9), the establishment of a more effective treatment method is desired. [Prior art documents] [Patent documents]

[0007] [Patent Document 1] International Publication No. 2005 / 023777 [Patent Document 2] International Publication No. 2015 / 005365 [Non-patent literature]

[0008] [Non-Patent Document 1] Chalasani N. et al. Hepatology, 55, 2005-23 (2012) [Non-Patent Document 2] Musso G. et al. Nat. Rev. Drug Discov. 15(4), 249-74 (2016.1) [Non-Patent Document 3] Japanese Society of Gastroenterology NAFLD / NASH Clinical Practice Guidelines 2014 [Non-Patent Document 4] Day CP. et al, Gastroenterology, 114(4), 842-5 (1998) [Non-Patent Document 5] Tilg H. et al. Hepatology, 52, 1836-46 (2010) [Non-Patent Document 6] Matteoni CA. et al. Gastroenterology, 116, 1413-9 (1999) [Non-Patent Document 7] Fernandez-Miranda C. et al. Dig. Liver Dis., 40, 200-5 (2008) [Non-Patent Document 8] Wilkison W. et al. Abstract O047. International Liver Congress; April 22-26, 2015 [Non-Patent Document 9] Fujii M. et al. Med. Mol. Morphol, 46, 141-52 (2013) Summary of the Invention [Problem to be solved by the invention]

[0009] The objective of the present invention is to provide a pharmaceutical composition or drug combination that has the effect of preventing and / or improving the enlargement and / or balloon degeneration of lipid droplets in hepatocytes and is capable of preventing and / or treating NAFLD and NASH. [Means for solving the problem]

[0010] In view of the current situation, the present inventors conducted intensive studies using NASH-HCC mice, an animal model for NASH, in order to find a method useful for preventing and / or treating nonalcoholic fatty liver disease (NAFLD), particularly nonalcoholic steatohepatitis (NASH), which has severe symptoms. As a result, they found that a combination of a PPARα agonist, (R)-2-[3-[[N-(benzoxazol-2-yl)-N-3-(4-methoxyphenoxy)propyl]aminomethyl]phenoxy]butyric acid (hereinafter sometimes referred to as compound 1), and an SGLT2 inhibitor (disclosed in Expert Opin. Investig. Drugs (2013) 22(4):463-486, etc.) has a strong effect of reducing the size of lipid droplets in hepatocytes and a strong effect of inhibiting balloon degeneration, and thus has an effect of preventing and / or treating NAFLD and NASH, thereby completing the present invention.

[0011] That is, the present invention relates to a composition, a kit, etc. that is characterized by a combination of a PPARα agonist and an SGLT2 inhibitor. More specifically, the present invention relates to the following [1] to

[52] . [1] A preventive and / or therapeutic agent for liver disease comprising a combination of a PPARα agonist and an SGLT2 inhibitor. [2] The preventive and / or therapeutic agent according to [1] above, wherein the liver disease is non-alcoholic fatty liver disease. [3] The preventive and / or therapeutic agent according to [2] above, wherein the non-alcoholic fatty liver disease is non-alcoholic steatohepatitis. [4] The preventive and / or therapeutic agent according to [2] above, which has an inhibitory effect on balloon degeneration of hepatocytes in non-alcoholic fatty liver disease. [5] The preventive and / or therapeutic agent according to [1] above, wherein the liver disease is cirrhosis or hepatocellular carcinoma. [6] PPARα agonists: The preventive and / or therapeutic agent according to any one of the above [1] to [5], which is (R)-2-[3-[[N-(benzoxazol-2-yl)-N-3-(4-methoxyphenoxy)propyl]aminomethyl]phenoxy]butyric acid, a salt thereof, or a solvate thereof. [7] The preventive and / or therapeutic agent according to any one of the above [1] to [6], wherein the SGLT2 inhibitor is dapagliflozin, canagliflozin, ipragliflozin, empagliflozin, luseogliflozin, tofogliflozin, ertugliflozin, sotagliflozin, bexagliflozin or remogliflozin. [8] The preventive and / or therapeutic agent according to any one of [1] to [7] above, which is a combination drug. [9] The preventive and / or therapeutic agent according to any one of [1] to [7] above, which is a kit.

[0012]

[10] A pharmaceutical composition for use in the prevention and / or treatment of liver disease, comprising a PPARα agonist in combination with an SGLT2 inhibitor.

[11] The pharmaceutical composition according to

[10] above, wherein the liver disease is non-alcoholic fatty liver disease.

[12] The pharmaceutical composition according to

[11] above, wherein the non-alcoholic fatty liver disease is non-alcoholic steatohepatitis.

[13] The pharmaceutical according to

[11] above, which has an inhibitory effect on balloon degeneration of hepatocytes in non-alcoholic fatty liver disease.

[14] The pharmaceutical composition according to

[10] above, wherein the liver disease is cirrhosis or hepatocellular carcinoma.

[15] The pharmaceutical agent according to any one of the above

[10] to

[14] , wherein the PPARα agonist is (R)-2-[3-[[N-(benzoxazol-2-yl)-N-3-(4-methoxyphenoxy)propyl]aminomethyl]phenoxy]butyric acid or a salt thereof, or a solvate thereof.

[16] The medicine according to any one of

[10] to

[15] , wherein the SGLT2 inhibitor is dapagliflozin, canagliflozin, ipragliflozin, empagliflozin, luseogliflozin, tofogliflozin, ertugliflozin, sotagliflozin, bexagliflozin or remogliflozin.

[17] The pharmaceutical agent according to any one of

[10] to

[16] above, which is a combination drug.

[18] The pharmaceutical according to any one of

[10] to

[16] above, which is a kit.

[0013]

[19] A pharmaceutical composition for the prevention and / or treatment of liver disease, comprising a PPARα agonist, an SGLT2 inhibitor, and a pharma- ceutical acceptable carrier.

[20] The pharmaceutical composition according to

[19] above, wherein the liver disease is non-alcoholic fatty liver disease.

[21] The pharmaceutical composition according to

[20] above, wherein the non-alcoholic fatty liver disease is non-alcoholic steatohepatitis.

[22] The pharmaceutical composition according to

[20] above, which has an inhibitory effect on balloon degeneration of hepatocytes in non-alcoholic fatty liver disease.

[23] The pharmaceutical composition described in

[19] above, wherein the liver disease is cirrhosis or hepatocellular carcinoma.

[24] The pharmaceutical composition according to any one of

[19] to

[23] above, wherein the PPARα agonist is (R)-2-[3-[[N-(benzoxazol-2-yl)-N-3-(4-methoxyphenoxy)propyl]aminomethyl]phenoxy]butyric acid or a salt thereof, or a solvate thereof.

[25] The pharmaceutical composition according to any one of

[19] to

[24] , wherein the SGLT2 inhibitor is dapagliflozin, canagliflozin, ipragliflozin, empagliflozin, luseogliflozin, tofogliflozin, ertugliflozin, sotagliflozin, bexagliflozin or remogliflozin.

[0014]

[26] A method for preventing and / or treating a liver disease, comprising the step of administering to a subject in need of treatment an effective amount of a PPARα agonist and an effective amount of an SGLT2 inhibitor.

[27] The method for prevention and / or treatment according to

[26] above, wherein the liver disease is non-alcoholic fatty liver disease.

[28] The method for preventing and / or treating non-alcoholic fatty liver disease according to

[27] above, wherein the non-alcoholic fatty liver disease is non-alcoholic steatohepatitis.

[29] The method for prevention and / or treatment according to

[27] above, which inhibits balloon degeneration of hepatocytes in non-alcoholic fatty liver disease.

[30] The method for prevention and / or treatment according to

[26] above, wherein the liver disease is cirrhosis or hepatocellular carcinoma.

[31] The method for prevention and / or treatment according to any one of the above

[26] to

[30] , wherein the PPARα agonist is (R)-2-[3-[[N-(benzoxazol-2-yl)-N-3-(4-methoxyphenoxy)propyl]aminomethyl]phenoxy]butyric acid or a salt thereof, or a solvate thereof.

[32] The method for prevention and / or treatment according to any one of

[26] to

[31] , wherein the SGLT2 inhibitor is dapagliflozin, canagliflozin, ipragliflozin, empagliflozin, luseogliflozin, tofogliflozin, ertugliflozin, sotagliflozin, bexagliflozin or remogliflozin.

[33] The method for prevention and / or treatment according to any one of

[26] to

[32] above, wherein a PPARα agonist and an SGLT2 inhibitor are administered simultaneously.

[34] The preventive and / or therapeutic method according to any one of

[26] to

[32] above, wherein the PPARα agonist and the SGLT2 inhibitor are administered separately with an interval in between.

[0015]

[35] Use of a PPARα agonist and an SGLT2 inhibitor for the manufacture of an agent for the prevention and / or treatment of liver disease.

[36] The use according to

[35] above, wherein the liver disease is non-alcoholic fatty liver disease.

[37] The use according to

[36] above, wherein the non-alcoholic fatty liver disease is non-alcoholic steatohepatitis.

[38] The use according to

[36] above, which has an inhibitory effect on balloon degeneration of hepatocytes in non-alcoholic fatty liver disease.

[39] The use described in

[35] above, wherein the liver disease is cirrhosis or hepatocellular carcinoma.

[40] The use according to any one of

[35] to

[39] above, wherein the PPARα agonist is (R)-2-[3-[[N-(benzoxazol-2-yl)-N-3-(4-methoxyphenoxy)propyl]aminomethyl]phenoxy]butyric acid or a salt thereof, or a solvate thereof.

[41] The use according to any one of

[35] to

[40] above, wherein the SGLT2 inhibitor is dapagliflozin, canagliflozin, ipragliflozin, empagliflozin, luseogliflozin, tofogliflozin, ertugliflozin, sotagliflozin, bexagliflozin or remogliflozin.

[42] The use according to any one of

[35] to

[41] above, wherein the prophylactic and / or therapeutic agent is a combination drug.

[43] The use according to any one of

[35] to

[41] above, wherein the preventive and / or therapeutic agent is a kit.

[0016]

[44] A combination of a PPARα agonist and an SGLT2 inhibitor for preventing and / or treating liver disease.

[45] The combination according to

[44] above, wherein the liver disease is non-alcoholic fatty liver disease.

[46] The combination according to

[45] above, wherein the non-alcoholic fatty liver disease is non-alcoholic steatohepatitis.

[47] The combination according to

[45] above, which has an inhibitory effect on balloon degeneration of hepatocytes in non-alcoholic fatty liver disease.

[48] ​​The combination according to

[44] above, wherein the liver disease is cirrhosis or hepatocellular carcinoma.

[49] The combination according to any one of

[44] to

[48] above, wherein the PPARα agonist is (R)-2-[3-[[N-(benzoxazol-2-yl)-N-3-(4-methoxyphenoxy)propyl]aminomethyl]phenoxy]butyric acid or a salt thereof, or a solvate thereof.

[50] The combination according to any one of

[44] to

[49] , wherein the SGLT2 inhibitor is dapagliflozin, canagliflozin, ipragliflozin, empagliflozin, luseogliflozin, tofogliflozin, ertugliflozin, sotagliflozin, bexagliflozin or remogliflozin.

[51] The combination according to any one of

[44] to

[50] above, which is a combination drug.

[52] The combination according to any one of

[44] to

[50] above, which is a kit. Effect of the Invention

[0017] The therapeutic agent, medicament, pharmaceutical composition, treatment method, use, or combination of the present invention can suppress lipid droplet hypertrophy and balloon-like degeneration of hepatocytes derived from nonalcoholic fatty liver disease (NAFLD) and nonalcoholic steatohepatitis (NASH), providing a new prevention and / or treatment for NAFLD and NASH, particularly for severe NASH. [Brief description of the drawings]

[0018] [Figure 1] FIG. 1 is a graph showing the lipid droplet size (μm2) in hepatocytes when compound 1 (0.1 mg / kg) of the present invention, tofogliflozin (10 mg / kg), or compound 1 (0.1 mg / kg) and tofogliflozin (10 mg / kg) were administered. [Diagram 2] FIG. 2 is a graph showing ballooning of hepatocytes when compound 1 (0.1 mg / kg), tofogliflozin (10 mg / kg), or compound 1 (0.1 mg / kg) and tofogliflozin (10 mg / kg) of the present invention were administered. [Diagram 3] FIG. 3 is a graph showing ballooning of hepatocytes when compound 1 (0.1 mg / kg), ipragliflozin (3 mg / kg), or compound 1 (0.1 mg / kg) and ipragliflozin (3 mg / kg) of the present invention were administered. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0019] In the present invention, PPARα agonist refers to a general term for compounds that activate PPARα type receptors, which are involved in fat oxidation among peroxisome proliferator-activated receptors (PPARs), which are one of the nuclear receptors.Specific examples include fibrates such as fenofibrate, clofibrate, bezafibrate, clinofibrate, ciprofibrate, etofibrate, and gemfibrozil, WY-14643 (pirinixic acid), GW-7647 (2-(4-(2-(1-1-cyclohexanebutyl)-3-cyclohexylureido)ethyl)phenylthio)-2-methylpropionic acid), and pemafibrate. The (R)-2-[3-[[N-(benzoxazol-2-yl)-N-3-(4-methoxyphenoxy)propyl]aminomethyl]phenoxy]butyric acid (compound 1) used in the present invention is also known as pemafibrate, and can be produced, for example, according to the method described in WO2005 / 023777, etc. It can also be produced according to known methods described in the literature.

[0020] In the present invention, the term "SGLT2 inhibitor" refers to a general term for compounds having inhibitory activity against sodium-glucose cotransporter 2 (SGLT2), which is involved in glucose reabsorption in the kidney. Specific examples include dapagliflozin, canagliflozin, ipragliflozin (ASP1941), empagliflozin (BI 10773), luseogliflozin (TS-071), tofogliflozin (CSG452), ertugliflozin (PF-04971729), sotagliflozin (LX-4211), bexagliflozin (EGT-1442), remogliflozin (KGT-1681), and the like. These compounds may be used as appropriate pharma- ceutically acceptable salts and / or solvates, all of which are included in the present invention.

[0021] Ipragliflozin is the generic name for the compound (1S)-1,5-anhydro-1-C-{3-[(1-benzothiophen-2-yl)methyl]-4-fluorophenyl}-D-glucitol. Ipragliflozin can be used, for example, as ipragliflozin L-proline (1:1).

[0022] Tofogliflozin is the generic name of the compound (1S,3'R,4'S,5'S,6'R)-6-[(4-ethylphenyl)methyl]-6'-(hydroxymethyl)-3',4',5',6'-tetrahydro-3H-spiro[2-benzofuran-1,2'-pyran]-3',4',5'-triol. Tofogliflozin can be used, for example, as tofogliflozin monohydrate.

[0023] Ipragliflozin can be produced, for example, according to the method described in WO2004 / 080990, etc. Ipragliflozin can also be produced in accordance with known methods described in the literature.

[0024] Tofogliflozin can be produced, for example, according to the method described in WO2006 / 080421 etc. Alternatively, it can be produced in accordance with known methods described in the literature.

[0025] In the present invention, liver diseases include fatty liver, hepatitis, NAFLD, NASH, liver cirrhosis, and liver cancer such as hepatocellular carcinoma.

[0026] The "salt" of the present invention is not particularly limited as long as it is pharma- ceutically acceptable, and examples thereof include alkali metal salts such as sodium salts and potassium salts; alkaline earth metal salts such as calcium salts and magnesium salts; inorganic base salts such as ammonium salts; organic base salts such as trialkylamine salts; mineral acid salts such as hydrochlorides and sulfates; and organic acid salts such as acetates.

[0027] The "solvate" of the present invention includes hydrates, alcohol solvates (for example, ethanol solvates), and the like.

[0028] A combination drug is a pharmaceutical product that contains multiple active ingredients in one formulation. In the present invention, an example is a single tablet that contains an effective amount of a PPARα agonist and an SGLT2 inhibitor.

[0029] A kit is a combination of medicines for taking or administering multiple drugs simultaneously or separately at intervals. In the present invention, an example is a combination of a medicine containing an effective amount of a PPARα agonist and a medicine containing an effective amount of an SGLT2 inhibitor.

[0030] In NASH, lipid droplets in hepatocytes become larger and characteristically have balloon-like degeneration (see Non-Patent Documents 3 and 6). As shown in the Examples below, the combined use of Compound 1 and Ipragliflozin significantly suppressed the balloon-like degeneration of hepatocytes in NASH-HCC mice, which are model animals of NASH. In addition, the combined use of Compound 1 and Tofogliflozin significantly suppressed the balloon-like degeneration of hepatocytes and suppressed the enlargement of lipid droplets in hepatocytes in NASH-HCC mice. Therefore, the combined use of the PPARα agonist and SGLT2 inhibitor of the present invention is useful as a preventive and / or therapeutic agent for NASH in mammals, including humans.

[0031] The therapeutic agent, medicine, etc. comprising a combination of the PPARα agonist and SGLT2 inhibitor of the present invention can be made into dosage forms such as tablets, capsules, granules, powders, lotions, ointments, injections, suppositories, etc., either alone or using other pharma- ceutically acceptable carriers, and these preparations can be manufactured by known methods.

[0032] The therapeutic agent, medicine, etc., which is a combination of the PPARα agonist and the SGLT2 inhibitor of the present invention, is administered orally or parenterally. The dosage can be appropriately set by a person skilled in the art depending on the patient's weight, age, sex, symptoms, etc., but when compound 1 is administered to an adult as a PPARα agonist, it is preferable to administer 0.01 to 1000 mg, preferably 0.01 to 10 mg, more preferably 0.05 to 5 mg, divided into 1 to 3 doses per day. When ipragliflozin is used as an SGLT2 inhibitor, it is preferable to administer 0.1 to 1000 mg, preferably 1 to 200 mg, per day, and when tofogliflozin is used, it is preferable to administer 0.1 to 500 mg, preferably 1 to 100 mg, per day, divided into 1 to 3 doses. EXAMPLES

[0033] The present invention will be described in more detail below with reference to examples, but the present invention is not limited to these examples in any way.

[0034] Example 1 Effect of Compound 1 and Tofogliflozin on NASH Mouse Model The effects of compound 1 and tofogliflozin were examined in NASH-HCC mice (Non-Patent Document 9), a mouse model that develops fatty liver, NASH, and then liver cirrhosis and subsequently hepatocellular carcinoma when ingesting a high-fat diet. In this test, Compound 1 was prepared and used according to the method described in the above-mentioned Patent Document 1. Tofogliflozin monohydrate (Chugai Pharmaceutical Co., Ltd.) was used as tofogliflozin.

[0035] 1) Animals used: NASH-HCC mice were created and subjected to the experiment with reference to Non-Patent Document 9. Specifically, 200 μg of streptozotocin was subcutaneously administered to male C57BL / 6J mice on the second day after birth, and then the mice were allowed to freely consume a high-fat diet (HFD-32, Nippon Clea) for 2 weeks from the age of 4 weeks. After that, the amount of food intake was measured every day from the 6th to 9th week of the administration period, and all groups were fed the same amount of food as the control group. 2) Group composition: To avoid any difference in body weight between the groups, NASH-HCC mice were divided into a control group, a compound 1 0.1 mg / kg administration group, a tofogliflozin 10 mg / kg administration group, and a combination group (compound 1 0.1 mg / kg and tofogliflozin 10 mg / kg) just before the start of the administration period (6 weeks of age) (n=5-7). 3) Drug Administration: The administration dose was 5 mL / kg body weight, and a 3% gum arabic solution, which is the drug solvent, was orally administered once a day to the control group, while the compound 1 administration group, tofogliflozin administration group, and compound 1 and tofogliflozin combined administration group were orally administered the respective drug solutions once a day. The administration period was 3 weeks from 6 weeks of age as described above.

[0036] 4) Observation and inspection methods: After administration, the liver was removed under anesthesia and fixed in 10% neutral buffered formalin solution, and hematoxylin-eosin stained specimens were prepared. The hematoxylin-eosin stained specimens were used to analyze the lipid droplet size in hepatocytes using the image analysis software Image J. The area of ​​6,000 to 10,000 lipid droplets was calculated from each specimen. The median of the lipid droplet area of ​​each individual was then calculated, and the median and quartile of each group were calculated based on the median of each individual within the group, and the data were displayed in a box plot (Figure 1). The ballooning of hepatocytes was scored under blinded conditions according to the following criteria (Kleiner et al. Hepatology 41, 1313-21, 2005), and the average score (ballooning score) for each group was calculated and the data was expressed (Figure 2). No balloon cells: 0 Few balloon cells: 1 Many or prominent ballooning cells (Ballooning): 2 Statistical processing was performed using EZR. EZR is a statistical software that extends the functions of R and R Commander, and is distributed free of charge on the website of the Saitama Medical Center, Jichi Medical University (Bone Marrow Transplantation (2013) 48, 452-458). When there was a significant difference from the control group at p<0.05 using the Steel test (N=5-7), an * was marked in the figure. When there was a significant difference from the control group at p<0.01, an ** was marked in the figure.

[0037] 5) Results As shown in Figure 1, the median lipid droplet size in the control group was 1.17 (μm 2 ) in the group administered 0.1 mg / kg of Compound 1, whereas the mean mean mean was 1.00 (μm 2 ), and 1.00 (μm 2 ), and although there was a tendency for the size of lipid droplets in hepatocytes to decrease, no statistically significant difference was observed. However, the median lipid droplet size in the combined treatment group was 0.89 (μm 2 ) and a significant decrease (p=0.037) was observed compared to the control group. As shown in Figure 2, the ballooning score of the control group was 1.33, whereas the ballooning score of the compound 1 0.1 mg / kg group was 0.60, and the tofogliflozin 10 mg / kg group was 0.71, and although a tendency to suppress ballooning degeneration of hepatocytes was observed, no statistically significant difference was observed. However, in the group administered both drugs in combination, the ballooning score was 0.14, and a significant suppression of ballooning degeneration of hepatocytes (p=0.0089) was observed compared to the control group.

[0038] Example 2 Effect of Compound 1 and Ipragliflozin on NASH Mouse Model The effects of Compound 1 and ipragliflozin in NASH-HCC mice (Non-Patent Document 9) were investigated. In this test, Compound 1 was prepared and used according to the method described in Patent Document 1. Ipragliflozin (Shanghai Haoyuan Chamexpress, Shanghai, China) was used as ipragliflozin.

[0039] 1) Animals used: NASH-HCC mice were prepared in the same manner as in Example 1, 1) and subjected to the experiment. 2) Group composition: To avoid any difference in body weight between the groups, NASH-HCC mice were divided into a control group, a group administered 0.1 mg / kg of compound 1, a group administered 3 mg / kg of ipragliflozin, and a combination group (0.1 mg / kg of compound 1 and 3 mg / kg of ipragliflozin) just before the start of the administration period (6 weeks of age) (n=8). 3) Drug Administration: The administration dose was 5mL / kg body weight, and the control group was orally administered a 3% gum arabic solution, which is the drug solvent, and the compound 1 administration group, ipragliflozin administration group, and the compound 1 and ipragliflozin combination administration group were orally administered the respective drug solutions once a day. The administration period was 3 weeks from the age of 6 weeks.

[0040] 4) Observation and inspection methods: Evaluation was carried out in the same manner as in Example 1, 4).

[0041] 5) Results As shown in Figure 3, the ballooning score of the control group was 1.50, whereas the ballooning score of the 0.1 mg / kg compound 1 group was 1.35, and the 3 mg / kg ipragliflozin group was 0.88. Although a tendency to suppress ballooning degeneration of hepatocytes was observed, no statistically significant difference was observed. However, the ballooning score of the combined use of both drugs was 0.38, and a significant suppression of ballooning degeneration of hepatocytes (p=0.009) was observed compared to the control group.

[0042] From the above results, the combined use of compound 1 of the present invention with SGLT2 inhibitors such as tofogliflozin and ipragliflozin significantly reduced the lipid droplet size of liver cells and inhibited balloon-like degeneration of liver cells in NASH mouse models. The reduction in lipid droplet size and inhibition of balloon-like degeneration of liver cells improves the pathology of NAFLD and NASH, and ultimately leads to the prevention of cirrhosis and hepatocellular carcinoma, which are the terminal pathologies of these diseases. [Industrial Applicability]

[0043] Since the combined use of a PPARα agonist and an SGLT2 inhibitor showed preventive and / or therapeutic effects against NAFLD and NASH, the present invention has industrial applicability.

Claims

[Claim 1] A preventive and / or therapeutic agent for liver disease comprising a PPARα agonist and an SGLT2 inhibitor in combination.

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