Pharmaceutical composition for preventing or treating metabolic dysfunction-related fatty liver disease comprising empagliflozin and telmisartan

Abstract

The present invention relates to a pharmaceutical composition for preventing or treating metabolic abnormality-related fatty liver diseases containing empagliflozin and telmisartan. The pharmaceutical composition has no issues of drug-drug interaction and exhibits a synergistic effect in the prevention or treatment of metabolic dysfunction-related fatty liver disease, thereby allowing effective application to patients with metabolic dysfunction-related fatty liver disease.

Description

Pharmaceutical composition for preventing or treating fatty liver disease associated with metabolic disorders containing empagliflozin and telmisartan

[0001] The present invention relates to a pharmaceutical composition for preventing or treating fatty liver disease related to metabolic disorders, containing empagliflozin and telmisartan.

[0002] Metabolic fatty liver disease (FLD) is diagnosed when intracellular fat deposition is observed on imaging or histological examination, and when there is no other cause of intracellular fat accumulation, including significant alcohol consumption, persistent medication use, or genetic disorders. This condition is known to encompass both simple fatty liver disease and metabolic steatohepatitis.

[0003] Fatty liver disease related to metabolic disorders is a very common disease, and its prevalence is rapidly increasing along with the increase in the incidence of various metabolic syndromes such as obesity and diabetes. It is known that the incidence of cirrhosis due to metabolic steatohepatitis is higher than that of simple fatty liver, and it has been reported that the risk of liver failure or hepatocellular carcinoma due to cirrhosis as well as liver-related mortality increases.

[0004] Fatty liver disease related to metabolic disorders is reported to increase with age and to show differences by gender. This is known to be due to a decrease in insulin sensitivity and hormonal changes with age.

[0005] Metabolic fatty liver disease (FLD) is known to be closely associated with metabolic syndrome, overweight and obesity, type 2 diabetes, and dyslipidemia. Ninety percent of patients with FLD have at least one component of FLD, and one-third have FLD. Overweight and obesity are risk factors for FLD, but central obesity is a particularly strong risk factor.

[0006] Accordingly, the inventors of the present invention conducted research to develop a combination therapy that can be effectively applied to the treatment of fatty liver disease related to metabolic disorders, and completed the present invention.

[0007] One object of the present invention is to provide a pharmaceutical composition for preventing or treating metabolic dysfunction-associated steatotic liver disease, comprising an angiotensin II receptor blocker or a pharmaceutically acceptable salt thereof as a first pharmacological ingredient; and a sodium-glucose cotransporter-2 (SGLT-2) inhibitor or a pharmaceutically acceptable salt thereof as a second pharmacological ingredient.

[0008] Another object of the present invention is to provide a method for preventing or treating fatty liver disease associated with metabolic disorders, comprising administering the pharmaceutical composition to a subject.

[0009] Another object of the present invention is to provide a kit comprising the composition.

[0010] One aspect of the present invention provides a pharmaceutical composition for preventing or treating metabolic dysfunction-associated steatotic liver disease, comprising an angiotensin II receptor blocker or a pharmaceutically acceptable salt thereof as a first pharmacological ingredient; and a sodium-glucose cotransporter-2 (SGLT-2) inhibitor or a pharmaceutically acceptable salt thereof as a second pharmacological ingredient.

[0011] According to one specific example of the present invention, the sodium-glucose cotransporter-2 (SGLT-2) inhibitor may be empagliflozin.

[0012] According to one specific example of the present invention, the angiotensin II receptor blocker may be telmisartan.

[0013] According to one specific example of the present invention, the fatty liver disease associated with metabolic dysfunction may be simple steatosis or metabolic dysfunction-associated steatohepatitis.

[0014] According to one specific example of the present invention, the weight ratio of the angiotensin II receptor blocker and the sodium glucose transporter-2 inhibitor may be 2:1 to 8:1.

[0015] Another aspect of the present invention provides a method for preventing or treating fatty liver disease associated with metabolic disorders, comprising administering the pharmaceutical composition to a subject.

[0016] Another aspect of the present invention provides a kit comprising the composition.

[0017] According to a pharmaceutical composition for preventing or treating fatty liver disease associated with metabolic disorders containing empagliflozin and telmisartan, there is no drug interaction problem and a synergistic effect is shown in the prevention or treatment of fatty liver disease associated with metabolic disorders, so it can be effectively applied to patients with fatty liver disease associated with metabolic disorders.

[0018] Figure 1 is a fluorescence micrograph showing (A) normal HepG2 cells, (B) HepG2 cells induced to have a metabolically abnormal fatty liver state by oleic acid (OA), and (C) HepG2 cells induced to have a metabolically abnormal fatty liver state by oleic acid and treated with 10 μM empagliflozin and 20 μM telmisartan.

[0019] Figure 2 is a graph showing the increase rate (fold change) of mRNA expression of PPAR-α compared to β-actin according to the treatment concentration (5 μM+5 μM, 5 μM+10 μM, 10 μM+20 μM) of empagliflozin (Em) and telmisartan (Telm) in HepG2 cells induced to a state of metabolic abnormality fatty liver by oleic acid (OA).

[0020] Figure 3 is a graph showing the increase rate (fold change) of mRNA expression of PPAR-γ compared to β-actin according to the treatment concentration of empagliflozin (Em) and telmisartan (Telm) (5 μM+5 μM, 5 μM+10 μM, 10 μM+20 μM) in HepG2 cells induced to a state of metabolic abnormality fatty liver by oleic acid (OA).

[0021] One aspect of the present invention provides a pharmaceutical composition for preventing or treating metabolic dysfunction-associated steatotic liver disease, comprising an angiotensin II receptor blocker or a pharmaceutically acceptable salt thereof as a first pharmacological ingredient; and a sodium-glucose cotransporter-2 (SGLT-2) inhibitor or a pharmaceutically acceptable salt thereof as a second pharmacological ingredient.

[0022] The SGLT-2 inhibitor included in the pharmaceutical composition of the present invention may be any one substance selected from the group consisting of dapagliflozin, empagliflozin, ipragliflozin, canagliflozin, luseogliflozin, and tofogliflozin, but empagliflozin is preferable in terms of combination compatibility and synergistic effect.

[0023] The dosage of the pharmaceutical composition according to one specific example of the present invention may be, for example, in the range of about 0.001 mg / kg to about 100 mg / kg, about 0.01 mg / kg to about 10 mg / kg, or about 0.1 mg / kg to about 1 mg / kg on an adult basis, and may be administered once a day, multiple times a day, or once a week, once every two weeks, once every three weeks, or once every four weeks to once a year, and may be divided into once a day or several times a day at regular intervals according to the judgment of a doctor or pharmacist.

[0024] The pharmaceutical composition of the present invention can be prepared by mixing an angiotensin II receptor blocker as a first pharmacological ingredient and an SGLT-2 inhibitor as a second pharmacological ingredient, and the mixing thereof can be performed simultaneously or sequentially, and can be performed using a method known in the art.

[0025] The pharmaceutical composition of the present invention may be, for example, a double-layer tablet comprising an angiotensin II receptor blocker as a first pharmacological ingredient and an SGLT-2 inhibitor as a second pharmacological ingredient, or may be an inner core tablet structure comprising an angiotensin II receptor blocker as a first pharmacological ingredient and an SGLT-2 inhibitor as a second pharmacological ingredient and an outer layer surrounding an inner core layer, but is not limited thereto.

[0026] The composition of the present invention may include a pharmaceutically acceptable carrier. Pharmaceutically acceptable carriers included in the composition of the present invention are those commonly used in the manufacture of pharmaceuticals, and include, but are not limited to, lactose, dextrose, sucrose, sorbitol, mannitol, starch, acacia gum, calcium phosphate, alginate, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrup, methyl cellulose, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and mineral oil. In addition to the above components, the pharmaceutical composition of the present invention may further include lubricants, wetting agents, sweetening agents, flavoring agents, emulsifiers, suspending agents, preservatives, and the like. Suitable pharmaceutically acceptable carriers and formulations are described in detail in Remington: the science and practice of pharmacy 22nd edition (2013).

[0027] A pharmaceutical composition according to one specific example of the present invention may be administered together with a substance exhibiting pharmacological activity against one or more metabolic abnormalities-related fatty liver diseases.

[0028] In addition, the pharmaceutical composition according to one embodiment of the present invention can be used alone or in combination with methods using surgery, hormone therapy, drug therapy, and / or biological response modifiers for the prevention or treatment of fatty liver disease related to metabolic disorders.

[0029] The composition of the present invention may include various bases and / or additives necessary and appropriate for the formulation of the dosage form, and may be manufactured by further including known compounds such as nonionic surfactants, silicone polymers, pigments, fragrances, preservatives, bactericides, oxidation stabilizers, organic solvents, ionic or nonionic thickeners, softeners, antioxidants, free radical scavengers, opacifiers, stabilizers, emollients, silicones, α-hydroxy acids, antifoaming agents, moisturizers, vitamins, insect repellents, fragrances, preservatives, surfactants, anti-inflammatory agents, substance P antagonists, fillers, polymers, propellants, alkalizing or acidifying agents, or colorants, within a range that does not reduce the effectiveness thereof.

[0030] The appropriate dosage of the composition of the present invention may be prescribed in various ways depending on factors such as the formulation method, administration method, patient age, weight, sex, pathological condition, food, administration time, administration route, excretion rate, and response sensitivity. The dosage of the composition of the present invention may be 0.001 to 1000 mg / kg for adults.

[0031] The composition of the present invention can be administered orally.

[0032] The composition of the present invention can be administered in various dosage forms when administered orally, such as tablets, pills, hard / soft capsules, liquids, suspensions, emulsifiers, syrups, granules, elixirs, troches, etc., and may further include various excipients, for example, wetting agents, sweeteners, fragrances, preservatives, etc. Specifically, when the composition of the present invention is formulated into an oral administration dosage form, it may further include appropriate carriers, excipients, and diluents commonly used in the manufacture thereof. Examples of the carrier, excipient and diluent may include, but are not limited to, lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methyl hydroxybenzoate, propyl hydroxybenzoate, talc, magnesium stearate and / or mineral oil. In addition, the formulation may be prepared by including diluents or excipients such as fillers, extenders, binders, wetting agents, disintegrants and surfactants commonly used in formulations, and in addition to the excipients, a lubricant such as magnesium stearate or talc may be further included.

[0033] The composition of the present invention may be composed of about 0.1% to 90%, preferably about 1% to about 80%, of a pharmacological ingredient. Pharmaceutical preparations for enteral or parenteral administration may be prepared by known methods, for example, conventional mixing, granulating, coating, solubilizing or freeze-drying methods. Pharmaceutical preparations for oral use may be obtained by mixing the active ingredient with a solid excipient, preferably granulating the resulting mixture, and, if necessary or essential, adding suitable auxiliary substances, and then processing the mixture or granules into tablets or coated tablet cores.

[0034] According to one specific example of the present invention, the sodium-glucose cotransporter-2 (SGLT-2) inhibitor may be empagliflozin.

[0035] As used herein, the term "Empagliflozin" refers to (1S)-1,5-Anhydro-1-C-[4-chloro-3-[[4-[[(3S)-tetrahydro-3-furanyl]oxy]phenyl]methyl]phenyl]-D-glucitol.

[0036] A preferred dosage unit form of the SGLT-2 inhibitor may be a tablet or capsule containing from about 5 mg to about 800 mg, preferably from about 50 mg to about 700 mg, more preferably from about 100 mg to about 600 mg, and most preferably from about 100 mg to about 300 mg, administered, for example, once daily.

[0037] According to one specific example of the present invention, the angiotensin II receptor blocker may be telmisartan.

[0038] As used herein, the term "telmisartan" refers to 4′-[(1,4′-dimethyl-2′-propyl[2,6′-bi-1H-benzimidazol]-1′-yl)methyl][1,1′-biphenyl]-2-carboxylic acid.

[0039] Telmisartan may be supplied in a suitable dosage unit form, such as a capsule or tablet, containing a therapeutically effective amount of telmisartan, e.g., about 20 to about 320 mg, that can be administered to a patient. The active ingredient may be administered up to three times daily, starting with a daily dose of 20 mg or 40 mg, increasing to a daily dose of 80 mg, and then increasing to a daily dose of 160 to 320 mg. For example, the dose may be administered in the morning, midday, or evening. In cases of fatty liver disease associated with metabolic abnormalities, once-daily or twice-daily administration is preferred.

[0040] The above empagliflozin or telmisartan may refer to both its active metabolites and prodrugs. The "metabolites" are each active derivative that can be produced when empagliflozin or telmisartan is metabolized, and the "prodrugs" refer to compounds that are metabolized to empagliflozin or telmisartan, or to the same metabolite(s) as empagliflozin or telmisartan. In addition, empagliflozin or telmisartan may include all of its pharmaceutically acceptable salts, crystal forms, hydrates, solvates, diastereomers, or enantiomers.

[0041] The composition of the present invention may additionally comprise vitamin D. A preferred dosage unit form of vitamin D is, for example, a tablet or capsule containing from about 0.5 mg to about 1,200 mg, preferably from about 5 mg to about 600 mg, and the daily dosage is preferably administered once or twice daily.

[0042] According to one specific example of the present invention, the fatty liver disease associated with metabolic dysfunction may be simple steatosis or metabolic dysfunction-associated steatohepatitis.

[0043] Recently, in major liver societies around the world, including the United States and Europe, the names of 'non-alcoholic fatty liver disease (NAFLD)' and 'non-alcoholic steatohepatitis (NASH)' have been changed to 'metabolic dysfunction-associated steatotic liver disease (MASLD)' and 'metabolic dysfunction-associated steatohepatitis (MASH)', respectively.

[0044] Accordingly, the term 'metabolic dysfunction-associated steatotic liver disease (MASLD)' used in the present invention is used with the same meaning as 'non-alcoholic fatty liver disease (NAFLD)'.

[0045] Additionally, 'metabolic dysfunction-associated steatohepatitis (MASH)' is used interchangeably with 'non-alcoholic steatohepatitis (NASH)'.

[0046] According to one specific example of the present invention, the weight ratio of the angiotensin II receptor blocker and the sodium glucose transporter-2 inhibitor may be 2:1 to 8:1.

[0047] If the angiotensin II receptor blocker is included in less than 2 parts by weight or more than 8 parts by weight based on 1 part by weight of the sodium glucose transporter-2 inhibitor, the effect of suppressing intracellular fat accumulation and weight reduction may not be sufficient, and thus the sufficient effect for preventing or treating fatty liver disease related to metabolic disorders may not be achieved.

[0048] Another aspect of the present invention provides a method for preventing or treating fatty liver disease associated with metabolic disorders, comprising administering the pharmaceutical composition to a subject.

[0049] The angiotensin II receptor blocker or a pharmaceutically acceptable salt thereof, and the SGLT-2 inhibitor or a pharmaceutically acceptable salt thereof, included in the pharmaceutical composition of the present invention, may be administered orally in an amount effective for the treatment or prevention of a subject or patient, depending on the purpose. It should be understood that the dosage for administration to a specific subject or patient should be determined based on various related factors such as the patient's body weight, age, race, sex, health condition, diet, administration time, administration method, and severity of the disease, and may be appropriately increased or decreased by a specialist. For example, a physician may start the dosage of the pharmaceutical composition of the present invention at a level lower than that required to achieve the desired therapeutic effect, and gradually increase the dosage until the desired effect is achieved, and may easily determine and prescribe the dosage as needed.

[0050] The method for preventing or treating fatty liver disease related to metabolic disorder of the present invention may be to administer therapeutically effective amounts of each pharmacological component of the composition simultaneously, sequentially, or in any order.

[0051] The corresponding active ingredient or its chemically acceptable salt may be used in the form of a hydrate or may include other solvents used for crystallization.

[0052] Another aspect of the present invention provides a kit comprising the composition.

[0053] The kit of the present invention may comprise an angiotensin II receptor blocker or a pharmaceutically acceptable salt thereof as a first pharmacological component; and a sodium-glucose cotransporter-2 (SGLT-2) inhibitor or a pharmaceutically acceptable salt thereof as a second pharmacological component. The kit may be in the form of a separate pharmacological component administered in the same dosage form or in different dosage forms (e.g., the first pharmacological component is parenteral and the second pharmacological component is oral), and may be particularly advantageous when administered at multiple dosage intervals.

[0054] The present invention will be described in more detail below through one or more examples. However, these examples are intended to exemplify the present invention and the scope of the present invention is not limited to these examples.

[0055]

[0056] Experimental Example 1. Confirmation of the excellent fat accumulation inhibition effect of the empagliflozin and telmisartan combination in cells induced with metabolic abnormalities and fatty liver disease.

[0057] 1-1. Oleic acid-induced fatty liver model and experimental component treatment

[0058] HepG2 cells were treated with oleic acid (OA) to induce a metabolic fatty liver state, and the inhibitory effect on fat accumulation was evaluated in cells in which metabolic fatty liver was induced by treating them with experimental components.

[0059] Specifically, HepG2 cells were cultured in an incubator at 37°C and 5% CO₂ using DMEM medium supplemented with 10% FBS and 1% penicillin-streptomycin. Cells were seeded at 1 × 10 per well in a 6-well plate. 5 The cells were seeded at a density of 100 μm and treated with a medium containing 0.5 mM oleic acid to induce fat accumulation after 24 hours.

[0060] Oleic acid was pre-prepared and used in the form of a complex with fatty acid-free BSA (OA: BSA = 6:1 molar ratio). The medium was replaced with fresh OA medium every day after treatment, and the cells were cultured for a total of 3 days. Simultaneously with the induction of fat accumulation, the experimental groups were treated with the experimental components (single and complex components) at various concentrations (Table 1). After 3 days of culture, the degree of intracellular triglyceride accumulation was evaluated.

[0061]

[0062] Distinctive treatment conditions Empagliflozin (Em, μM) Telmisartan (Telm, μM) Total concentration (μM) A Normal 000 BOA only (0.5 mM, 72h) 000 COA + Em 50 5 DOA + Em 10 010 EOA + Em 20 020 FOA + Telm 05 5 GOA + Telm 01 010 HOA + Telm 02 020 IOA + Em + Telm 55 10 JOA + Em + Telm 51 015 KOA + Em + Telm 52 025 LOA + Em + Telm 10 1020 MOA + Em + Telm 10 2030

[0063]

[0064] 1-2. Quantitative analysis of neutral fat in HepG2 cells using the AdipoRed Assay

[0065] In order to evaluate the effect of inhibiting fat accumulation in oleic acid-induced fatty liver cells treated with the test compound in Experimental Example 1-1, the intracellular neutral fat content was measured using the AdipoRed Assay.

[0066] Specifically, AdipoRed Assay Reagent (Lonza) was used to quantify intracellular triglyceride content. After treatment with the experimental components, the culture medium was removed, the cells were washed twice with PBS, and each well was fixed with 4% paraformaldehyde for 15 minutes at room temperature. After removing the fixative, the cells were washed twice with PBS, and 30 μl of AdipoRed reagent was directly added to each well. The wells were incubated in a darkroom at room temperature for 10 minutes. Fluorescence signals were measured using a multifunctional microplate reader under excitation conditions of 485 nm and emission conditions of 535 nm.

[0067]

[0068] Classification RFU (mean ± standard deviation) Inhibition rate (%) A675 ± 193 - B4, 778 ± 981 - C4, 301 ± 1,023 10.0 D3, 997 ± 990 16.4 E3, 701 ± 957 22.5 F4, 001 ± 886 16.3 G3, 623 ± 623 24.2 H3, 022 ± 610 36.8 I3, 745 ± 54 121.6 J3, 267 ± 477 31.6 K2, 799 ± 31 141.4 L2, 100 ± 153 56.0 M1, 923 ± 168 59.8

[0069]

[0070] As a result, the empagliflozin and telmisartan combination treatment group showed a significantly higher neutral fat suppression effect (p<0.05) compared to the single-component treatment group, and the suppression rate was confirmed to be highest at high concentrations. In addition, the combination treatment group showed a dose-dependent response, in which the suppression effect gradually increased with increasing concentration.

[0071] Through these results, it was confirmed that the empagliflozin and telmisartan combination effectively suppressed oleic acid-induced intrahepatocyte neutral fat accumulation and that there was a synergistic effect between the two components.

[0072]

[0073] 1-3. Evaluation of neutral fat accumulation in HepG2 cells using Oil Red O staining

[0074] In order to evaluate the intracellular neutral fat suppression effect in oleic acid-induced fatty liver cells treated with the experimental component in Experimental Example 1-1, the intracellular accumulated neutral fat was visually evaluated through Oil Red O staining.

[0075] Specifically, for groups A, B, and M in Table 1, the medium was removed after cell culture, the cells were washed twice with PBS, and then fixed with 4% paraformaldehyde solution for 15 minutes at room temperature. After fixation, the cells were washed twice with PBS, pretreated with 60% isopropanol for 5 minutes, and 1 mL of Oil Red O working solution (0.3% Oil Red O in isopropanol, diluted 3:2 with distilled water) was added to each well, followed by staining for 15 minutes at room temperature. Thereafter, the staining solution was removed, washed three times with distilled water to remove excess staining, and observed and images were taken under an optical microscope.

[0076] As a result, in group A, which was the normal control group, intracellular fat accumulation was hardly observed, but in group B, which was treated with only oleic acid, it was confirmed that neutral fat was widely accumulated. On the other hand, it was confirmed that intracellular fat accumulation was significantly reduced in group M, which was treated with a combination of 10 μM empagliflozin and 20 μM telmisartan, compared to group B (Fig. 1).

[0077] Through these results, it was confirmed that the combination of empagliflozin and telmisartan has the effect of inhibiting fat accumulation in cells.

[0078]

[0079] 1-4. Measurement of mRNA expression levels of PPAR-α and PPAR-γ

[0080] In order to investigate the molecular biological mechanism of the metabolic abnormality fatty liver improvement effect in the metabolic abnormality fatty liver induced cells treated with the experimental component in Example 1-1, the relative level of mRNA expression for genes related to intracellular fatty acid synthesis and oxidation was measured using the Real-time PCR method.

[0081] Specifically, for groups A, B, I, J, and M in Table 1, cells were washed twice with PBS, and then total RNA was extracted using an RNA Mini Kit. 1 μg of the extracted RNA was used to synthesize cDNA, and the synthesized cDNA was diluted 1 / 10 and used in qPCR reactions (1 μl each). Real-time PCR was performed with 12.5 μl of SYBR Green Master Mix (2X), 0.5 μl of each primer (10 pmole / μl) in Table 3, and distilled water in a total volume of 25 μl. β-actin was used as a control gene, and the fluorescent signal was quantitatively analyzed using the MAX Pro program.

[0082]

[0083] Primer name Nucleic acid sequence (5'→3') Sequence number PPAR-alphaforwardTCCTGAGCCATGCAGAATTTACSequence number 1reverseAGTCTAAGGCCTCGCTGGTGSequence number 2 PPAR-gammaforwardATTCCATTCACAAGAACAGATCCAGSequence number 3reverseTTTATCTCCACAGACACGACATTCASequence number 4beta-actinforwardCCTGGCACCCAGCACAATSequence number 5reverseGCCGATCCACACACGGAGTACTSequence number 6

[0084]

[0085] As a result, in group B treated with only oleic acid, the expression of PPAR-α mRNA was significantly decreased compared to the normal control group, whereas in groups I, J, and M treated with the combination of empagliflozin and telmisartan, PPAR-α expression was significantly increased (Fig. 2). Meanwhile, PPAR-γ mRNA expression was increased compared to the normal group by oleic acid treatment, but it was confirmed that expression showed a concentration-dependent decrease when treated with the combination of empagliflozin and telmisartan. In particular, the most prominent inhibitory effect was observed in group M (Fig. 3).

[0086] Through these results, it was confirmed that the empagliflozin and telmisartan complex can effectively suppress intracellular neutral fat accumulation by inducing the expression of PPAR-α in a fat accumulation environment, promoting fatty acid oxidation, and simultaneously suppressing the expression of PPAR-γ, thereby limiting fat synthesis.

[0087]

[0088] Experimental Example 2. Confirmation of the optimal combination and formulation of an angiotensin II receptor blocker and sodium glucose transporter-2 complex.

[0089] 2-1. Confirmation of the optimal combination of angiotensin II receptor blocker and sodium glucose transporter-2 combination drug based on drug compatibility testing.

[0090] The optimal combination of an angiotensin II receptor blocker (ARB) and a sodium-glucose cotransporter-2 (SGLT-2) inhibitor that does not decompose and has high stability even after long-term storage was identified by evaluating drug-drug compatibility test.

[0091] Specifically, after storing a mixture of ARB and SGLT-2 inhibitor for 4 weeks under accelerated stability test conditions (40±2℃ / 75±5% relative humidity), the changes in the content and appearance (color, clumping, etc.) of each component and related substances were analyzed. Olmesartan, valsartan, and telmisartan were used as ARBs, and empagliflozin, dapagliflozin propanediol monohydrate, and canagliflozin were used as SGLT-2 inhibitors, and the contents of each component are as shown in Table 4.

[0092]

[0093] Classification Ingredients ARBSGLT-2 InhibitorExample 3 Telmisartan 40 mg Empagliflozin 40 mgExample 4 Telmisartan 40 mg Dapagliflozin 40 mgComparative Example 16 Telmisartan 80 mg - Comparative Example 17 Empagliflozin 80 mgComparative Example 18 Dapagliflozin 80 mgComparative Example 19 Olmesartan 80 mg - Comparative Example 20 Valsartan 80 mg - Comparative Example 21 Canagliflozin 80 mgComparative Example 22 Telmisartan 40 mg Canagliflozin 40 mgComparative Example 23 Olmesartan 40 mg Empagliflozin 40 mgComparative Example 24 Olmesartan 40 mg Dapagliflozin 40 mgComparative Example 25 Valsartan 40 mg Empagliflozin 40 mg Comparative Example 26 Valsartan 40 mg Dapagliflozin 40 mg

[0094]

[0095] In order to analyze the content of the main ingredient and the degree of formation of flexible substances, 10T tablets of a mixture of ARB and SGLT-2 inhibitor stored for 1, 2, 3, and 4 weeks were placed in three 100 mL flasks, and a solution of 0.01 M hydrochloric acid aqueous solution and acetonitrile mixed in an appropriate ratio was added to extract the main ingredient, stirred, and then analyzed by HPLC.

[0096] In order to observe the properties and viscosity of the main ingredients, 10 g of a mixture of ARB and SGLT-2 inhibitor stored for 1 week, 2 weeks, 3 weeks, and 4 weeks was taken and exposed to 100 ml of purified water at room temperature for 10 minutes, and then the properties and viscosity were checked.

[0097] As a result, in the 3rd week of acceleration, all indices were confirmed to be unsuitable in Comparative Examples 22 to 26 among the mixtures of ARB and SGLT-2 inhibitors, whereas the contents of telmisartan and empagliflozin (Example 3) and the mixture of telmisartan and dapagliflozin (Example 4) were 98.6% / 92.1% and 98.9 / 99.0%, respectively, showing content changes of less than 10% in all, and the total amount of flexible substances was also found to be appropriate, confirming that the combination of drugs was the most appropriate. However, the properties were found to be unsuitable from the 3rd week of acceleration, confirming that a formulation capable of maintaining the properties was necessary (Table 5).

[0098]

[0099] Classification Test Item Initial Acceleration 1 week Acceleration 2 weeks Acceleration 3 weeks Acceleration 4 weeks Example 3 Content 99.9 / 100 99.5 / 97.6 99.4 / 95.1 99.2 / 92.199 / 89.8 Flexible material 0.1 or less 1.0 or more 1.0 or more 1.0 or more 5.0 or more Property Suitable Suitable Suitable Unsuitable Unsuitable Example 4 Content 99.9 / 100.1 99.4 / 99.999.2 / 99.2 98.9 / 9998.6 / 98.9 Flexible material 0.1 or less 0.1 or more 0.1 or more 1.0 or less 1.0 or more Property Suitable Suitable Suitable Unsuitable Unsuitable Comparative Example 16 Content 100.1 100.1 100 100 100 Flexible material 0.1 or less 0.1 or less 0.1 or less 0.1 or less 0.1 or less 0.1 Below Property Suitable Suitable Suitable Suitable Suitable Comparison Example 17 Content 99.997.395.185.480.2 Flexible material 0.1 or less 1.0 or less 1.0 or more 5.0 or more 5.0 or more Property Suitable Suitable Suitable Unsuitable Unsuitable Comparison Example 18 Content 100.3100.1100.299.899 Flexible material 0.1 or less 0.1 or less 0.1 or less 0.1 or less 1.0 or less Property Suitable Suitable Suitable Unsuitable Unsuitable Comparison Example 19 Content 99.999.799.799.599.4 Flexible material 0.1 or less 0.1 or more 0.1 or more 0.1 or more Property Suitable Suitable Suitable Suitable Suitable Comparison Example 20 Content 99.799.499.29998.7 Flexible material 0.1 or less 0.1 Above 0.1 Above 0.1 Above 0.1 Above Property Suitable Suitable Suitable Suitable Comparison Example 21 Content 99.5 95.28 9.28 5.28 0.1 Flexible material 0.1 or less 1.0 or less 5.0 or more 5.0 or more 5.0 or more 5.0 or more Above 5.0 Above Property Suitable Unsuitable Unsuitable Unsuitable Comparison Example 22 Content 99.5 / 92.1 99.1 / 89.1 98.9 / 82.3 94.5 / 74.4 92.3 / 62.3 Flexible material 0.1 or less 5.0 or more 5.0 or more 5.0 or more Above Property Suitable Unsuitable Unsuitable Unsuitable Comparison Example 23 Content 100.1 / 99.99 9.1 / 90 95.4 / 84.6 92.3 / 72.8 90.1 / 60.5 Flexible material 0.1 or less 5.0 Above 5.0 Above 5.0 Above 5.0 Above Property Suitable Suitable Unsuitable Unsuitable Comparison Example 24 Content 100.1 / 100 97.8 / 89.195 / 79.79 1.9 / 69.9 89.9 / 60 Flexible Material 0.1 Below 5.0 Above 5.0 Above 5.0 Above 5.0 Above Property Suitable Suitable Unsuitable Unsuitable Comparison Example 25 Content 100.2 / 99.99 5.5 / 95.493.3 / 90.190.8 / 85.590 / 80.7Flexible material 0.1 or less 1.0 or less 5.0 or more 5.0 or more 5.0 or more Property Suitable Inappropriate Inappropriate Inappropriate Inappropriate Comparison Example 26 Content 100.3 / 99.795.5 / 95.791.1 / 92.389.1 / 87.987.1 / 81.1Flexible material 0.1 or less 1.0 or more 1.0 or more 5.0 or more 5.0 or more Property Suitable Inappropriate Inappropriate Inappropriate Inappropriate Inappropriate.

[0100]

[0101] 2-2. Manufacturing of a bilayer tablet containing angiotensin II receptor blocker and sodium glucose transporter-2 using the direct tableting process.

[0102] The ARB granule layer according to Table 6 was manufactured through wet granulation, and the granules were manufactured using a semi-aqueous method with the main ingredient, mannitol, meglumine, and sodium hydroxide as a solubilizing agent, and then dried to manufacture the granules. All the components of the ARB layer and the SGLT-2 inhibitor layer, except for the lubricants sodium stearyl fumarate and magnesium stearate, were sieved through a 20-mesh standard sieve, and then pre-mixed with each layer composition for the first time and mixed for at least 10 minutes using a turbula mixer. Thereafter, the lubricant was added to each powder mixture and mixed again for 5 minutes. Finally, the mixture forming each layer was injected into each hopper of a two-layer tablet press, compressed, and then coated with a coating substrate to manufacture a two-layer tablet. Olmesartan, valsartan, and telmisartan were used as ARBs, and empagliflozin, dapagliflozin propanediol monohydrate, and canagliflozin were used as SGLT-2 inhibitors. The combination of each ingredient is as shown in Table 7.

[0103]

[0104] Classification Functional Ingredients Weight Part ARB Layer API ARB Granules 16.8 Excipient Mannitol 28.2 Lubricant Sodium stearyl fumarate and magnesium stearate 2.5 SGLT-2 Inhibitor Layer API SGLT-2 Inhibitor 2.6 Excipient Mannitol 42.5 Disintegrant Polyvinylpyrrolidone 6.3 Lubricant Magnesium stearate 1.1 Total 100

[0105]

[0106] Distinctive ingredient ARBSGLT-2 inhibitor Example 5 Telmisartan empagliflozin Example 6 Telmisartan dapagliflozin Comparative example 27 Telmisartan canagliflozin Comparative example 28 Valsartan empagliflozin Comparative example 29 Valsartan dapagliflozin Comparative example 30 Olmesartan empagliflozin Comparative example 31 Olmesartan dapagliflozin

[0107]

[0108] 2-3. Confirmation of the optimal combination of angiotensin II receptor blocker and sodium glucose transporter-2 complex formulation based on formulation stability analysis.

[0109] By evaluating the stability of the formulation, the optimal combination of an angiotensin II receptor blocker (ARB) and a sodium-glucose cotransporter-2 (SGLT-2) inhibitor formulation with high stability and no degradation even after long-term storage was identified.

[0110] Specifically, the ARB and SGLT-2 inhibitor composite bilayer tablet manufactured in Experimental Example 2-2 was stored for 6 months under accelerated stability test conditions (40±2℃ / 75±5% relative humidity) or long-term stability test conditions (25±2℃ / 60±5% relative humidity), and then the changes in the content of each component and flexible substance, changes in appearance (color, agglomeration, etc.), and changes in dissolution (interference, changes in dissolution, etc.) were analyzed.

[0111] Changes in the content of each component and flexible substance, and changes in appearance (color, clumping, etc.) were analyzed using the same method as in Experimental Example 2-1, and analysis of dissolution changes was performed using the method in Table 8 for the ARB and SGLT-2 inhibitor composite two-layer tablet manufactured in Experimental Example 2-2.

[0112]

[0113] Dissolution test conditions Dissolution method Dissolution test method 2 of the Korean Pharmacopoeia (paddle method) Dissolution solution pH 1.2 hydrochloric acid (HCl) solution Dissolution amount 900 ml Dissolution apparatus temperature 37.5 ℃ Paddle speed 50 rpm Test group 4 Sample collection time 5, 10, 15, 30, 45, 60, 90, 120 min Sample analysis conditions Detector Ultraviolet spectrophotometer (measurement wavelength: 229 nm) Column C18 (5 μm, 4.6 × 250 mm) Mobile phase Weigh 2.0 g of ammonium dihydrogen phosphate, dissolve in water to make exactly 1.0 L, and then use a solution prepared by mixing 300 mL of a solution adjusted to pH 3.0 with 1 mol / L phosphoric acid and 700 mL of methanol. Flow rate 0.7 ml / min Column temperature 35 ℃

[0114]

[0115] As a result, most of the indices of the ARB and SGLT-2 inhibitor combination bilayer tablets in Comparative Examples 27 to 31 were found to be unsuitable at accelerated 6-month and long-term 6-month treatment, whereas the telmisartan and empagliflozin combination bilayer tablet (Example 5) showed all indices to be suitable, confirming excellent formulation stability (Table 9). In particular, the properties that were found to be unsuitable in the drug-to-drug compatibility test were also confirmed to be suitable after being formulated as a combination bilayer tablet.

[0116]

[0117] Category Test Item Initial Acceleration 1 month Acceleration 3 months Acceleration 6 months Long-term 1 month Long-term 3 months Long-term 6 months Example 5 Content 100.3% 100.1% 100.1% 99.7% 100.3% 99.9% 100.1% Flexible substance Total 0.1% or less Total 0.1% or less Total 0.15% or less Total 0.2% or less Total 0.1% or less Total 0.1% or less Property Suitable Suitable Suitable Suitable Suitable Suitable Suitable Suitable Elution Suitable Suitable Suitable Suitable Suitable Suitable Example 6 Content 100.1% 99.9% 100.0% 99.8% 100.1% 100.1% 100.2% Flexible substance Total 0.1% or less Total 0.1% or less Total 0.1% or less Total 0.2% or less Total 0.1% or lessTotal 0.1% or lessTotal 0.1% or lessPropertySuitableSuitableSuitableSuitableSuitableElutionSuitableSuitableSuitableSuitableComparative Example 27Content99.8%95.0%90.1%82.5%99.0%96.9%94.3%Flexible SubstanceTotal 0.1% or lessTotal 1.0% or lessTotal 3.0% or lessTotal 5% or moreTotal 0.1% or lessTotal 1.0% or lessTotal 1.0% or lessPropertySuitableNotSuitableNotSuitableNotSuitableNotSuitableElutionSuitableNotSuitableNotSuitableNotSuitableNotSuitableComparative Example 28Content99.9%97.4%89.7%80.4%99.2%95.4%95.0%Flexible SubstanceTotal 0.1% or lessTotal 1.0% or lessTotal 5% or moreTotal 5% or moreTotal 0.1% or lessTotal 1.0% or lessTotal 3.0% or lessPropertySuitableSuitableUnsuitableUnsuitableUnsuitableUnsuitableUnsuitableDissolutionSuitableUnsuitableUnsuitableUnsuitableUnsuitableUnsuitableComparison Example 29Content 100.2%100.1%87.7%79.5%98.6%94.8%95.3%Flexible SubstanceTotal 0.1% or lessTotal 0.1% or lessTotal 5% or moreTotal 5% or moreTotal 0.1% or lessTotal 1.0% or lessTotal 3.0% or lessPropertySuitableUnsuitableUnsuitableUnsuitableUnsuitableDissolutionSuitableUnsuitableUnsuitableUnsuitableUnsuitableUnsuitableComparison Example 30 Content 100.1% 98.7% 97.7% 95.8% 99.5% 98.5% 97.3% Flexible Substance Total 0.1% or less Total 0.1% or less Total 1.0% or less Total 3.0% or less Total 0.1% or less Total 1.0% or less Total 1.0% or less Abnormal Properties Suitable Suitable Suitable Unsuitable Unsuitable Unsuitable Unsuitable Dissolution Suitable Suitable Unsuitable Unsuitable Unsuitable Unsuitable Comparison Example 31 Content 99.9% 98.9% 98.1% 94.3% 99.6% 98.9% 97.9% Flexible Substance Total 0.1% or less Total 0.Less than 1% Total Less than 1.0% Total Less than 3.0% Total Less than 0.1% Total Less than 1.0% Total Above 1.0% Characteristics Suitable Suitable Suitable Unsuitable Unsuitable Unsuitable Unsuitable Dissolution Suitable Suitable Unsuitable Unsuitable Unsuitable Unsuitable Unsuitable.

[0118]

[0119] The present invention has been described above, focusing on specific embodiments thereof. Those skilled in the art will appreciate that the present invention can be implemented in modified forms without departing from its essential characteristics. Therefore, the disclosed embodiments should be considered illustrative rather than limiting. The scope of the present invention is set forth in the claims, not the foregoing description, and all differences within the scope equivalent thereto should be construed as being encompassed by the present invention.

Claims

1. Angiotensin II receptor blocker or a pharmaceutically acceptable salt thereof as the first pharmacological ingredient; and As a second pharmacological ingredient, a sodium-glucose cotransporter-2 (SGLT-2) inhibitor or a pharmaceutically acceptable salt thereof A pharmaceutical composition for preventing or treating metabolic dysfunction-associated steatotic liver disease.

2. A pharmaceutical composition for preventing or treating fatty liver disease related to metabolic disorder, wherein the sodium-glucose cotransporter-2 (SGLT-2) inhibitor in paragraph 1 is empagliflozin.

3. A pharmaceutical composition for preventing or treating fatty liver disease related to metabolic disorder, wherein the angiotensin II receptor blocker in paragraph 1 is telmisartan.

4. A pharmaceutical composition for preventing or treating fatty liver disease associated with metabolic dysfunction, wherein the fatty liver disease associated with metabolic dysfunction in the first paragraph is simple steatosis or metabolic dysfunction-associated steatohepatitis.

5. A pharmaceutical composition for preventing or treating fatty liver disease related to metabolic disorder, wherein the weight ratio of the angiotensin II receptor blocker and the sodium glucose transporter-2 inhibitor in paragraph 1 is 2:1 to 8:

1.

6. A method for preventing or treating fatty liver disease associated with metabolic disorders, comprising administering to a subject the pharmaceutical composition of claim 1.

7. A kit comprising the composition of paragraph 1.

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