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WO2023182735A1 - Composition for preventing or treating macular degeneration, comprising compound that induces expression of anti-aging gene klotho - Google Patents

Composition for preventing or treating macular degeneration, comprising compound that induces expression of anti-aging gene klotho Download PDF

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Publication number
WO2023182735A1
WO2023182735A1 PCT/KR2023/003592 KR2023003592W WO2023182735A1 WO 2023182735 A1 WO2023182735 A1 WO 2023182735A1 KR 2023003592 W KR2023003592 W KR 2023003592W WO 2023182735 A1 WO2023182735 A1 WO 2023182735A1
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formula
macular degeneration
composition
compound represented
preventing
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PCT/KR2023/003592
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French (fr)
Korean (ko)
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정동주
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주식회사 클로소사이언스
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Priority claimed from KR1020220187495A external-priority patent/KR20230137816A/en
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Publication of WO2023182735A1 publication Critical patent/WO2023182735A1/en

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    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES, NOT OTHERWISE PROVIDED FOR; PREPARATION OR TREATMENT THEREOF
    • A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
    • A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/41—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
    • A61K31/42—Oxazoles
    • A61K31/423—Oxazoles condensed with carbocyclic rings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P27/00—Drugs for disorders of the senses
    • A61P27/02—Ophthalmic agents

Definitions

  • the present invention relates to a composition for preventing or treating macular degeneration containing a compound that induces the expression of the anti-aging gene klotho.
  • the macula is located in the center of the retina at the back of the eye, where images of objects are formed, and is where visual cells are concentrated.
  • Age-related macular degeneration occurs when the macular area degenerates. Lack of oxygen is known to be the main cause of degeneration, and as a result, Vascular Endothelial Growth Factor (VEGF) increases and new blood vessels are formed in areas of the retina where there were no blood vessels, resulting in retinal degeneration.
  • VEGF Vascular Endothelial Growth Factor
  • the outer layer, where the retina's visual cells are gathered, is surrounded by polarized epithelial cells called retinal pigment epithelium (RPE).
  • RPE retinal pigment epithelium
  • RPE cells The apical side of these RPE cells is in direct contact with the visual cells, and the basal side is in contact with Burch's membrane located inside the choroid layer.
  • the function of RPE cells plays an important role in maintaining vision. Its function is to remove old visual cells and move waste materials discharged from visual cells to the choroid for cleaning.
  • Age-Related Macular Degeneration is a disease that occurs due to aging. It is a disease with a high incidence in the elderly, with one in three adults over the age of 70 suffering from symptoms. Age-related macular degeneration is a representative eye disease with a higher occurrence frequency than the combined incidence of other eye diseases, glaucoma and cataract. If the disease continues to progress, vision is lost.
  • Age-related macular degeneration occurs due to loss of RPE cells and visual cells in the macula.
  • the histological characteristic of age-related macular degeneration disease is the accumulation of crystals called drusen between the RPE cell layer and Burch's membrane. Most of the components of these crystals are formed through oxidative stress and inflammation. Therefore, oxidative stress or inflammation can be considered to be the cause of age-related macular degeneration.
  • the treatment for this mainly uses laser or drug administration to remove malignant blood vessels. However, this does not provide a fundamental treatment and is only a way to alleviate the worsening symptoms.
  • mice that later increased the expression of this gene had an increase in lifespan of 20.0 to 30.8% in males and 18.8 to 19.0% in females. This served as an opportunity to inform the world for the first time that the lifespan of mice can be increased or decreased depending on the expression of a single gene.
  • the base sequence of the Klotho gene is very similar between animals, and it has been reported that there is about 98% identity between mice and humans. This indicates that even in humans, lifespan can be regulated by the expression of the klotho gene.
  • the klotho gene is located on chromosome 13 and produces a membrane protein with a base sequence similar to that of ⁇ -glucosidase.
  • Klotho protein is mainly expressed in renal tubular epithelial cells and brain choroid plexus, and has been reported to be expressed in some parathyroid glands.
  • the Klotho gene is a gene involved in various aging phenotypes. In mice deficient in the klotho gene, the aging process includes reduced lifespan, reduced activity, growth retardation, atherosclerosis, arterial calcification, osteoporosis, reproductive immaturity, infertility, skin atrophy, and emphysema. A similar syndrome occurs.
  • Klotho mRNA expression is significantly higher in kidney tissue than in other tissues, but expression of klotho mRNA is reduced in the kidneys of mouse disease models for hypertension, type 2 diabetes, diabetic nephropathy, and chronic renal failure.
  • NO Nitric Oxide
  • OLETF Oleuka Long-Evans Tokushima fatty rats
  • the klotho gene also affects glucose and insulin metabolism in mice, and statins, a representative hypercholesterolemia treatment, increase the expression of klotho mRNA in kidney proximal tubule cells.
  • statins a representative hypercholesterolemia treatment
  • osteopenia with low bone turnover occurs due to impaired differentiation of both osteoblasts and osteoclasts, which is similar to the characteristics of age-related bone loss and senile osteoporosis in humans.
  • Klotho mutant mice abnormal elongation of trabecular bone in the epiphyseal region and abnormal trabecular bone tissue findings are observed in microcomputed tomography, which is due to a disruption in the bone resorption process.
  • KL-VS functional variant of klotho
  • klotho which has mutations in three regions of the Klotho gene exon2
  • lipid metabolism blood pressure, lifespan, cognitive function, coronary artery disease, and cerebrovascular disease
  • microsatellites of the Klotho gene polymorphisms and single nucleotide polymorphisms have been associated with bone mineral density, and it has also been reported that single nucleotide polymorphisms in the Klotho gene are associated with risk factors for cardiovascular disease and bone mineral density in healthy adult women.
  • Recently, the association between the Klotho gene and Alzheimer's has been reported in several papers.
  • the present inventor has continuously conducted research on compounds that induce klotho expression, and as a result, the new compound developed by the present inventor has excellent stability and has a protective effect against oxidative stress (oxidative toxicity) in retinal pigment epithelial cells (RPE). It was experimentally confirmed that it was excellent, and the present invention was completed.
  • RPE retinal pigment epithelial cells
  • the purpose of the present invention is to provide a pharmaceutical composition for preventing or treating macular degeneration containing the compound represented by Formula 1 or a pharmaceutically acceptable salt thereof as an active ingredient.
  • Another object of the present invention is to provide a food composition for preventing or improving macular degeneration containing the compound represented by Formula 1 or a pharmaceutically acceptable salt thereof as an active ingredient.
  • the present invention provides a pharmaceutical composition for preventing or treating macular degeneration, comprising a compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient.
  • the present invention provides a food composition or health functional food composition for preventing or improving macular degeneration, comprising a compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient.
  • R 1 and R 2 are each -H, halogen or C 1-6 straight or branched alkyl
  • R 3 and R 4 may each be -H or halogen.
  • the compound represented by Formula 1 may be a compound represented by Formula 1-1 below.
  • R 1 and R 2 may each be -H, halogen, or C 1-6 straight or branched alkyl.
  • the composition can increase the expression of the Klotho gene.
  • the composition can protect retinal pigment epithelial cells from oxidative stress.
  • the present invention provides a method for preventing or treating macular degeneration, comprising administering to a subject a composition containing the compound represented by Formula 1 or a pharmaceutically acceptable salt thereof as an active ingredient.
  • the present invention provides a use for preventing or treating macular degeneration of a composition containing the compound represented by Formula 1 or a pharmaceutically acceptable salt thereof as an active ingredient.
  • the compound according to the present invention has the effect of improving the expression level of the Klotho gene, a gene related to aging, and has a protective effect against oxidative stress in retinal pigment epithelial cells (RPE), and is a pharmaceutical composition for preventing or treating macular degeneration. , or can be usefully used as a health functional food composition.
  • RPE retinal pigment epithelial cells
  • Figure 1 shows the results of confirming the stability of KS compounds according to Example 1 of the present invention by MS/MS analysis: KS101 (compound of Formula 1-1A), KS102 (compound of Formula 1-1B), KS103 (compound of Formula 1-1C) ), KS104 (Formula 1-1D compound).
  • Figure 2 shows the results of confirming the expression level of secreted klotho and membrane klotho genes by the KS compound according to Example 1 of the present invention by PCR.
  • Figure 3 shows the results of confirming changes in survival rate in cells (ARPE: retinal pigment epithelial cells) treated with the KS compound for a long time according to Example 1 of the present invention.
  • FIG 4 shows the results of confirming changes in reactive oxygen species (ROS) in cells (ARPE: retinal pigment epithelial cells) treated with the KS compound for a long time according to Example 1 of the present invention.
  • ROS reactive oxygen species
  • FIG. 5 shows the results of confirming changes in reactive oxygen species (ROS) in cells (ARPE: retinal pigment epithelial cells) treated with the KS compound for a short period of time (6 hours) according to Example 1 of the present invention.
  • ROS reactive oxygen species
  • Figure 6 shows the results of confirming the cytotoxicity of the KS compound according to Example 1 of the present invention.
  • Figure 7 shows the results confirming the cell protection effect of KS compounds from oxidative stress induced by BSO.
  • Figure 8 shows the results confirming the cell protection effect of KS compounds from oxidative stress induced by NaIO 3 .
  • the present invention provides a composition for preventing, improving or treating macular degeneration comprising a compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient.
  • R 1 and R 2 may each be -H, halogen, or C 1-6 straight or branched alkyl, and R 3 and R 4 may each be -H or halogen.
  • the compound represented by Formula 1 may be a compound represented by Formula 1-1 below.
  • R 1 and R 2 may each be -H, halogen, or C 1-6 straight or branched alkyl.
  • the compound represented by Formula 1-1 is:
  • the macular degeneration may be age-related macular degeneration.
  • composition of the present invention can increase the expression level of the Klotho gene.
  • composition of the present invention can protect retinal pigment epithelial cells from oxidative stress.
  • the composition may be a pharmaceutical composition, food composition, or health functional food composition.
  • the composition may contain 1 to 99% by weight, 5 to 95% by weight, or 10 to 90% by weight of the active ingredient based on the total weight of the composition, but is not limited thereto.
  • prevention refers to any action that delays the onset of macular degeneration by administration of the composition of the present invention
  • treatment and “improvement” refers to the improvement or beneficial change in symptoms of macular degeneration by administration of the composition of the present invention. It means all actions.
  • the compound of the present invention can be used in the form of a pharmaceutically acceptable salt, and an acid addition salt formed by a pharmaceutically acceptable free acid is useful as the salt.
  • pharmaceutically acceptable salt refers to any organic or organic salt of the base compound of Formula 1 where side effects due to the salt do not reduce the beneficial effects of the base compound of Formula 1, at a concentration that is relatively non-toxic and harmless to the patient and has an effective effect. It means inorganic addition salt. For these salts, inorganic acids and organic acids can be used as free acids.
  • Hydrochloric acid, hydrobromic acid, nitric acid, sulfuric acid, perchloric acid, and phosphoric acid can be used as inorganic acids, and citric acid, acetic acid, lactic acid, maleic acid, and fumarine can be used as organic acids.
  • Acids gluconic acid, methanesulfonic acid, glyconic acid, succinic acid, tartaric acid, galacturonic acid, embonic acid, glutamic acid, aspartic acid, oxalic acid, (D) or (L) malic acid, maleic acid, methanesulfonic acid, ethane sulfuric acid Fonic acid, 4-toluenesulfonic acid, salicylic acid, citric acid, benzoic acid, or malonic acid can be used.
  • these salts include alkali metal salts (sodium salts, potassium salts, etc.) and alkaline earth metal salts (calcium salts, magnesium salts, etc.).
  • acid addition salts include acetate, aspartate, benzate, besylate, bicarbonate/carbonate, bisulfate/sulfate, borate, camsylate, citrate, edisylate, esylate, formate, fumarate, Gluceptate, gluconate, glucuronate, hexafluorophosphate, hybenzate, hydrochloride/chloride, hydrobromide/bromide, hydroiodide/iodide, isethionate, lactate, maleate, maleate , malonate, mesylate, methyl sulfate, naphthylate, 2-naphsylate, nicotinate, nitrate, orotate, oxalate, palmitate, pamoate, phosphate/hydrogen phosphate/dihydrogen phosphate, saccharate.
  • stearate, succinate, tartrate, tosylate, trifluoroacetate aluminum, arginine, benzathine, calcium, choline, diethylamine, diolamine, glycine, lysine, magnesium, meglumine, olamine, potassium, Sodium, tromethamine, zinc salt, etc. may be included, and among these, hydrochloride or trifluoroacetate is preferable.
  • the compound of the present invention includes not only pharmaceutically acceptable salts, but also all salts, isomers, hydrates and solvates that can be prepared by conventional methods.
  • the addition salt according to the present invention can be prepared by conventional methods.
  • the compound is dissolved in a water-miscible organic solvent, such as acetone, methanol, ethanol, or acetonitrile, and an excess amount of organic acid is added or an aqueous acid solution of an inorganic acid is added. It can be manufactured by adding and then precipitating or crystallizing. Then, the solvent or excess acid in this mixture is evaporated and dried to obtain an addition salt, or the precipitated salt can be prepared by suction filtration.
  • a water-miscible organic solvent such as acetone, methanol, ethanol, or acetonitrile
  • the compounds of the present invention can be usefully used as a preventive, ameliorating, or therapeutic agent for macular degeneration disease. .
  • compositions of the present invention can be prepared as pharmaceutical compositions.
  • the pharmaceutical composition of the present invention may include a pharmaceutically acceptable carrier.
  • the composition of the present invention comprises (a) a pharmaceutically effective amount of the above-described compound of the present invention or a pharmaceutically acceptable salt thereof; and (b) a pharmaceutically acceptable carrier.
  • pharmaceutically effective amount refers to an amount sufficient to achieve the efficacy or activity of the above-described compound of the present invention or a pharmaceutically acceptable salt thereof.
  • Pharmaceutically acceptable carriers are those commonly used in preparation, such as lactose, dextrose, sucrose, sorbitol, mannitol, starch, gum acacia, calcium phosphate, alginate, gelatin, calcium silicate, microcrystalline cellulose, poly Includes, but is not limited to, vinylpyrrolidone, cellulose, water, syrup, methyl cellulose, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and mineral oil.
  • the pharmaceutical composition of the present invention may further include lubricants, wetting agents, sweeteners, flavoring agents, emulsifiers, suspending agents, preservatives, etc. Suitable pharmaceutically acceptable carriers and formulations are described in detail in Remington's Pharmaceutical Sciences (19th ed., 1995).
  • the administration method of the pharmaceutical composition of the present invention is not particularly limited and may follow methods commonly used in the art. More specifically, the pharmaceutical composition of the present invention can be administered in various oral and parenteral dosage forms during clinical administration, and when formulated, commonly used fillers, extenders, binders, wetting agents, disintegrants, surfactants, etc. It may be prepared using diluents or excipients.
  • the pharmaceutical composition may be formulated in any one dosage form selected from the group consisting of eye drops, injections, granules, tablets, troches, pills, capsules, gels, syrups, suspensions, emulsions, drops, and solutions.
  • Solid preparations for oral administration include tablets, pills, powders, granules, capsules, troches, etc. These solid preparations include one or more compounds of the present invention and at least one excipient such as starch, calcium carbonate, water, etc. It is prepared by mixing sucrose, lactose, or gelatin. Additionally, in addition to simple excipients, lubricants such as magnesium styrate talc are also used.
  • Liquid preparations for oral administration include suspensions, oral solutions, emulsions, or syrups. In addition to the commonly used simple diluents such as water and liquid paraffin, they contain various excipients such as wetting agents, sweeteners, fragrances, and preservatives. You can.
  • Preparations for parenteral administration include sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, suppositories, etc.
  • Non-aqueous solvents and suspensions may include propylene glycol, polyethylene glycol, vegetable oil such as olive oil, and injectable ester such as ethyl oleate.
  • injectable ester such as ethyl oleate.
  • As a base for suppositories witepsol, macrogol, tween 61, cacao, laurel, glycerol, gelatin, etc. can be used.
  • parenteral administration it can be administered by injection in the form of intravenous injection, subcutaneous injection, intramuscular injection, intraperitoneal injection, transdermal administration, ocular administration, or topical ocular administration.
  • Ocular topical administration for example, can be administered directly intraocularly, periocularly, retrobulbally, subretinal, central retinal, outside the fovea, subconjunctival, or intravitreous. ), intracameral, or suprachoroidal administration.
  • the effective dosage for the human body of the composition of the present invention may vary depending on the patient's age, weight, gender, dosage form, health condition, and degree of disease, and the amount of the active ingredient contained in the composition of the present invention is generally about 0.001-100 mg/kg/day, preferably 0.01-35 mg/kg/day. Based on an adult patient weighing 70 kg, the dose is generally 0.07-7000 mg/day, preferably 0.7-2500 mg/day, and is administered once a day at regular intervals depending on the judgment of the doctor or pharmacist. It may be administered in several divided doses.
  • composition of the present invention may be provided as a food composition or health functional food composition.
  • Examples of foods to which the active ingredient of the present invention can be added include drinks, meat, sausages, bread, biscuits, rice cakes, chocolate, candies, snacks, confectionery, pizza, ramen, other noodles, gum, dairy products including ice cream, There are various soups, beverages, alcoholic beverages, vitamin complexes, dairy products and milk products, and it includes all foods, health foods and health functional foods in the conventional sense.
  • Food and health functional food compositions containing the active ingredients according to the present invention can be added as is to food or used together with other foods or food ingredients, and can be used appropriately according to conventional methods.
  • the mixing amount of the active ingredient can be appropriately determined depending on the purpose of use (prevention or improvement).
  • the amount of the active ingredient of the present invention may be added in an amount of 0.1 to 90 parts by weight based on the total weight of the food (based on 100 parts by weight in total).
  • the amount may be below the above range, and since there is no problem in terms of safety, the active ingredient may be used in an amount above the above range.
  • the food and health functional food composition of the present invention has no particular restrictions on other ingredients other than containing the active ingredient of the present invention as an essential ingredient in the indicated ratio, and contains various flavoring agents or natural carbohydrates as additional ingredients like a conventional beverage. can do.
  • natural carbohydrates include monosaccharides such as glucose, fructose, etc.; Disaccharides such as maltose, sucrose, etc.; and polysaccharides, such as common sugars such as dextrin and cyclodextrin, and sugar alcohols such as xylitol, sorbitol, and erythritol.
  • natural flavoring agents thaumatin, stevia extract (e.g., rebaudioside A, glycyrrhizin, etc.)
  • synthetic flavoring agents sacharin, aspartame, etc.
  • the ratio of the natural carbohydrate is generally about 1 to 20 g, preferably about 5 to 12 g, per 100 g of the food or health functional food composition of the present invention.
  • food and health functional food compositions containing the active ingredients of the present invention include various nutrients, vitamins, minerals (electrolytes), flavoring agents such as synthetic and natural flavors, colorants and enhancers (cheese, chocolate, etc.), It may contain pectic acid and its salts, alginic acid and its salts, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohol, carbonating agents used in carbonated beverages, etc.
  • the food and health functional food composition of the present invention may contain pulp for the production of natural fruit juice, fruit juice drinks, and vegetable drinks.
  • the ratio of these additives is not that important, but is generally selected in the range of 0.1 to about 20 parts by weight per 100 parts by weight of the food and health functional food composition containing the active ingredient of the present invention.
  • Step 1 1-(3,4-difluorophenyl)-3-(2-hydroxyphenyl)thiourea (1-(3,4-difluorophenyl)-3-(2-hydroxyphenyl)thiourea ,FCCS-19025 -2-1) Manufacturing
  • reaction mixture was purified by silica-gel column chromatography (20% Acetone / n-hexane) to obtain 1-(3,4-difluorophenyl)-3-(2-hydroxyphenyl)thiourea (1 -(3,4-difluorophenyl)-3-(2-hydroxyphenyl)thiourea, FCCS-19025-2-1) was obtained as a pale yellow solid (354 mg, 92%).
  • Step 2 N-(3,4-difluorophenyl)benzo[d]oxazol-2-amine (FCCS-19025) manufacturing
  • FCCS-19025-2-1 (224 mg, 0.80 mmol) and potassium peroxide (KO 2 ) (284 mg, 4.00 mmol) obtained in step 1 were mixed with acetonitrile (MeCN) (25) under Ar gas atmosphere. mL) and stirred at room temperature for 14 hours. After confirming the completion of the reaction by TLC (thin layer chromatography), acetonitrile (MeCN) was removed under reduced pressure.
  • reaction mixture was purified by silica-gel column chromatography (10 ⁇ 20% Ethyl acetate / n-hexane) to obtain the target compound N-(3,4-difluorophenyl)benzo[d]oxazole-2- Amine (N-(3,4-difluorophenyl)benzo[d]oxazol-2-amine, FCCS-19025) was obtained as a white solid (160 mg, 82%).
  • 3,4-difluorophenyl isothiocyanate (210mg, 1.23mmol) and 2-amino-5-fluorophenol (2-amino-5) obtained in Example 1-2 -fluorophenol, 130 mg, 1.02 mmol) was reacted to produce KS103 compound ( N -(3,4-difluorophenyl)-6-fluorobenzo[ d ]oxazol-2-amine), represented by the following formula 1-1C, as an ivory solid (203). mg, 75%).
  • 3,4-difluorophenyl isothiocyanate (212 mg, 1.24 mmol) and 2-amino-4,5-fluorophenol (2- amino-4,5-difluorophenol, 150 mg, 1.03 mmol) was reacted to produce KS104 compound ( N -(3,4-difluorophenyl)-5,6-difluorobenzo[ d ]oxazol-2, represented by the following formula 1-1D -amine) was obtained as a light purple solid (187 mg, 64%).
  • KS compound in Example 1 was used dissolved in 10mM DMSO, and tolbutamide, used as a standard substance, was dissolved in methanol at a concentration of 10mM.
  • each KS compound solution in 50 ⁇ l to a final concentration of 1 ⁇ M, pre-incubate for 3 minutes in a shaking water bath (37°C, 40 rpm), and then add 25 ⁇ l of 10 mM NADPH. Incubation was performed for 30 minutes under conditions with or without addition.
  • Figure 1 shows quantitative confirmation of KS compounds cultured with human liver microsomes through MS/MS analysis. Microsome enzymes operate dependent on the coenzyme NADPH(+), so compound metabolism occurred only in reactions where NADPH(+) was added. KS compounds showed higher stability than tolbutamide used as a control, and in particular, KS104 was confirmed to have the best microsomal stability.
  • KS101, KS102, KS103, KS104 compounds of Example 1 was added to Retinal Pigment Epithelium (RPE cells), whose aging was accelerated by adding BSO (Buthionine sulfoximine) at a concentration of 1 ⁇ M to the medium during the culture process. ) were added at 2.5 ⁇ M each and cultured for 8 hours. All RNA expressed in the cells was collected, and changes in expression of only klotho mRNA were selectively confirmed through PCR. As a result of the experiment, all KS compounds induced an increase in the expression of secreted klotho and membrane klotho. In particular, KS104 had a higher ability to induce secreted klotho expression than other compounds ( Figure 2).
  • KS101, KS102, KS103, and KS104 compounds were added to the retinal pigment epithelial cells (AEPE) in culture, and the cells were subcultured for 10 times. Then, BSO was added at a concentration of 1 ⁇ M and cultured for an additional 20 passages. Cell viability was measured. The proportion of living cells among cultured cells was measured using a LUNA-IITM automatic cell counter. LUNA-IITM automatic cell counter uses a reagent called tryphan blue. Living cells do not accept this reagent, but only dead cells are stained, indicating the ratio of live cells among all cells. As a result, it was confirmed that the survival rate of retinal pigment epithelial cells grown in medium containing KS compound was statistically higher ( p ⁇ 0.05) than that of cells grown in control conditions containing DMSO ( Fig. 3 ).
  • each of the KS compounds of Example 1 (KS101, KS102, KS103, and KS104 compounds) were added to the retinal pigment epithelial cells in culture, and the cells were subcultured for 10 passages.
  • BSO was added at a concentration of 1 ⁇ M, and BSO was added at a concentration of 1 ⁇ M for an additional 20 passages.
  • Activity in the cells Oxygen (Reactive Oxygen Species, ROS) concentration was measured.
  • ROS detection reagents from Molecular Probes were used as measurement reagents, and the concentration was measured by measuring the absorbance of the pigment produced by reacting with active oxygen using a spectrophotometer according to the method required by the manufacturer.
  • KS101, KS102, KS103, KS104 compounds 2.5 ⁇ M each of the KS compounds (KS101, KS102, KS103, KS104 compounds) of Example 1 were added to the retinal pigment epithelial cells in culture, and the concentration of reactive oxygen species (ROS) in the cells was tested after culturing for an additional 6 hours. Measurements were made in the same way as in 4. Through the experiment, it was confirmed that the amount of intracellular reactive oxygen species in retinal pigment epithelial cells grown in medium supplemented with KS compound for a short period of time was statistically lower ( p ⁇ 0.05) than in cells grown in control conditions to which DMSO was added. Through this, it was confirmed that KS compounds can reduce the amount of reactive oxygen species in cells even with a short treatment time of 6 hours ( Figure 4).
  • KS101, KS102, KS103, and KS104 compounds were compared using compound H, which was the basis for the development of these compounds. Cytotoxicity was measured using a reagent called EZ-CYTOX, a product of Dugen Biotechnology Research Institute. Using this reagent, the formation of an orange-colored water-soluble substance through dehydrogenase, which is active only in living cells, was quantified by measuring the absorbance at 405 nm. As a result of the experiment, it was confirmed that compound H showed clear cytotoxicity even at a concentration of about 5 ⁇ M, while KS compounds did not show significant toxicity even at a concentration of 10 ⁇ M. In particular, in the case of KS104, it was confirmed that cytotoxicity was not high even at a concentration of 40 ⁇ M ( Figure 6).
  • BSO a substance that causes cellular oxidative stress, inhibits the production of glutathione (GSH), an intracellular oxygen radical scavenging substance.
  • GSH glutathione
  • BSO a substance that causes cellular oxidative stress, inhibits the production of glutathione (GSH), an intracellular oxygen radical scavenging substance.
  • KS101, KS102, KS103, KS104 compounds 2.5 ⁇ M each of the KS compounds (KS101, KS102, KS103, KS104 compounds) of Example 1 were added to retinal pigment epithelial cells in culture, and cultured for 3 hours. . These cells were additionally treated with BSO (50mM) at a concentration that induces cytotoxicity, which was confirmed through preliminary experiments. After 6 hours of incubation, cytotoxicity was measured using EZ-CYTOX reagent. As a result, it was confirmed that the cytoprotective effect of all KS compounds was statistically higher ( p ⁇ 0.05) than the DMSO-treated control group
  • NaIO 3 a substance that causes intracellular oxidative stress, is known to be a substance that causes age-related macular degeneration (AMD) in retinal pigment epithelial cells.
  • AMD age-related macular degeneration
  • KS compounds of Example 1 2.5 ⁇ M each of the KS compounds of Example 1 (KS101, KS102, KS103, and KS104 compounds) were added to the retinal pigment epithelial cells and cultured for 3 hours.
  • NaIO 3 was added at a concentration of 5 mM to the cells and incubated for 24 hours. Additional culture was performed. Afterwards, the toxicity protection effect in cells was measured using EZ-CYTOX reagent. As a result, it was confirmed that the cytoprotective effect of all KS compounds was statistically higher ( p ⁇ 0.05) than the DMSO-treated control group (FIG. 8).

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Abstract

The present invention relates to a composition for preventing or treating macular degeneration, comprising a compound that induces the expression of anti-aging gene klotho. The compound according to the present invention has an excellent effect of enhancing the expression level of the age-associated gene Klotho, and also has a protective effect against oxidative stress in retinal pigment epithelium (RPE) cells, and thus can be advantageously used as a pharmaceutical composition or a health function food composition for preventing or treating macular degeneration.

Description

항노화 유전자 KLOTHO의 발현을 유도하는 화합물을 포함하는 황반변성의 예방 또는 치료용 조성물Composition for preventing or treating macular degeneration containing a compound that induces expression of the anti-aging gene KLOTHO

본 출원은 2022년 3월 22일 출원된 대한민국 특허출원 제10-2022-0035142호 및 2022년 12월 28일 출원된 대한민국 특허출원 제10-2022-0187495호를 우선권으로 주장하고, 상기 명세서 전체는 본 출원의 참고문헌이다. This application claims priority to Republic of Korea Patent Application No. 10-2022-0035142, filed on March 22, 2022, and Republic of Korea Patent Application No. 10-2022-0187495, filed on December 28, 2022, and the entire above specification is It is a reference to this application.

본 발명은 항노화 유전자 klotho의 발현을 유도하는 화합물을 포함하는 황반변성(Macular Degeneration)의 예방 또는 치료용 조성물에 관한 것이다.The present invention relates to a composition for preventing or treating macular degeneration containing a compound that induces the expression of the anti-aging gene klotho.

황반(macula)은 안구의 뒤쪽 망막의 중심부에 위치하여 물체의 상이 맺히는 곳으로, 시각세포가 집중적으로 모여있는 장소이다. 노인성황반변성은 이 황반 부위가 퇴화함으로써 발생한다. 퇴화하는 주원인으로 산소의 부족이 알려져 있으며, 이로 인해 혈관내피성장인자(Vascular Endothelial Growth Factor, VEGF)가 증가하여 혈관이 없던 망막 부위에 새로운 혈관이 형성됨으로 망막의 변성을 초래하게 된다. 망막의 시각세포가 모여 있는 바깥층에는 망막색소상피세포(Retinal Pigment Epithelium, RPE)이라는 극성을 나타내는 상피세포로 싸여 있다. 이 RPE세포의 극쪽(Apical)은 시각세포와 직접 접해 있으며 바닥쪽(Basal)은 맥락막(Choroid)층의 안쪽에 위치한 Burch's membrane과 접촉하고 있다. RPE세포의 기능은 시각 유지에 중요한 역할을 맡고 있다. 그 기능으로 시각세포 중 오래된 것을 제거하고 시각세포로부터 배출된 쓰레기 물질들을 맥락막으로 이동시켜 청소한다.The macula is located in the center of the retina at the back of the eye, where images of objects are formed, and is where visual cells are concentrated. Age-related macular degeneration occurs when the macular area degenerates. Lack of oxygen is known to be the main cause of degeneration, and as a result, Vascular Endothelial Growth Factor (VEGF) increases and new blood vessels are formed in areas of the retina where there were no blood vessels, resulting in retinal degeneration. The outer layer, where the retina's visual cells are gathered, is surrounded by polarized epithelial cells called retinal pigment epithelium (RPE). The apical side of these RPE cells is in direct contact with the visual cells, and the basal side is in contact with Burch's membrane located inside the choroid layer. The function of RPE cells plays an important role in maintaining vision. Its function is to remove old visual cells and move waste materials discharged from visual cells to the choroid for cleaning.

노인성황반변성(Age-Related Macular Degeneration, AMD)은 노화가 발생함으로 생기는 질환으로, 70세 이상 성인의 3명 중 1명이 증상을 겪고 있을 정도로 노령자에게서 발생률 높은 질환이다. 노인성황반변성의 발생 빈도는 다른 안구질환인 녹내장(Glaucoma)과 백내장(Cataract)의 발생 빈도를 합한 것보다 많을 정도로 높을 발생 빈도를 나타내는 대표적 안과 질환으로 계속 병이 진행할 경우 시력을 잃게 된다. Age-Related Macular Degeneration (AMD) is a disease that occurs due to aging. It is a disease with a high incidence in the elderly, with one in three adults over the age of 70 suffering from symptoms. Age-related macular degeneration is a representative eye disease with a higher occurrence frequency than the combined incidence of other eye diseases, glaucoma and cataract. If the disease continues to progress, vision is lost.

노인성황반변성의 경우 황반에서 이들 RPE세포와 시각세포의 손실이 발생함으로써 일어난다. 노인성황반변성 질환의 조직학적 특징은 RPE세포층과 Burch's membrane 사이에 드루센(Drusen)이라는 결정이 축적되는 것인데, 이 결정의 성분은 대부분은 산화스트레스와 염증을 통해 만들어지는 것들이다. 그러므로 산화스트레스나 염증이 노인성황반변성 발생에 원인이 된다고 볼 수 있다. 드루센의 축적과 이 부위에 새로운 악성 혈관들이 발생함으로써 시각 능력이 점차 소실되게 되는데, 물체가 휘어 보이는 변형시각이나 시야의 중심부위에 암점이 발생하여 볼 수 없게 되는 등의 현상이 나타나기 시작한다. 치료하지 않고 그냥 두면 시력이 점차 소실되어 결국 실명하게 된다. 현재 이를 위한 치료법으로는 주로 레이저나 약물 투여를 통한 악성 혈관의 제거를 이용하고 있다. 하지만 이는 근원적인 치료가 되지 않고 증상이 악화하는 것을 완화시키는 방법밖에는 되지 않는다. Age-related macular degeneration occurs due to loss of RPE cells and visual cells in the macula. The histological characteristic of age-related macular degeneration disease is the accumulation of crystals called drusen between the RPE cell layer and Burch's membrane. Most of the components of these crystals are formed through oxidative stress and inflammation. Therefore, oxidative stress or inflammation can be considered to be the cause of age-related macular degeneration. Due to the accumulation of drusen and the development of new malignant blood vessels in this area, visual ability is gradually lost. Phenomenons such as distorted vision where objects appear curved or scotomas in the center of the field of vision, making it impossible to see, begin to appear. If left untreated, vision will gradually decrease and eventually lead to blindness. Currently, the treatment for this mainly uses laser or drug administration to remove malignant blood vessels. However, this does not provide a fundamental treatment and is only a way to alleviate the worsening symptoms.

한편, 동물의 노화를 조절할 수 있는 유전자가 존재할 수 있다는 사실은 1981년에 보고된 SAM(senescence-accelerated mice)에서 알려지게 되었다. AKR/J 계열의 쥐를 교배하는 도중 우연히 만들어진 이 쥐는 같은 계열의 쥐보다 훨씬 노화가 빨랐는데, 이 쥐는 여러 유전자에 변이를 가지고 있는 것이 확인되었다. 이후 단일 그룹의 노화 관련 유전자 발견은 1990년대에 보고되었다. 보고된 유전자는 RecQ family로 DNA helicase라는 효소를 발현시키는 유전자들이었다. 이 유전자들에 변이가 발생하면 조기 노화가 발생하거나 암이 생성되는 결과가 보고되었는데, 이는 DNA의 수리에 영향을 미쳐서 발생하는 것으로 알려졌다. 단일 유전자로 노화와 관련된 것은 1997년에 보고가 된 klotho 유전자이다. Klotho 유전자는 고혈압 쥐의 형질변경 동물모델을 만들던 중 우연히 발견되었는데, 이 유전자를 발현하지 못하게 된 쥐에서는 조기 노화 현상이 발생하고 생명이 짧아졌다. 더 흥미로운 사실은 나중에 이 유전자의 발현을 증가시킨 쥐는 수컷의 경우 수명이 20.0~30.8%가 증가했고 암컷에서는 18.8 ~ 19.0% 증가하였다. 이는 단일 유전자의 발현에 따라 쥐의 수명이 늘어날 수도, 줄어들 수도 있다는 사실을 처음으로 세상에 알리게 된 계기가 되었다. 그리고 Klotho 유전자의 염기서열은 동물 간에 매우 유사하여 쥐와 사람의 경우는 98% 정도나 일치하는 것으로 보고되었다. 이는 사람에서도 klotho 유전자의 발현에 따라 수명이 조절될 수 있음을 나타낸다. Meanwhile, the fact that genes that can control aging in animals may exist became known from SAM (senescence-accelerated mice) reported in 1981. This mouse, which was accidentally created while crossing AKR/J mice, aged much faster than mice of the same line, and it was confirmed that this mouse had mutations in several genes. Later, the discovery of a single group of aging-related genes was reported in the 1990s. The genes reported were RecQ family genes that express an enzyme called DNA helicase. Mutations in these genes have been reported to result in premature aging or cancer, which is known to occur by affecting DNA repair. The single gene associated with aging is the klotho gene, which was reported in 1997. The Klotho gene was accidentally discovered while creating a genetically modified animal model for hypertensive rats. In rats that were unable to express this gene, premature aging occurred and their lives were shortened. What's more interesting is that mice that later increased the expression of this gene had an increase in lifespan of 20.0 to 30.8% in males and 18.8 to 19.0% in females. This served as an opportunity to inform the world for the first time that the lifespan of mice can be increased or decreased depending on the expression of a single gene. And the base sequence of the Klotho gene is very similar between animals, and it has been reported that there is about 98% identity between mice and humans. This indicates that even in humans, lifespan can be regulated by the expression of the klotho gene.

사람에서 klotho 유전자는 13번째 염색체에 위치하며 β-glucosidase와 염기서열이 유사한 막 단백질을 생산한다. Klotho 단백질은 신장의 세뇨관 상피세포와 뇌의 맥락총(choroid plexus)에서 주로 발현되며 일부 부갑상샘에서 발현되는 것으로 보고되었다. Klotho 유전자는 다양한 노화의 표현형에 관련된 유전자로, klotho 유전자가 결핍된 쥐에서는 수명감소, 활동성저하, 성장지연, 죽상경화증, 동맥의 석회화, 골다공증, 생식기미숙, 불임, 피부위축, 폐기종 등의 노화과정과 유사한 증후군이 발생한다. Klotho 변이 쥐에서는 인간에서 노화에 의하여 발생하는 중막 석회증(Monckeberg type)의 동맥경화증과 유사한 양상의 동맥경화증이 대동맥부터 소동맥까지 모든 동맥에서 관찰되고, 혈관형성(angiogenesis)과 맥관형성(vasculogenesis)에 장애가 발생한다. In humans, the klotho gene is located on chromosome 13 and produces a membrane protein with a base sequence similar to that of β-glucosidase. Klotho protein is mainly expressed in renal tubular epithelial cells and brain choroid plexus, and has been reported to be expressed in some parathyroid glands. The Klotho gene is a gene involved in various aging phenotypes. In mice deficient in the klotho gene, the aging process includes reduced lifespan, reduced activity, growth retardation, atherosclerosis, arterial calcification, osteoporosis, reproductive immaturity, infertility, skin atrophy, and emphysema. A similar syndrome occurs. In Klotho mutant mice, arteriosclerosis similar to the arteriosclerosis of the media (Monckeberg type) that occurs due to aging in humans is observed in all arteries from the aorta to the small artery, and is affected by angiogenesis and vasculogenesis. Disruption occurs.

Klotho mRNA는 신장 조직에서 타조직보다 발현이 현저히 높으나, 고혈압, 제2형 당뇨병, 당뇨병성 신증, 만성신부전의 질환 모델 쥐의 신장에서는 klotho mRNA의 발현이 감소된다. Klotho 발현 저하 쥐에서는 혈관내피유래 이완인자인 NO(Nitric Oxide) 생산이 감소하고, 많은 심혈관계질환의 위험인자를 동시에 가진 OLETF(Otsuka Long-Evans Tokushima fatty rat) 쥐에 klotho 유전자를 바이러스 유전자 전달체를 이용하여 주입하면 혈관내막 기능이상이 호전되고 NO 생산을 증가시키며 혈관 비후와 섬유화를 억제하여 혈압을 강하시킨다. 또한 klotho 유전자는 쥐에서 당과 인슐린대사에도 영향을 미치며, 대표적인 고콜레스테롤혈증 치료제인 스타틴은 신장 근위세뇨관 세포에서 klotho mRNA의 발현을 증가시킨다. Klotho 발현 저하 쥐에서는 조골세포와 파골세포 모두의 분화장애로 인한 낮은 골교체 상태의 골감소증이 발생하고 이는 인간에서 연령증가에 따른 골 소실 및 노인성 골다공증의 특징과 유사하다. 또한 Klotho 변이 쥐에서는 골단 부위의 소주골의 비정상적인 신장과 미세 전산화단층촬영에서 비정상적인 소주골 조직 소견이 관찰되며 이는 골 흡수 과정의 장애에 기인한다. 사람에서 관찰되는 Klotho 유전자 돌연변이에 의한 임상적 표현형의 변화는 다양하다. Klotho 유전자 exon2의 세 부위의 돌연변이를 가진 KL-VS(functional variant of klotho) 변형은 지질대사, 혈압, 수명, 인지기능, 관상동맥질환 및 뇌혈관 질환과 관련되며, Klotho 유전자의 미세부수체(microsatellite) 다형성과 단일염기 유전자 다형성이 골밀도와 연관되었고, 건강한 성인 여성에서 Klotho 유전자의 단일염기 유전자 다형성이 심혈관계질환의 위험인자 및 골밀도와 관련됨도 발표된 바 있다. 최근에는 Klotho 유전자와 알츠하이머와의 연관성도 여러 논문을 통해 보고되었다. 알츠하이머 치매 생쥐모델에서 klotho를 과발현시킬 경우 쥐의 수명이 30% 증가하고 인지기능 저하가 억제되는 것이 보고되었다. 아울러 klotho 발현에 의해 뇌 내에서 아밀로이드 베타 단백질의 생성이 50% 감소하는 것이 관찰되었다. 사람에서도 klotho의 발현량은 알츠하이머 질환의 진행 정도와 반비례하고, klotho 단백질이 알츠하이머 질환자의 혈액에서 염증성 사이토킨의 양을 감소시킨다는 보고가 제출되었다. Klotho는 망막색소상피(RPE)의 기능을 증진시키고 산화스트레스로부터 RPE세포를 보호하는 효과를 가지고 있다는 여러 연구 결과도 보고되었다. Klotho 유전자를 없앤(KL-/-) 쥐의 망막에서는 광수용체세포(photoreceptor)들의 퇴행과 RPE세포에서의 색소 생성이 감소된 것이 나타나는 것이 알려졌다. 배양된 RPE세포에서도 klotho가 발현된다는 것이 확인되었고, 발현된 klotho 단백질은 L-3,4-dihydorxyphenylalanine(L-DOPA)의 합성을 증가하고, 기저막으로부터 혈관내피성장인자(vascular endothelial growth factor, VEGF)의 생성을 억제시키는 것이 실험을 통해 확인되었다. 또한 RPE세포에서 klotho는 활성산소(reactive oxygen species, ROS)의 생성을 감소시켜 RPE세포를 산화스트레스로부터 보호하는 기능이 있음이 연구결과로 확인되었다. 노인성황반변성(AMD)을 가진 환자들을 대상으로 실험한 결과 안구 방수액(aqueous humor)에서 klotho 발현이 감소되어 있었는데, 이 감소 효과는 산화스트레스 및 염증과 연관되어 있음이 확인되었다.Klotho mRNA expression is significantly higher in kidney tissue than in other tissues, but expression of klotho mRNA is reduced in the kidneys of mouse disease models for hypertension, type 2 diabetes, diabetic nephropathy, and chronic renal failure. In mice with low Klotho expression, the production of NO (Nitric Oxide), a vascular endothelium-derived relaxation factor, is reduced, and the klotho gene was transferred to OLETF (Otsuka Long-Evans Tokushima fatty rats) mice, which have many risk factors for cardiovascular disease, using a viral gene delivery system. When injected, it improves vascular intimal dysfunction, increases NO production, and suppresses vascular thickening and fibrosis, thereby lowering blood pressure. Additionally, the klotho gene also affects glucose and insulin metabolism in mice, and statins, a representative hypercholesterolemia treatment, increase the expression of klotho mRNA in kidney proximal tubule cells. In mice with low Klotho expression, osteopenia with low bone turnover occurs due to impaired differentiation of both osteoblasts and osteoclasts, which is similar to the characteristics of age-related bone loss and senile osteoporosis in humans. Additionally, in Klotho mutant mice, abnormal elongation of trabecular bone in the epiphyseal region and abnormal trabecular bone tissue findings are observed in microcomputed tomography, which is due to a disruption in the bone resorption process. The clinical phenotypic changes caused by Klotho gene mutations observed in humans are diverse. KL-VS (functional variant of klotho), which has mutations in three regions of the Klotho gene exon2, is associated with lipid metabolism, blood pressure, lifespan, cognitive function, coronary artery disease, and cerebrovascular disease, and is associated with microsatellites of the Klotho gene. ) polymorphisms and single nucleotide polymorphisms have been associated with bone mineral density, and it has also been reported that single nucleotide polymorphisms in the Klotho gene are associated with risk factors for cardiovascular disease and bone mineral density in healthy adult women. Recently, the association between the Klotho gene and Alzheimer's has been reported in several papers. It has been reported that when klotho is overexpressed in an Alzheimer's dementia mouse model, the lifespan of the mouse increases by 30% and cognitive decline is suppressed. In addition, it was observed that the production of amyloid beta protein in the brain was reduced by 50% due to klotho expression. In humans, it has been reported that the expression level of klotho is inversely proportional to the progression of Alzheimer's disease, and that klotho protein reduces the amount of inflammatory cytokines in the blood of Alzheimer's disease patients. Several studies have reported that Klotho improves the function of retinal pigment epithelium (RPE) and has the effect of protecting RPE cells from oxidative stress. It has been known that the retina of mice with the Klotho gene deleted (KL -/- ) shows degeneration of photoreceptors and decreased pigment production in RPE cells. It was confirmed that klotho is expressed in cultured RPE cells, and the expressed klotho protein increases the synthesis of L-3,4-dihydorxyphenylalanine (L-DOPA) and vascular endothelial growth factor (VEGF) from the basement membrane. It was confirmed through experiments that the production of was suppressed. Additionally, research results have confirmed that klotho has the function of protecting RPE cells from oxidative stress by reducing the production of reactive oxygen species (ROS) in RPE cells. As a result of experiments on patients with age-related macular degeneration (AMD), klotho expression was decreased in the aqueous humor of the eye, and this decrease was confirmed to be associated with oxidative stress and inflammation.

본 발명자는 klotho 발현을 유도하는 화합물에 대한 연구를 지속적으로 수행한 결과, 본 발명자가 개발한 신규 화합물이 안정성이 우수하며 망막색소상피세포(RPE)에서 산화스트레스(산화독성)에 대한 보호 효과가 우수하다는 것을 실험적으로 확인하고, 본 발명을 완성하였다.The present inventor has continuously conducted research on compounds that induce klotho expression, and as a result, the new compound developed by the present inventor has excellent stability and has a protective effect against oxidative stress (oxidative toxicity) in retinal pigment epithelial cells (RPE). It was experimentally confirmed that it was excellent, and the present invention was completed.

본 발명의 목적은 화학식 1로 표시되는 화합물 또는 이의 약학적으로 허용가능한 염을 유효성분으로 포함하는 황반변성 예방 또는 치료용 약학적 조성물을 제공하는 데 있다. The purpose of the present invention is to provide a pharmaceutical composition for preventing or treating macular degeneration containing the compound represented by Formula 1 or a pharmaceutically acceptable salt thereof as an active ingredient.

본 발명의 다른 목적은 화학식 1로 표시되는 화합물 또는 이의 약학적으로 허용가능한 염을 유효성분으로 포함하는 황반변성 예방 또는 개선용 식품 조성물을 제공하는 데 있다. Another object of the present invention is to provide a food composition for preventing or improving macular degeneration containing the compound represented by Formula 1 or a pharmaceutically acceptable salt thereof as an active ingredient.

그러나, 본 발명이 이루고자 하는 기술적 과제는 이상에서 언급한 과제에 제한되지 않으며, 언급되지 않은 또 다른 과제들은 아래의 기재로부터 당업자에게 명확하게 이해될 수 있을 것이다.However, the technical problem to be achieved by the present invention is not limited to the problems mentioned above, and other problems not mentioned will be clearly understood by those skilled in the art from the description below.

상기 목적을 달성하기 위해, To achieve the above purpose,

본 발명은 하기 화학식 1로 표시되는 화합물 또는 이의 약학적으로 허용가능한 염을 유효성분으로 포함하는, 황반변성 예방 또는 치료용 약학적 조성물을 제공한다.The present invention provides a pharmaceutical composition for preventing or treating macular degeneration, comprising a compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient.

또한, 본 발명은 하기 화학식 1로 표시되는 화합물 또는 이의 약학적으로 허용가능한 염을 유효성분으로 포함하는, 황반변성 예방 또는 개선용 식품 조성물 또는 건강기능식품 조성물을 제공한다.In addition, the present invention provides a food composition or health functional food composition for preventing or improving macular degeneration, comprising a compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient.

[화학식 1][Formula 1]

Figure PCTKR2023003592-appb-img-000001
Figure PCTKR2023003592-appb-img-000001

상기 화학식 1에서, In Formula 1,

R1 및 R2는 각각 -H, 할로겐 또는 C1-6의 직쇄 또는 측쇄 알킬이고,R 1 and R 2 are each -H, halogen or C 1-6 straight or branched alkyl,

R3 및 R4는 각각 -H 또는 할로겐일 수 있다.R 3 and R 4 may each be -H or halogen.

본 발명의 바람직한 일실시예에 있어서, 상기 화학식 1로 표시되는 화합물은 하기 화학식 1-1로 표시되는 화합물일 수 있다.In a preferred embodiment of the present invention, the compound represented by Formula 1 may be a compound represented by Formula 1-1 below.

[화학식 1-1][Formula 1-1]

Figure PCTKR2023003592-appb-img-000002
Figure PCTKR2023003592-appb-img-000002

상기 화학식 1-1에서 R1 및 R2는 각각 -H, 할로겐 또는 C1-6의 직쇄 또는 측쇄 알킬일 수 있다.In Formula 1-1, R 1 and R 2 may each be -H, halogen, or C 1-6 straight or branched alkyl.

본 발명의 바람직한 다른 일실시예에 있어서, 상기 화학식 1-1로 표시되는 화합물은 In another preferred embodiment of the present invention, the compound represented by Formula 1-1 is

하기 화학식 1-1A로 표시되는 화합물;A compound represented by the following formula 1-1A;

[화학식 1-1A][Formula 1-1A]

Figure PCTKR2023003592-appb-img-000003
Figure PCTKR2023003592-appb-img-000003

하기 화학식 1-1B로 표시되는 화합물;A compound represented by the following formula 1-1B;

[화학식 1-1B][Formula 1-1B]

Figure PCTKR2023003592-appb-img-000004
Figure PCTKR2023003592-appb-img-000004

하기 화학식 1-1C로 표시되는 화합물; 및A compound represented by the following formula 1-1C; and

[화학식 1-1C][Formula 1-1C]

Figure PCTKR2023003592-appb-img-000005
Figure PCTKR2023003592-appb-img-000005

하기 화학식 1-1D로 표시되는 화합물;Compounds represented by the following formula 1-1D;

[화학식 1-1D][Formula 1-1D]

Figure PCTKR2023003592-appb-img-000006
Figure PCTKR2023003592-appb-img-000006

로 구성된 군에서 선택된 어느 하나 이상일 수 있다.It may be any one or more selected from the group consisting of.

본 발명의 바람직한 또 다른 일실시예에 있어서, 상기 조성물은 클로소(Klotho) 유전자의 발현을 증가시킬 수 있다. In another preferred embodiment of the present invention, the composition can increase the expression of the Klotho gene.

본 발명이 바람직한 또 다른 일실시예에 있어서, 상기 조성물은 산화스트레스로부터 망막색소상피세포를 보호할 수 있다.In another preferred embodiment of the present invention, the composition can protect retinal pigment epithelial cells from oxidative stress.

또한, 본 발명은 상기 화학식 1로 표시되는 화합물 또는 이의 약학적으로 허용가능한 염을 유효성분으로 포함하는 조성물을 개체에 투여하는 단계를 포함하는 황반변성 예방 또는 치료 방법을 제공한다.Additionally, the present invention provides a method for preventing or treating macular degeneration, comprising administering to a subject a composition containing the compound represented by Formula 1 or a pharmaceutically acceptable salt thereof as an active ingredient.

또한, 본 발명은 상기 화학식 1로 표시되는 화합물 또는 이의 약학적으로 허용가능한 염을 유효성분으로 포함하는 조성물의 황반변성 예방 또는 치료 용도를 제공한다.In addition, the present invention provides a use for preventing or treating macular degeneration of a composition containing the compound represented by Formula 1 or a pharmaceutically acceptable salt thereof as an active ingredient.

본 발명에 따른 화합물은 노화와 관련된 유전자인 Klotho 유전자의 발현량을 향상시키는 효과가 있으며, 망막색소상피세포(RPE)에서 산화스트레스에 대한 보호 효과를 가지고 있어, 황반변성 예방 또는 치료용 약학적 조성물, 또는 건강기능식품 조성물로 유용하게 사용할 수 있다.The compound according to the present invention has the effect of improving the expression level of the Klotho gene, a gene related to aging, and has a protective effect against oxidative stress in retinal pigment epithelial cells (RPE), and is a pharmaceutical composition for preventing or treating macular degeneration. , or can be usefully used as a health functional food composition.

도 1은 본 발명의 실시예 1에 따른 KS화합물의 안정성을 MS/MS분석으로 확인한 결과이다: KS101(화학식1-1A 화합물), KS102(화학식1-1B 화합물), KS103(화학식1-1C 화합물), KS104(화학식1-1D 화합물).Figure 1 shows the results of confirming the stability of KS compounds according to Example 1 of the present invention by MS/MS analysis: KS101 (compound of Formula 1-1A), KS102 (compound of Formula 1-1B), KS103 (compound of Formula 1-1C) ), KS104 (Formula 1-1D compound).

도 2는 본 발명의 실시예 1에 따른 KS화합물에 의한 secreted(분비) klotho 및 membrane(막) klotho 유전자의 발현 정도를 PCR로 확인한 결과이다. Figure 2 shows the results of confirming the expression level of secreted klotho and membrane klotho genes by the KS compound according to Example 1 of the present invention by PCR.

도 3은 본 발명의 실시예 1에 따른 KS화합물을 장시간 처리한 세포(ARPE: 망막색소상피세포)에서의 생존율 변화를 확인한 결과이다.Figure 3 shows the results of confirming changes in survival rate in cells (ARPE: retinal pigment epithelial cells) treated with the KS compound for a long time according to Example 1 of the present invention.

도 4는 본 발명의 실시예 1에 따른 KS화합물을 장시간 처리한 세포(ARPE: 망막색소상피세포)에서의 활성산소(Reactive Oxygen Species, ROS) 변화를 확인한 결과이다.Figure 4 shows the results of confirming changes in reactive oxygen species (ROS) in cells (ARPE: retinal pigment epithelial cells) treated with the KS compound for a long time according to Example 1 of the present invention.

도 5는 본 발명의 실시예 1에 따른 KS화합물을 단시간(6시간) 처리한 세포(ARPE: 망막색소상피세포)에서의 활성산소(Reactive Oxygen Species, ROS) 변화를 확인한 결과이다.Figure 5 shows the results of confirming changes in reactive oxygen species (ROS) in cells (ARPE: retinal pigment epithelial cells) treated with the KS compound for a short period of time (6 hours) according to Example 1 of the present invention.

도 6은 본 발명의 실시예 1에 따른 KS화합물의 세포독성을 확인한 결과이다. Figure 6 shows the results of confirming the cytotoxicity of the KS compound according to Example 1 of the present invention.

도 7은 BSO로 유발된 산화스트레스로부터 KS화합물들의 세포 보호 효과를 확인한 결과이다.Figure 7 shows the results confirming the cell protection effect of KS compounds from oxidative stress induced by BSO.

도 8은 NaIO3로 유발된 산화스트레스로부터 KS화합물들의 세포 보호 효과를 확인한 결과이다.Figure 8 shows the results confirming the cell protection effect of KS compounds from oxidative stress induced by NaIO 3 .

이하, 본 발명을 상세하게 설명한다. Hereinafter, the present invention will be described in detail.

본 발명은 하기 화학식 1로 표시되는 화합물 또는 이의 약학적으로 허용가능한 염을 유효성분으로 포함하는 황반변성 예방, 개선 또는 치료용 조성물을 제공한다. The present invention provides a composition for preventing, improving or treating macular degeneration comprising a compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient.

[화학식 1][Formula 1]

Figure PCTKR2023003592-appb-img-000007
Figure PCTKR2023003592-appb-img-000007

(상기 화학식 1에서, R1 및 R2는 각각 -H, 할로겐 또는 C1-6의 직쇄 또는 측쇄 알킬이고, R3 및 R4는 각각 -H 또는 할로겐일 수 있다.)(In Formula 1 above, R 1 and R 2 may each be -H, halogen, or C 1-6 straight or branched alkyl, and R 3 and R 4 may each be -H or halogen.)

바람직하게, 상기 화학식 1로 표시되는 화합물은 하기 화학식 1-1로 표시되는 화합물일 수 있다.Preferably, the compound represented by Formula 1 may be a compound represented by Formula 1-1 below.

[화학식 1-1][Formula 1-1]

Figure PCTKR2023003592-appb-img-000008
Figure PCTKR2023003592-appb-img-000008

(상기 화학식 1-1에서 R1 및 R2는 각각 -H, 할로겐 또는 C1-6의 직쇄 또는 측쇄 알킬일 수 있다.)(In Formula 1-1, R 1 and R 2 may each be -H, halogen, or C 1-6 straight or branched alkyl.)

더 바람직하게, 상기 화학식 1-1로 표시되는 화합물은, More preferably, the compound represented by Formula 1-1 is:

하기 화학식 1-1A로 표시되는 화합물;A compound represented by the following formula 1-1A;

[화학식 1-1A][Formula 1-1A]

Figure PCTKR2023003592-appb-img-000009
Figure PCTKR2023003592-appb-img-000009

하기 화학식 1-1B로 표시되는 화합물;A compound represented by the following formula 1-1B;

[화학식 1-1B][Formula 1-1B]

Figure PCTKR2023003592-appb-img-000010
Figure PCTKR2023003592-appb-img-000010

하기 화학식 1-1C로 표시되는 화합물; 및A compound represented by the following formula 1-1C; and

[화학식 1-1C][Formula 1-1C]

Figure PCTKR2023003592-appb-img-000011
Figure PCTKR2023003592-appb-img-000011

하기 화학식 1-1D로 표시되는 화합물;Compounds represented by the following formula 1-1D;

[화학식 1-1D][Formula 1-1D]

Figure PCTKR2023003592-appb-img-000012
Figure PCTKR2023003592-appb-img-000012

로 구성된 군에서 선택된 어느 하나 이상일 수 있다.It may be any one or more selected from the group consisting of.

상기 황반변성은 노인성 황반변성일 수 있다.The macular degeneration may be age-related macular degeneration.

본 발명의 조성물은 클로소(Klotho) 유전자의 발현정도를 증가시킬 수 있다. The composition of the present invention can increase the expression level of the Klotho gene.

또한, 본 발명의 조성물은 산화스트레스로부터 망막색소상피 세포를 보호할 수 있다.Additionally, the composition of the present invention can protect retinal pigment epithelial cells from oxidative stress.

상기 조성물은 약학적 조성물, 식품 조성물 또는 건강기능식품 조성물일 수 있다.The composition may be a pharmaceutical composition, food composition, or health functional food composition.

상기 조성물은, 조성물 전체 중량에 대하여 상기 유효성분을 1 내지 99 중량 %, 5 내지 95 중량%, 또는 10 내지 90 중량%로 포함할 수 있으나, 이에 제한되는 것은 아니다. The composition may contain 1 to 99% by weight, 5 to 95% by weight, or 10 to 90% by weight of the active ingredient based on the total weight of the composition, but is not limited thereto.

본 명세서 내 "예방"은 본 발명의 조성물의 투여로 황반변성의 발병을 지연시키는 모든 행위를 의미하고, "치료" 및 "개선"은 본 발명의 조성물의 투여로 황반변성의 증세가 호전 또는 이롭게 변경되는 모든 행위를 의미한다.As used herein, “prevention” refers to any action that delays the onset of macular degeneration by administration of the composition of the present invention, and “treatment” and “improvement” refers to the improvement or beneficial change in symptoms of macular degeneration by administration of the composition of the present invention. It means all actions.

본 발명의 상기 화합물은 약학적으로 허용가능한 염의 형태로 사용할 수 있으며, 염으로는 약학적으로 허용가능한 유리산(free acid)에 의해 형성된 산부가염이 유용하다. 약학적으로 허용가능한 염이란 표현은 환자에게 비교적 비독성이고 무해한 유효작용을 갖는 농도로서 이 염에 기인한 부작용이 화학식 1의 염기 화합물의 이로운 효능을 떨어뜨리지 않는 화학식 1의 염기 화합물의 어떠한 유기 또는 무기 부가염을 의미한다. 이들 염은 유리산으로는 무기산과 유기산을 사용할 수 있으며, 무기산으로는 염산, 브롬산, 질산, 황산, 과염소산, 인산 등을 사용할 수 있고, 유기산으로는 구연산, 초산, 젖산, 말레산, 푸마린산, 글루콘산, 메탄설폰산, 글리콘산, 숙신산, 타타르산, 갈룩투론산, 엠본산, 글루탐산, 아스파르트산, 옥살산, (D) 또는 (L) 말산, 말레산, 메테인설폰산, 에테인설폰산, 4-톨루엔술폰산, 살리실산, 시트르산, 벤조산 또는 말론산 등을 사용할 수 있다. 또한, 이들 염은 알칼리 금속염(나트륨염, 칼륨염 등) 및 알칼리 토금속염(칼슘염, 마그네슘염 등) 등을 포함한다. 예를 들면, 산부가염으로는 아세테이트, 아스파테이트, 벤즈에이트, 베실레이트, 바이카보네이트/카보네이트, 바이설페이트/설페이트, 보레이트, 캄실레이트, 시트레이트, 에디실레이트, 에실레이트, 포메이트, 퓨마레이트, 글루셉테이트, 글루코네이트, 글루큐로네이트, 헥사플루오르포스페이트, 하이벤제이트, 하이드로클로라이드/클로라이드, 하이드로브로마이드/브로마이드, 하이드로요오디드/요오디드, 이세티오네이트, 락테이트, 말레이트, 말리에이트, 말로네이트, 메실레이트, 메틸설페이트, 나프틸레이트, 2-나프실레이트, 니코티네이트, 나이트레이트, 오로테이트, 옥살레이트, 팔미테이트, 파모에이트, 포스페이트/수소 포스페이트/이수소 포스페이트, 사카레이트, 스테아레이트, 석시네이트, 타르트레이트, 토실레이트, 트리플루오르아세테이트, 알루미늄, 알기닌, 벤자틴, 칼슘, 콜린, 디에틸아민, 디올아민, 글라이신, 라이신, 마그네슘, 메글루민, 올아민, 칼륨, 나트륨, 트로메타민, 아연염 등이 포함될 수 있으며, 이들 중 하이드로클로라이드 또는 트리플루오르아세테이트가 바람직하다.The compound of the present invention can be used in the form of a pharmaceutically acceptable salt, and an acid addition salt formed by a pharmaceutically acceptable free acid is useful as the salt. The expression “pharmaceutically acceptable salt” refers to any organic or organic salt of the base compound of Formula 1 where side effects due to the salt do not reduce the beneficial effects of the base compound of Formula 1, at a concentration that is relatively non-toxic and harmless to the patient and has an effective effect. It means inorganic addition salt. For these salts, inorganic acids and organic acids can be used as free acids. Hydrochloric acid, hydrobromic acid, nitric acid, sulfuric acid, perchloric acid, and phosphoric acid can be used as inorganic acids, and citric acid, acetic acid, lactic acid, maleic acid, and fumarine can be used as organic acids. Acids, gluconic acid, methanesulfonic acid, glyconic acid, succinic acid, tartaric acid, galacturonic acid, embonic acid, glutamic acid, aspartic acid, oxalic acid, (D) or (L) malic acid, maleic acid, methanesulfonic acid, ethane sulfuric acid Fonic acid, 4-toluenesulfonic acid, salicylic acid, citric acid, benzoic acid, or malonic acid can be used. Additionally, these salts include alkali metal salts (sodium salts, potassium salts, etc.) and alkaline earth metal salts (calcium salts, magnesium salts, etc.). For example, acid addition salts include acetate, aspartate, benzate, besylate, bicarbonate/carbonate, bisulfate/sulfate, borate, camsylate, citrate, edisylate, esylate, formate, fumarate, Gluceptate, gluconate, glucuronate, hexafluorophosphate, hybenzate, hydrochloride/chloride, hydrobromide/bromide, hydroiodide/iodide, isethionate, lactate, maleate, maleate , malonate, mesylate, methyl sulfate, naphthylate, 2-naphsylate, nicotinate, nitrate, orotate, oxalate, palmitate, pamoate, phosphate/hydrogen phosphate/dihydrogen phosphate, saccharate. , stearate, succinate, tartrate, tosylate, trifluoroacetate, aluminum, arginine, benzathine, calcium, choline, diethylamine, diolamine, glycine, lysine, magnesium, meglumine, olamine, potassium, Sodium, tromethamine, zinc salt, etc. may be included, and among these, hydrochloride or trifluoroacetate is preferable.

또한, 본 발명의 상기 화합물은 약학적으로 허용되는 염뿐만 아니라, 통상의 방법에 의해 제조될 수 있는 모든 염, 이성질체, 수화물 및 용매화물을 모두 포함한다.In addition, the compound of the present invention includes not only pharmaceutically acceptable salts, but also all salts, isomers, hydrates and solvates that can be prepared by conventional methods.

본 발명에 따른 부가염은 통상의 방법으로 제조할 수 있으며, 예를 들면 화합물을 수혼화성 유기용매, 예를 들면 아세톤, 메탄올, 에탄올, 또는 아세토니트릴 등에 녹이고 과량의 유기산을 가하거나 무기산의 산 수용액의 가한 후 침전시키거나 결정화시켜서 제조할 수 있다. 이어서 이 혼합물에서 용매나 과량의 산을 증발시킨 후 건조시켜서 부가염을 얻거나 또는 석출된 염을 흡인 여과시켜 제조할 수 있다.The addition salt according to the present invention can be prepared by conventional methods. For example, the compound is dissolved in a water-miscible organic solvent, such as acetone, methanol, ethanol, or acetonitrile, and an excess amount of organic acid is added or an aqueous acid solution of an inorganic acid is added. It can be manufactured by adding and then precipitating or crystallizing. Then, the solvent or excess acid in this mixture is evaporated and dried to obtain an addition salt, or the precipitated salt can be prepared by suction filtration.

본 발명에서는, 실험을 통해, 상기 화합물 클로소(Klotho) 유전자의 발현을 증가시킬 수 있음을 확인하였다. In the present invention, it was confirmed through experiments that the compound can increase the expression of the Klotho gene.

또한, 망막색소상피를 이용한 산화스트레스 실험을 통해 상기 화합물들이 산화스트레스로부터 세포를 보호하는 효과를 나타내는 것을 확인하였는 바, 본 발명의 화합물들은 황반변성 질환의 예방, 개선 또는 치료제로 유용하게 이용할 수 있다.In addition, through an oxidative stress experiment using retinal pigment epithelium, it was confirmed that the compounds exhibit an effect of protecting cells from oxidative stress. Therefore, the compounds of the present invention can be usefully used as a preventive, ameliorating, or therapeutic agent for macular degeneration disease. .

황반변성 예방 또는 치료용 약학적 조성물Pharmaceutical composition for preventing or treating macular degeneration

본 발명의 조성물은 약학적 조성물로 제조될 수 있다.The compositions of the present invention can be prepared as pharmaceutical compositions.

본 발명의 조성물이 약학적 조성물로 제조되는 경우, 본 발명의 약학적 조성물은 약학적으로 허용되는 담체를 포함할 수 있다.When the composition of the present invention is prepared as a pharmaceutical composition, the pharmaceutical composition of the present invention may include a pharmaceutically acceptable carrier.

본 발명의 바람직한 구현예에 따르면, 본 발명의 조성물은 (a) 상술한 본 발명의 화합물 또는 이의 약학적으로 허용가능한 염의 약학적 유효량; 및 (b) 약학적으로 허용되는 담체를 포함하는 약학적 조성물일 수 있다. 본 명세서에서 용어 "약학적 유효량"은 상술한 본 발명의 화합물 또는 이의 약학적으로 허용가능한 염의 효능 또는 활성을 달성하는 데 충분한 양을 의미한다.According to a preferred embodiment of the present invention, the composition of the present invention comprises (a) a pharmaceutically effective amount of the above-described compound of the present invention or a pharmaceutically acceptable salt thereof; and (b) a pharmaceutically acceptable carrier. As used herein, the term “pharmaceutically effective amount” refers to an amount sufficient to achieve the efficacy or activity of the above-described compound of the present invention or a pharmaceutically acceptable salt thereof.

약학적으로 허용되는 담체는 제제시에 통상적으로 이용되는 것으로서, 락토스, 덱스트로스, 수크로스, 솔비톨, 만니톨, 전분, 아카시아 고무, 인산 칼슘, 알기네이트, 젤라틴, 규산 칼슘, 미세결정성 셀룰로스, 폴리비닐피롤리돈, 셀룰로스, 물, 시럽, 메틸 셀룰로스, 메틸히드록시벤조에이트, 프로필히드록시벤조에이트, 활석, 스테아르산 마그네슘 및 미네랄 오일 등을 포함하나, 이에 한정되는 것은 아니다. 본 발명의 약제학적 조성물은 상기 성분들 이외에 윤활제, 습윤제, 감미제, 향미제, 유화제, 현탁제, 보존제 등을 추가로 포함할 수 있다. 적합한 약제학적으로 허용되는 담체 및 제제는 Remington's Pharmaceutical Sciences (19th ed., 1995)에 상세히 기재되어 있다.Pharmaceutically acceptable carriers are those commonly used in preparation, such as lactose, dextrose, sucrose, sorbitol, mannitol, starch, gum acacia, calcium phosphate, alginate, gelatin, calcium silicate, microcrystalline cellulose, poly Includes, but is not limited to, vinylpyrrolidone, cellulose, water, syrup, methyl cellulose, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and mineral oil. In addition to the above ingredients, the pharmaceutical composition of the present invention may further include lubricants, wetting agents, sweeteners, flavoring agents, emulsifiers, suspending agents, preservatives, etc. Suitable pharmaceutically acceptable carriers and formulations are described in detail in Remington's Pharmaceutical Sciences (19th ed., 1995).

본 발명의 약학적 조성물의 투여 방식은 특별히 제한되지 않으며, 당해 기술 분야에서 통상적으로 사용되는 방식에 따를 수 있다. 보다 구체적으로, 본 발명의 약학적 조성물은 임상 투여시에 경구 및 비경구의 여러 가지 제형으로 투여될 수 있으며, 제제화할 경우에는 보통 사용하는 충진제, 증량제, 결합제, 습윤제, 붕해제, 계면활성제 등의 희석제 또는 부형제를 사용하여 제조될 수 있다.The administration method of the pharmaceutical composition of the present invention is not particularly limited and may follow methods commonly used in the art. More specifically, the pharmaceutical composition of the present invention can be administered in various oral and parenteral dosage forms during clinical administration, and when formulated, commonly used fillers, extenders, binders, wetting agents, disintegrants, surfactants, etc. It may be prepared using diluents or excipients.

상기 약학조성물은 점안제, 주사제, 과립제, 정제, 트로키제, 환제, 캡슐제, 겔, 시럽, 현탁제, 유제, 점적제 및 액제 등으로 이루어진 군에서 선택된 어느 하나의 제형으로 제제화될 수 있다.The pharmaceutical composition may be formulated in any one dosage form selected from the group consisting of eye drops, injections, granules, tablets, troches, pills, capsules, gels, syrups, suspensions, emulsions, drops, and solutions.

경구투여를 위한 고형 제제에는 정제, 환제, 산제, 과립제, 캡슐제, 트로키제 등이 포함되며, 이러한 고형 제제는 하나 이상의 본 발명의 화합물에 적어도 하나 이상의 부형제 예를 들면, 전분, 탄산칼슘, 수크로스(sucrose), 락토오스(lactose) 또는 젤라틴 등을 섞어 조제된다. 또한, 단순한 부형제 외에 마그네슘 스티레이트 탈크 같은 윤활제들도 사용된다. 경구 투여를 위한 액상 제제로는 현탁제, 내용액제, 유제 또는 시럽제 등이 해당되는데, 흔히 사용되는 단순 희석제인 물, 리퀴드 파라핀 이외에 여러 가지 부형제, 예를 들면 습윤제, 감미제, 방향제, 보존제 등이 포함될 수 있다.Solid preparations for oral administration include tablets, pills, powders, granules, capsules, troches, etc. These solid preparations include one or more compounds of the present invention and at least one excipient such as starch, calcium carbonate, water, etc. It is prepared by mixing sucrose, lactose, or gelatin. Additionally, in addition to simple excipients, lubricants such as magnesium styrate talc are also used. Liquid preparations for oral administration include suspensions, oral solutions, emulsions, or syrups. In addition to the commonly used simple diluents such as water and liquid paraffin, they contain various excipients such as wetting agents, sweeteners, fragrances, and preservatives. You can.

비경구 투여를 위한 제제에는 멸균된 수용액, 비수성용제, 현탁용제, 유제, 동결건조제제, 좌제 등이 포함된다. 비수성용제, 현탁용제로는 프로필렌글리콜, 폴리에틸렌 글리콜, 올리브 오일과 같은 식물성 기름, 에틸올레이트와 같은 주사 가능한 에스테르 등이 사용될 수 있다. 좌제의 기제로는 위텝솔(witepsol), 마크로골, 트윈(tween) 61, 카카오지, 라우린지, 글리세롤, 젤라틴 등이 사용될 수 있다.Preparations for parenteral administration include sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, suppositories, etc. Non-aqueous solvents and suspensions may include propylene glycol, polyethylene glycol, vegetable oil such as olive oil, and injectable ester such as ethyl oleate. As a base for suppositories, witepsol, macrogol, tween 61, cacao, laurel, glycerol, gelatin, etc. can be used.

비경구 투여인 경우, 정맥내 주입, 피하 주입, 근육 주입, 복강 주입, 경피 투여, 안구 투여 또는 안구 국소 투여 등의 형태로 주사 투여할 수 있다. In the case of parenteral administration, it can be administered by injection in the form of intravenous injection, subcutaneous injection, intramuscular injection, intraperitoneal injection, transdermal administration, ocular administration, or topical ocular administration.

안구 국소 투여는 예를 들어, 직접적으로 안구내 투약되거나, 안구주위, 안구뒤, 망막하(subretinal), 망막중심 (central retinal), 중심와(fovea) 외부, 결막하 (subconjunctival), 유리체내(intravitreous), 전방내 (intracameral), 또는 맥락막위(suprachoroidal) 등에 투여하는 것을 포함한다.Ocular topical administration, for example, can be administered directly intraocularly, periocularly, retrobulbally, subretinal, central retinal, outside the fovea, subconjunctival, or intravitreous. ), intracameral, or suprachoroidal administration.

또한, 본 발명의 조성물의 인체에 대한 효과적인 투여량은 환자의 나이, 몸무게, 성별, 투여형태, 건강상태 및 질환 정도에 따라 달라질 수 있으며, 본 발명의 조성물에 포함되는 유효성분의 양은 일반적으로 약 0.001-100 mg/kg/일이며, 바람직하게는 0.01-35 mg/kg/일이다. 몸무게가 70㎏인 성인 환자를 기준으로 할 때, 일반적으로 0.07-7000 mg/일이며, 바람직하게는 0.7-2500 ㎎/일이며, 의사 또는 약사의 판단에 따라 일정시간 간격으로 1일 1회 내지 수회로 분할 투여할 수도 있다.In addition, the effective dosage for the human body of the composition of the present invention may vary depending on the patient's age, weight, gender, dosage form, health condition, and degree of disease, and the amount of the active ingredient contained in the composition of the present invention is generally about 0.001-100 mg/kg/day, preferably 0.01-35 mg/kg/day. Based on an adult patient weighing 70 kg, the dose is generally 0.07-7000 mg/day, preferably 0.7-2500 mg/day, and is administered once a day at regular intervals depending on the judgment of the doctor or pharmacist. It may be administered in several divided doses.

황반변성 예방 또는 개선용 식품 조성물Food composition for preventing or improving macular degeneration

본 발명의 조성물은 식품 조성물 또는 건강기능식품 조성물로 제공될 수 있다. The composition of the present invention may be provided as a food composition or health functional food composition.

식품의 종류에는 특별한 제한은 없다. 본 발명의 유효성분을 첨가할 수 있는 식품의 예로는 드링크제, 육류, 소시지, 빵, 비스킷, 떡, 초콜릿, 캔디류, 스낵류, 과자류, 피자, 라면, 기타 면류, 껌류, 아이스크림류를 포함한 낙농제품, 각종 스프, 음료수, 알코올 음료 및 비타민 복합제, 유제품 및 유가공 제품 등이 있으며, 통상적인 의미에서의 식품, 건강식품 및 건강기능성 식품을 모두 포함한다.There are no special restrictions on the type of food. Examples of foods to which the active ingredient of the present invention can be added include drinks, meat, sausages, bread, biscuits, rice cakes, chocolate, candies, snacks, confectionery, pizza, ramen, other noodles, gum, dairy products including ice cream, There are various soups, beverages, alcoholic beverages, vitamin complexes, dairy products and milk products, and it includes all foods, health foods and health functional foods in the conventional sense.

본 발명에 따른 유효성분을 함유하는 식품 및 건강기능성 식품 조성물은 식품에 그대로 첨가하거나 다른 식품 또는 식품 성분과 함께 사용될 수 있고, 통상적인 방법에 따라 적절하게 사용될 수 있다. 유효성분의 혼합량은 그의 사용 목적(예방 또는 개선용)에 따라 적합하게 결정될 수 있다. 일반적으로, 식품 또는 건강기능성 식품 중, 본 발명의 유효성분의 양은 전체 식품 중량(총 100 중량부 기준)의 0.1 내지 90 중량부로 첨가될 수 있다. 그러나 건강 유지를 목적으로 하거나 또는 건강 조절을 목적으로 하는 장기간의 섭취의 경우에는 상기 양은 상기 범위 이하일 수 있으며, 안전성 면에서 아무런 문제가 없기 때문에 유효성분은 상기 범위 이상의 양으로도 사용될 수 있다.Food and health functional food compositions containing the active ingredients according to the present invention can be added as is to food or used together with other foods or food ingredients, and can be used appropriately according to conventional methods. The mixing amount of the active ingredient can be appropriately determined depending on the purpose of use (prevention or improvement). Generally, in food or health functional food, the amount of the active ingredient of the present invention may be added in an amount of 0.1 to 90 parts by weight based on the total weight of the food (based on 100 parts by weight in total). However, in the case of long-term intake for the purpose of maintaining health or regulating health, the amount may be below the above range, and since there is no problem in terms of safety, the active ingredient may be used in an amount above the above range.

본 발명의 식품 및 건강기능식품 조성물은 지시된 비율로 필수 성분으로서 본 발명 유효성분을 함유하는 외에는 다른 성분에는 특별한 제한이 없으며 통상의 음료와 같이 여러 가지 향미제 또는 천연 탄수화물 등을 추가 성분으로서 함유할 수 있다. 상술한 천연 탄수화물의 예는 모노사카라이드, 예를 들어, 포도당, 과당 등; 디사카라이드, 예를 들어 말토스, 슈크로스 등; 및 폴리사카라이드, 예를 들어 덱스트린, 시클로덱스트린 등과 같은 통상적인 당, 및 자일리톨, 소르비톨, 에리트라이톨 등의 당알코올이다. 상술한 것 이외의 향미제로서 천연 향미제(타우마틴, 스테비아 추출물(예를 들어 레바우디오시드 A, 글리시르히진등) 및 합성 향미제(사카린, 아스파르탐 등)를 유리하게 사용할 수 있다. 상기 천연 탄수화물의 비율은 본 발명의 식품 또는 건강기능식품 조성물 100 g당 일반적으로 약 1 내지 20 g, 바람직하게는 약 5 내지 12 g이다.The food and health functional food composition of the present invention has no particular restrictions on other ingredients other than containing the active ingredient of the present invention as an essential ingredient in the indicated ratio, and contains various flavoring agents or natural carbohydrates as additional ingredients like a conventional beverage. can do. Examples of the above-mentioned natural carbohydrates include monosaccharides such as glucose, fructose, etc.; Disaccharides such as maltose, sucrose, etc.; and polysaccharides, such as common sugars such as dextrin and cyclodextrin, and sugar alcohols such as xylitol, sorbitol, and erythritol. As flavoring agents other than those mentioned above, natural flavoring agents (thaumatin, stevia extract (e.g., rebaudioside A, glycyrrhizin, etc.)) and synthetic flavoring agents (saccharin, aspartame, etc.) can be advantageously used. The ratio of the natural carbohydrate is generally about 1 to 20 g, preferably about 5 to 12 g, per 100 g of the food or health functional food composition of the present invention.

상기 외에 본 발명의 유효성분을 함유하는 식품 및 건강기능식품 조성물은 여러 가지 영양제, 비타민, 광물(전해질), 합성 풍미제 및 천연 풍미제 등의 풍미제, 착색제 및 증진제(치즈, 초콜릿 등), 펙트산 및 그의 염, 알긴산 및 그의 염, 유기산, 보호성 콜로이드 증점제, pH 조절제, 안정화제, 방부제, 글리세린, 알코올, 탄산음료에 사용되는 탄산화제 등을 함유할 수 있다. 그 밖에 본 발명의 식품 및 건강기능성식품 조성물은 천연 과일쥬스 및 과일쥬스 음료 및 야채 음료의 제조를 위한 과육을 함유할 수 있다.In addition to the above, food and health functional food compositions containing the active ingredients of the present invention include various nutrients, vitamins, minerals (electrolytes), flavoring agents such as synthetic and natural flavors, colorants and enhancers (cheese, chocolate, etc.), It may contain pectic acid and its salts, alginic acid and its salts, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohol, carbonating agents used in carbonated beverages, etc. In addition, the food and health functional food composition of the present invention may contain pulp for the production of natural fruit juice, fruit juice drinks, and vegetable drinks.

이러한 성분은 독립적으로 또는 조합하여 사용할 수 있다. 이러한 첨가제의 비율은 그렇게 중요하진 않지만 본 발명의 유효성분을 함유하는 식품 및 건강기능식품 조성물 100 중량부 당 0.1 내지 약 20 중량부의 범위에서 선택되는 것이 일반적이다.These ingredients can be used independently or in combination. The ratio of these additives is not that important, but is generally selected in the range of 0.1 to about 20 parts by weight per 100 parts by weight of the food and health functional food composition containing the active ingredient of the present invention.

이하, 본 발명을 하기의 실시예에 의하여 더욱 상세히 설명한다. 단, 하기의 실시예는 본 발명을 예시하는 것일 뿐, 본 발명의 내용이 하기의 실시예에 의해 한정되는 것은 아니다.Hereinafter, the present invention will be described in more detail through the following examples. However, the following examples are merely illustrative of the present invention, and the content of the present invention is not limited by the following examples.

실시예 1. 화합물 합성Example 1. Compound synthesis

1-1 : KS101 화합물 합성1-1: KS101 compound synthesis

단계 1: 1-(3,4-디플루오로페닐)-3-(2-하이드록시페닐)티오우레아 (1-(3,4-difluorophenyl)-3-(2-hydroxyphenyl)thiourea ,FCCS-19025-2-1)의 제조Step 1: 1-(3,4-difluorophenyl)-3-(2-hydroxyphenyl)thiourea (1-(3,4-difluorophenyl)-3-(2-hydroxyphenyl)thiourea ,FCCS-19025 -2-1) Manufacturing

Ar 가스 분위기 하에서 2-아미노페놀(2-Aminophenol) (150 mg, 1.37 mmol)을 메탄올(methanol) (8 mL)에 녹인 후, 3,4-디플루오로페닐 이소티오시아네이트(3,4-difluorophenyl isothiocyanate) (224 ㎕, 1.65 mmol)를 천천히 가한 후, 실온에서 13시간동안 교반하였다. TLC(thin layer chromatography)로 반응종결을 확인한 후, 감압하에서 메탄올(Methanol)을 제거하였다. 반응 혼합물을 크로마토그래피(Silica-gel column chromatrography) (20% Acetone / n-hexane)로 정제하여 1-(3,4-디플루오로페닐)-3-(2-하이드록시페닐)티오우레아 (1-(3,4-difluorophenyl)-3-(2-hydroxyphenyl)thiourea ,FCCS-19025-2-1)를 옅은 노란색의 고체 (354 mg, 92%)로 얻었다.After dissolving 2-Aminophenol (150 mg, 1.37 mmol) in methanol (8 mL) under Ar gas atmosphere, 3,4-difluorophenyl isothiocyanate (3,4- difluorophenyl isothiocyanate) (224 ㎕, 1.65 mmol) was slowly added and stirred at room temperature for 13 hours. After confirming the completion of the reaction by TLC (thin layer chromatography), methanol was removed under reduced pressure. The reaction mixture was purified by silica-gel column chromatography (20% Acetone / n-hexane) to obtain 1-(3,4-difluorophenyl)-3-(2-hydroxyphenyl)thiourea (1 -(3,4-difluorophenyl)-3-(2-hydroxyphenyl)thiourea, FCCS-19025-2-1) was obtained as a pale yellow solid (354 mg, 92%).

1H-NMR (400MHz, MeOH-d4) δ 7.62 (d, J = 8.0 Hz, 1H), 7.55 (ddd, J= 2.4 Hz, 1H), 7.25-7.13 (m, 2H), 7.11-7.05 (m, 1H), 6.92-6.82 (m, 2H); ESI-(+) 281.3 [M+H]+. 1 H-NMR (400 MHz, MeOH-d 4 ) δ 7.62 (d, J = 8.0 Hz, 1H), 7.55 (ddd, J = 2.4 Hz, 1H), 7.25-7.13 (m, 2H), 7.11-7.05 ( m, 1H), 6.92-6.82 (m, 2H); ESI-(+) 281.3 [M+H] + .

단계 2: N-(3,4-디플로오로페닐)벤조[d]옥사졸-2-아민 (N-(3,4-difluorophenyl)benzo[d]oxazol-2-amine ,FCCS-19025)의 제조Step 2: N-(3,4-difluorophenyl)benzo[d]oxazol-2-amine (FCCS-19025) manufacturing

Ar 가스 분위기 하에서 상기 단계 1에서 얻은 FCCS-19025-2-1 (224 mg, 0.80 mmol), 과산화칼륨(Potassium superoxide, KO2) (284 mg, 4.00 mmol)을 아세토니트릴(acetonitrile, MeCN) (25 mL)에 녹인 후, 상온에서 14시간동안 교반하였다. TLC(thin layer chromatography)로 반응종결을 확인한 후, 감압하에서 아세토니트릴(acetonitrile, MeCN)을 제거하였다. 반응 혼합물을 크로마토그래피(Silica-gel column chromatrography) (10 ~ 20% Ethyl acetate / n-hexane)으로 정제하여 목적 화합물 N-(3,4-디플로오로페닐)벤조[d]옥사졸-2-아민 (N-(3,4-difluorophenyl)benzo[d]oxazol-2-amine, FCCS-19025)을 흰색의 고체 (160 mg, 82%)로 얻었다.FCCS-19025-2-1 (224 mg, 0.80 mmol) and potassium peroxide (KO 2 ) (284 mg, 4.00 mmol) obtained in step 1 were mixed with acetonitrile (MeCN) (25) under Ar gas atmosphere. mL) and stirred at room temperature for 14 hours. After confirming the completion of the reaction by TLC (thin layer chromatography), acetonitrile (MeCN) was removed under reduced pressure. The reaction mixture was purified by silica-gel column chromatography (10 ~ 20% Ethyl acetate / n-hexane) to obtain the target compound N-(3,4-difluorophenyl)benzo[d]oxazole-2- Amine (N-(3,4-difluorophenyl)benzo[d]oxazol-2-amine, FCCS-19025) was obtained as a white solid (160 mg, 82%).

[화학식 1-1A][Formula 1-1A]

Figure PCTKR2023003592-appb-img-000013
Figure PCTKR2023003592-appb-img-000013

1H-NMR (400MHz, MeOH-d4) δ 7.82 (ddd, J= 2.8 Hz, 1H), 7.42 (d, J = 7.6 Hz, 1H), 7.36 (d, J = 8.0 Hz, 1H), 7.34-7.29 (m, 1H), 7.28-7.18 (m, 2H), 7.16-7.10 (m, 1H); ESI-(+) 247.2 [M+H]+. 1 H-NMR (400 MHz, MeOH-d 4 ) δ 7.82 (ddd, J = 2.8 Hz, 1H), 7.42 (d, J = 7.6 Hz, 1H), 7.36 (d, J = 8.0 Hz, 1H), 7.34 -7.29 (m, 1H), 7.28-7.18 (m, 2H), 7.16-7.10 (m, 1H); ESI-(+) 247.2 [M+H] + .

1-2 : KS102 화합물 합성1-2: KS102 compound synthesis

THF(2 mL/mmol)에 2-아미노페놀(2-aminophenol, 1.0 equiv.) 및 3,4-디플루오로페닐 이소티오시아네이트(3,4-difluorophenyl isothiocyanate, 1.2 equiv.)를 혼합한 다음, 실온에서 교반하였다. 3시간 동안 교반한 후, 염화제2철(ferric Chloride; FeCl3·6H2O)를 상기 용액에 첨가하고 반응 혼합물을 90℃에서 12시간 동안 교반하였다. 반응 혼합물을 실온으로 냉각시키고 에틸아세테이트(Ethyl acetate, EtOAc)로 추출하였다. 혼합한 유기층을 황산마그네슘(Magnesium sulfate; MgSO4)상에서 건조시키고, 진공에서 농축시켰다. 잔류물을 디클로로메탄(Dichloromethane; CH2Cl2)로 분쇄하고, 디클로로메탄으로 여과 및 세척하여 원하는 생성물(3,4-difluorophenyl isothiocyanate)을 고체로 수득하였다. Mix 2-aminophenol (1.0 equiv.) and 3,4-difluorophenyl isothiocyanate (1.2 equiv.) in THF (2 mL/mmol). , and stirred at room temperature. After stirring for 3 hours, ferric chloride (FeCl 3 ·6H 2 O) was added to the solution, and the reaction mixture was stirred at 90°C for 12 hours. The reaction mixture was cooled to room temperature and extracted with ethyl acetate (EtOAc). The mixed organic layer was dried over magnesium sulfate (MgSO 4 ) and concentrated in vacuum. The residue was triturated with dichloromethane (CH 2 Cl 2 ), filtered and washed with dichloromethane to obtain the desired product (3,4-difluorophenyl isothiocyanate) as a solid.

그 다음, 상기 3,4-디플루오로페닐 이소티오시아네이트(3,4-difluorophenyl isothiocyanate, 210mg, 1.23mmol) 및 2-아미노-4-플루오로페놀(2-amino-4-fluorophenol, 130mg, 1.02mmol)을 반응시켜 하기 화학식 1-1B로 표시되는, KS102 화합물(N-(3,4-difluorophenyl)-5-fluorobenzo[d]oxazol-2-amine)을 밝은 갈색 고체(195mg, 72%)로 얻었다.Next, the 3,4-difluorophenyl isothiocyanate (210 mg, 1.23 mmol) and 2-amino-4-fluorophenol (2-amino-4-fluorophenol, 130 mg, 1.02 mmol) was reacted to produce KS102 compound ( N -(3,4-difluorophenyl)-5-fluorobenzo[ d ]oxazol-2-amine), represented by the following formula 1-1B, as a light brown solid (195 mg, 72%). got it with

[화학식 1-1B][Formula 1-1B]

Figure PCTKR2023003592-appb-img-000014
Figure PCTKR2023003592-appb-img-000014

m.p. 222.9-223.9 ℃; Rf 0.40 (EtOAc/n-hexane = 1:3); HPLC RT 5.15 min (purity: 99.7%); 1H-NMR (DMSO-d6, 800 MHz) δ 6.97 (ddd, J = 9.8, 8.7, 2.6 Hz, 1H), 7.36 (dd, J = 9.0, 2.6 Hz, 1H), 7.43-7.48 (m, 2H), 7.52 (dd, J = 8.7, 4.4 Hz, 1H), 7.91 (ddd, J = 12.8, 7.1, 2.3 Hz, 1H), 11.0 (s, 1H); 13C-NMR (DMSO-d6, 200 MHz) 159.4 (JCF = 234.9 Hz), 159.1, 149.2 (JCF = 241.9, 13.0 Hz), 144.7 (JCF = 239.0, 12.4 Hz), 143.4, 143.2 (JCF = 13.7 Hz), 135.5 (JCF = 9.2, 2.2 Hz), 117.8 (JCF = 17.9 Hz), 114.1 (JCF = 5.8, 3.1 Hz), 109.5 (JCF = 10.3 Hz), 108.5 (JCF = 25.6 Hz), 106.7 (JCF = 22.1 Hz), 103.9 (JCF = 26.3 Hz) ppm, LR-MS (FAB) m/z 265 (M+H)+; HR-MS (FAB) calcd for C13H8F3N2O (M+H)+ 265.0589, found 265.0588.mp 222.9-223.9℃; R f 0.40 (EtOAc/n-hexane = 1:3); HPLC R T 5.15 min (purity: 99.7%); 1 H-NMR (DMSO-d 6 , 800 MHz) δ 6.97 (ddd, J = 9.8, 8.7, 2.6 Hz, 1H), 7.36 (dd, J = 9.0, 2.6 Hz, 1H), 7.43-7.48 (m, 2H), 7.52 (dd, J = 8.7, 4.4 Hz, 1H), 7.91 (ddd, J = 12.8, 7.1, 2.3 Hz, 1H), 11.0 (s, 1H); 13 C-NMR (DMSO-d 6 , 200 MHz) 159.4 (J CF = 234.9 Hz), 159.1, 149.2 (J CF = 241.9, 13.0 Hz), 144.7 (J CF = 239.0, 12.4 Hz), 143.4, 143.2 ( J CF = 13.7 Hz), 135.5 (J CF = 9.2, 2.2 Hz), 117.8 (J CF = 17.9 Hz), 114.1 (J CF = 5.8, 3.1 Hz) , 109.5 (J CF = 10.3 Hz), 108.5 (J CF = 25.6 Hz), 106.7 (J CF = 22.1 Hz), 103.9 (J CF = 26.3 Hz) ppm, LR-MS (FAB) m/z 265 (M+H) + ; HR-MS (FAB) calcd for C 13 H 8 F 3 N 2 O (M+H) + 265.0589, found 265.0588.

1-3 : KS103 화합물 합성1-3: KS103 compound synthesis

상기 실시예 1-2에서 수득한 3,4-디플루오로페닐 이소티오시아네이트(3,4-difluorophenyl isothiocyanate, 210mg, 1.23mmol) 및 2-아미노-5-플루오로페놀(2-amino-5-fluorophenol, 130mg, 1.02mmol)을 반응시켜 하기 화학식 1-1C로 표시되는, KS103 화합물(N-(3,4-difluorophenyl)-6-fluorobenzo[d]oxazol-2-amine)을 상아색 고체(203 mg, 75%)로 얻었다.3,4-difluorophenyl isothiocyanate (210mg, 1.23mmol) and 2-amino-5-fluorophenol (2-amino-5) obtained in Example 1-2 -fluorophenol, 130 mg, 1.02 mmol) was reacted to produce KS103 compound ( N -(3,4-difluorophenyl)-6-fluorobenzo[ d ]oxazol-2-amine), represented by the following formula 1-1C, as an ivory solid (203). mg, 75%).

[화학식 1-1C][Formula 1-1C]

Figure PCTKR2023003592-appb-img-000015
Figure PCTKR2023003592-appb-img-000015

m.p. 231.7-232.6 ℃; Rf 0.40 (EtOAc/n-hexane = 1:3); HPLC RT 5.12 min (purity: 98.4%); 1H-NMR (DMSO-d6, 800 MHz) δ 7.10 (ddd, J = 10.1, 8.6, 2.5 Hz, 1H), 7.42-7.45 (m, 2H), 7.47 (dd, J = 8.6, 4.9 Hz, 1H), 7.54 (dd, J = 8.4, 2.5 Hz, 1H), 7.91 (ddd, J = 12.6, 7.0, 2.1 Hz, 1H), 10.9 (s, 1H); 13C-NMR (DMSO-d6, 200 MHz) 158.1, 158.0 (JCF = 235.9 Hz), 149.2 (JCF = 241.8, 13.2 Hz), 146.8 (JCF = 15.0 Hz), 144.6 (JCF = 238.8, 12.6 Hz), 138.4. 135.7 (JCF = 7.3 Hz), 117.8 (JCF = 17.9 Hz), 116.8 (JCF = 9.6 Hz), 113.8 (JCF = 5.5, 3.3 Hz), 111.0 (JCF = 23.7 Hz), 106.4 (JCF = 22.1 Hz), 98.0 (JCF = 28.9 Hz) ppm, LR-MS (FAB) m/z 265 (M+H)+; HR-MS (FAB) calcd for C13H8F3N2O (M+H)+ 265.0589, found 265.0586.mp 231.7-232.6℃; R f 0.40 (EtOAc/n-hexane = 1:3); HPLC R T 5.12 min (purity: 98.4%); 1 H-NMR (DMSO-d 6 , 800 MHz) δ 7.10 (ddd, J = 10.1, 8.6, 2.5 Hz, 1H), 7.42-7.45 (m, 2H), 7.47 (dd, J = 8.6, 4.9 Hz, 1H), 7.54 (dd, J = 8.4, 2.5 Hz, 1H), 7.91 (ddd, J = 12.6, 7.0, 2.1 Hz, 1H), 10.9 (s, 1H); 13 C-NMR (DMSO-d 6 , 200 MHz) 158.1, 158.0 (J CF = 235.9 Hz), 149.2 (J CF = 241.8, 13.2 Hz), 146.8 (J CF = 15.0 Hz), 144.6 (J CF = 238.8 , 12.6 Hz), 138.4. 135.7 (J CF = 7.3 Hz), 117.8 (J CF = 17.9 Hz), 116.8 (J CF = 9.6 Hz), 113.8 (J CF = 5.5, 3.3 Hz) , 111.0 (J CF = 23.7 Hz), 106.4 (J CF = 22.1 Hz), 98.0 (J CF = 28.9 Hz) ppm, LR-MS (FAB) m/z 265 (M+H) + ; HR-MS (FAB) calcd for C 13 H 8 F 3 N 2 O (M+H) + 265.0589, found 265.0586.

1-4 : KS104 화합물 합성1-4: KS104 compound synthesis

상기 실시예 1-2에서 수득한 3,4-디플루오로페닐 이소티오시아네이트(3,4-difluorophenyl isothiocyanate, 212 mg, 1.24 mmol) 및 2-아미노-4,5-플루오로페놀(2-amino-4,5-difluorophenol, 150 mg, 1.03 mmol)을 반응시켜 하기 화학식 1-1D로 표시되는, KS104 화합물(N-(3,4-difluorophenyl)-5,6-difluorobenzo[d]oxazol-2-amine)을 밝은 보라색 고체(187 mg, 64%)로 얻었다.3,4-difluorophenyl isothiocyanate (212 mg, 1.24 mmol) and 2-amino-4,5-fluorophenol (2- amino-4,5-difluorophenol, 150 mg, 1.03 mmol) was reacted to produce KS104 compound ( N -(3,4-difluorophenyl)-5,6-difluorobenzo[ d ]oxazol-2, represented by the following formula 1-1D -amine) was obtained as a light purple solid (187 mg, 64%).

[화학식 1-1D][Formula 1-1D]

Figure PCTKR2023003592-appb-img-000016
Figure PCTKR2023003592-appb-img-000016

m.p. 242.1-243.2 ℃; Rf 0.40 (EtOAc/n-hexane = 1:3); HPLC RT 5.44 min (purity: 99.1%); 1H-NMR (DMSO-d6, 800 MHz) δ 7.41-7.42 (m, 1H), 7.46 (dd, J = 19.3, 9.0 Hz, 1H), 7.62 (dd, J = 10.5, 7.5 Hz, 1H), 7.83 (dd, J = 9.8, 6.9 Hz, 1H), 7.89 (ddd, J = 13.0, 7.2, 2.5 Hz, 1H), 11.0 (s, 1H); 13C-NMR (DMSO-d6, 200 MHz) 158.9, 149.2 (JCF = 242.0, 13.1 Hz), 147.3 (JCF = 237.0, 13.7 Hz), 145.5 (JCF = 238.0, 14.7 Hz), 144.8 (JCF = 239.0. 13.5 Hz), 142.1 (JCF = 12.6 Hz), 138.0 (JCF = 11.5, 1.5 Hz), 135.4 (JCF = 13.2, 1.8 Hz), 117.8 (JCF = 17.8 Hz), 114.0 ((JCF = 5.7, 3.1 Hz), 106.6 (JCF = 22.1 Hz), 104.9 (JCF = 21.7 Hz), 99.5 (JCF = 24.1 Hz) ppm, LR-MS (FAB) m/z 283 (M+H)+; HR-MS (FAB) calcd for C13H7F4N2O (M+H)+ 283.0495, found 283.0505.mp 242.1-243.2℃; R f 0.40 (EtOAc/n-hexane = 1:3); HPLC R T 5.44 min (purity: 99.1%); 1 H-NMR (DMSO-d 6 , 800 MHz) δ 7.41-7.42 (m, 1H), 7.46 (dd, J = 19.3, 9.0 Hz, 1H), 7.62 (dd, J = 10.5, 7.5 Hz, 1H) , 7.83 (dd, J = 9.8, 6.9 Hz, 1H), 7.89 (ddd, J = 13.0, 7.2, 2.5 Hz, 1H), 11.0 (s, 1H); 13 C-NMR (DMSO-d 6 , 200 MHz) 158.9, 149.2 (J CF = 242.0, 13.1 Hz), 147.3 (J CF = 237.0, 13.7 Hz), 145.5 (J CF = 238.0, 14.7 Hz), 144.8 ( J CF = 239.0. 13.5 Hz), 142.1 (J CF = 12.6 Hz), 138.0 (J CF = 11.5, 1.5 Hz), 135.4 (J CF = 13.2, 1.8 Hz), 117.8 (J CF = 17.8 Hz), 114.0 ((J CF = 5.7, 3.1 Hz), 106.6 (J CF = 22.1 Hz), 104.9 (J CF = 21.7 Hz), 99.5 (J CF = 24.1 Hz) ppm, LR-MS (FAB) m/z 283 ( M+H) + ; HR-MS (FAB) calcd for C 13 H 7 F 4 N 2 O (M+H) + 283.0495, found 283.0505.

실험예 1. 화합물의 안정성 평가Experimental Example 1. Stability evaluation of compounds

실시예 1의 각 KS화합물은 10mM DMSO에 녹여 사용하였고, 표준물질로 사용된 톨부타마이드(tolbutamide)는 10mM농도로 메탄올(methanol)에 녹여 사용하였다.Each KS compound in Example 1 was used dissolved in 10mM DMSO, and tolbutamide, used as a standard substance, was dissolved in methanol at a concentration of 10mM.

유리시험관(13×100 mm)에 0.1 M KH2PO4 (147.5 ㎕), 0.1 M MgCl2 (25 ㎕), corning life science에서 구입한 0.5 mg/ml 농도의 사람 간 마이크로좀(mixed-gender pool) 50 ㎕에 각 KS화합물 용액을 최종 1 μM이 되도록 혼합하여 진탕 항온 수조(shaking water bath, 37℃, 40rpm)에서 3분간 전배양(pre-incubation)한 다음, 10 mM NADPH, 25 ㎕를 가하거나 가하지 않은 조건에서 30분간 배양(incubation) 하였다. In a glass test tube ( 13 ) Mix each KS compound solution in 50 μl to a final concentration of 1 μM, pre-incubate for 3 minutes in a shaking water bath (37°C, 40 rpm), and then add 25 μl of 10 mM NADPH. Incubation was performed for 30 minutes under conditions with or without addition.

반응 전과 반응 후 50 ㎕씩 시료를 취한 다음, 2 ㎖의 아세토니트릴(acetonitrile)를 첨가하여 반응을 멈추고 1분간 볼텍싱(vortex)을 하였다. 혼합액을 20분간 원심분리(4,000rpm, 4℃)후 각각의 상층액을 새로운 유리시험관으로 옮겨 담고, 원심농축기를 사용하여 2시간 동안 건조시켰다. 완전히 건조된 것을 확인하고 각각의 유리시험관에 200 ㎕의 40% 아세토니트릴을 첨가한 후 1분간 볼텍싱하여 재부유 시켰다. 재부유 용액은 10분간 원심분리(4,000rpm, 4℃)후 150 ㎕의 상층액을 취하여 HPLC-UV 분석에 사용하였다. HPLC-UV 분석은 Agilient 1200 series를 사용하였고, MS/MS 분석은 Agilient 6410을 사용하였다. 분석에 사용된 조건은 아래와 같다. 50 μl of sample was taken before and after the reaction, then 2 ml of acetonitrile was added to stop the reaction and vortexed for 1 minute. After centrifuging the mixed solution for 20 minutes (4,000 rpm, 4°C), each supernatant was transferred to a new glass test tube and dried for 2 hours using a centrifugal concentrator. After confirming that it was completely dried, 200 ㎕ of 40% acetonitrile was added to each glass test tube and vortexed for 1 minute to resuspend. The resuspension solution was centrifuged for 10 minutes (4,000 rpm, 4°C), and then 150 ㎕ of the supernatant was taken and used for HPLC-UV analysis. HPLC-UV analysis used the Agilent 1200 series, and MS/MS analysis used the Agilent 6410. The conditions used in the analysis are as follows.

[표 1][Table 1]

HPLC로 분석하기 위한 조건Conditions for analysis by HPLC

Figure PCTKR2023003592-appb-img-000017
Figure PCTKR2023003592-appb-img-000017

[표 2][Table 2]

LC-MS/MS로 분석하기 위한 조건Conditions for analysis by LC-MS/MS

Figure PCTKR2023003592-appb-img-000018
Figure PCTKR2023003592-appb-img-000018

도 1은 사람의 간 마이크로좀과 함께 배양된 KS화합물을 MS/MS분석을 통해 정량적으로 확인한 것이다. Microsome의 효소는 조효소 NADPH(+)에 의존적으로 작동하므로 NADPH(+)가 첨가된 반응에서만 화합물의 대사가 일어났다. KS 화합물들은 control로 사용된 tolbutamide보다 높은 안정성을 나타내었으며, 특히 KS104의 microsomal stability가 가장 우수한 것으로 확인되었다.Figure 1 shows quantitative confirmation of KS compounds cultured with human liver microsomes through MS/MS analysis. Microsome enzymes operate dependent on the coenzyme NADPH(+), so compound metabolism occurred only in reactions where NADPH(+) was added. KS compounds showed higher stability than tolbutamide used as a control, and in particular, KS104 was confirmed to have the best microsomal stability.

실험예 2. KS화합물 처리에 의한 망막색소상피세포 내 Klotho 유전자 발현 효과 확인Experimental Example 2. Confirmation of Klotho gene expression effect in retinal pigment epithelial cells by KS compound treatment

배지에 1μM 농도의 BSO(Buthionine sulfoximine)를 배양과정 중 첨가하여 노화를 촉진 시킨 망막색소상피세포 (Retinal Pigment Epithelium, RPE세포)에, 실시예 1의 각 KS화합물 (KS101, KS102, KS103, KS104 화합물)을 각각 2.5 μM 씩 첨가하여 8시간 배양한 뒤 세포 내 발현된 모든 RNA를 모아 klotho mRNA만 선택적으로 PCR을 통해 발현 변화를 확인하였다. 실험 결과, KS화합물은 모두 secreted (분비) klotho와 membrane (막) klotho의 발현증가를 유도하였다. 특히 KS104의 경우는 secreted klotho 발현유도 능력이 다른 화합물보다 높았다(도 2).Each KS compound (KS101, KS102, KS103, KS104 compounds of Example 1) was added to Retinal Pigment Epithelium (RPE cells), whose aging was accelerated by adding BSO (Buthionine sulfoximine) at a concentration of 1 μM to the medium during the culture process. ) were added at 2.5 μM each and cultured for 8 hours. All RNA expressed in the cells was collected, and changes in expression of only klotho mRNA were selectively confirmed through PCR. As a result of the experiment, all KS compounds induced an increase in the expression of secreted klotho and membrane klotho. In particular, KS104 had a higher ability to induce secreted klotho expression than other compounds (Figure 2).

실험예 3. KS화합물을 장시간 처리한 세포에서의 생존율 변화 확인Experimental Example 3. Confirmation of changes in survival rate in cells treated with KS compound for a long time

배양 중인 망막색소상피세포(AEPE)에 실시예 1의 KS화합물 (KS101, KS102, KS103, KS104 화합물)을 각각 2.5 μM 씩 첨가하여 10 계대 배양한 뒤 BSO를 1μM 농도로 첨가하여 추가적으로 20 계대 배양한 세포의 생존율(viability)를 측정하였다. 배양된 세포 중 살아있는 세포의 비율은 LUNA-II™ 자동세포카운터를 이용하여 측정하였다. LUNA-II™ 자동세포카운터는 tryphan blue라는 시약을 이용하여 살아있는 세포는 이 시약을 받아들이지 않고, 죽은 세포만 받아들여 염색이 되는 것을 이용하여 전체 세포 중 살아있는 세포의 비율을 나타낸다. 그 결과 KS화합물이 첨가된 배지에서 자란 망막색소상피세포는 모두 DMSO가 첨가된 컨트롤 조건에서 자란 세포보다 생존율이 통계적으로 높은 것(p <0.05)을 확인하였다 (도 3).2.5 μM each of the KS compounds of Example 1 (KS101, KS102, KS103, and KS104 compounds) were added to the retinal pigment epithelial cells (AEPE) in culture, and the cells were subcultured for 10 times. Then, BSO was added at a concentration of 1 μM and cultured for an additional 20 passages. Cell viability was measured. The proportion of living cells among cultured cells was measured using a LUNA-II™ automatic cell counter. LUNA-II™ automatic cell counter uses a reagent called tryphan blue. Living cells do not accept this reagent, but only dead cells are stained, indicating the ratio of live cells among all cells. As a result, it was confirmed that the survival rate of retinal pigment epithelial cells grown in medium containing KS compound was statistically higher ( p <0.05) than that of cells grown in control conditions containing DMSO ( Fig. 3 ).

실험예 4. KS화합물을 장시간 처리한 세포 내에서의 활성산소 변화 확인Experimental Example 4. Confirmation of changes in active oxygen in cells treated with KS compound for a long time

배양 중인 망막색소상피세포에 실시예 1의 KS화합물 (KS101, KS102, KS103, KS104 화합물)을 각각 2.5 μM 씩 첨가하여 10 계대 배양한 뒤 BSO를 1μM 농도로 첨가하여 추가적으로 20 계대 배양한 세포 내의 활성산소(Reactive Oxygen Species, ROS) 농도를 측정하였다. 측정시약은 Molecular Probes 사의 ROS detection reagents를 이용하였고, 제조사에서 요구하는 방법을 통해 활성산소와 반응하여 생성되는 색소를 분광광도계를 이용하여 흡광도를 측정하여 농도를 측정하였다. 실험을 통해 장기간(10 계대 배양 기간) KS화합물이 첨부된 배지에서 자란 망막색소상피세포에서는 DMSO가 첨가된 컨트롤 조건에서 자란 세포보다 세포 내 활성산소량이 통계적으로 낮은 것(p <0.05)을 확인하였다 (도 4). 2.5 μM each of the KS compounds of Example 1 (KS101, KS102, KS103, and KS104 compounds) were added to the retinal pigment epithelial cells in culture, and the cells were subcultured for 10 passages. BSO was added at a concentration of 1 μM, and BSO was added at a concentration of 1 μM for an additional 20 passages. Activity in the cells Oxygen (Reactive Oxygen Species, ROS) concentration was measured. ROS detection reagents from Molecular Probes were used as measurement reagents, and the concentration was measured by measuring the absorbance of the pigment produced by reacting with active oxygen using a spectrophotometer according to the method required by the manufacturer. Through experiments, it was confirmed that the amount of intracellular reactive oxygen species was statistically lower ( p <0.05) in retinal pigment epithelial cells grown in medium supplemented with KS compounds for a long period of time (10 subculture periods) than in cells grown in control conditions supplemented with DMSO. (Figure 4).

실험예 5. KS화합물을 단시간 처리한 세포 내에서의 활성산소 변화 확인Experimental Example 5. Confirmation of changes in active oxygen in cells treated with KS compound for a short period of time

배양 중인 망막색소상피세포에 실시예 1의 KS화합물 (KS101, KS102, KS103, KS104 화합물)을 각각 2.5 μM 씩 첨가하고 6시간 추가 배양한 뒤에 세포 내의 활성산소(Reactive Oxygen Species, ROS) 농도를 실험 4에서와 동일하게 측정하였다. 실험을 통해 단시간 KS화합물이 첨부된 배지에서 자란 망막색소상피세포에서는 DMSO가 첨가된 컨트롤 조건에서 자란 세포보다 세포 내 활성산소의 양이 통계적으로 낮은 것(p <0.05)을 확인하였다. 이를 통해 6시간이라는 짧은 처리 시간에서도 KS화합물들이 세포 내 활성산소량을 감소시킬 수 있음을 확인하였다 (도 4).2.5 μM each of the KS compounds (KS101, KS102, KS103, KS104 compounds) of Example 1 were added to the retinal pigment epithelial cells in culture, and the concentration of reactive oxygen species (ROS) in the cells was tested after culturing for an additional 6 hours. Measurements were made in the same way as in 4. Through the experiment, it was confirmed that the amount of intracellular reactive oxygen species in retinal pigment epithelial cells grown in medium supplemented with KS compound for a short period of time was statistically lower ( p <0.05) than in cells grown in control conditions to which DMSO was added. Through this, it was confirmed that KS compounds can reduce the amount of reactive oxygen species in cells even with a short treatment time of 6 hours (Figure 4).

실험예 6. 세포독성(cytotoxicity) 개선 효과 확인Experimental Example 6. Confirmation of cytotoxicity improvement effect

실시예 1의 KS화합물 (KS101, KS102, KS103, KS104 화합물)의 세포독성을 평가하기 위해 이들 화합물 개발의 모체가 되었던 compound H를 이용하여 비교하였다. 세포독성은 두젠생명공학연구소 제품인 EZ-CYTOX라는 시약을 이용하여 측정하였다. 이 시약을 통해 살아있는 세포에서만 활성이 있는 dehydrogenase를 통해 오랜지색 수용성 물질 생기는 것을 405 nm에서 흡광도를 측정하여 정량하였다. 실험결과 compound H의 경우 5 μM 정도의 농도에서도 세포독성이 뚜렷하게 나타나는 반면에, KS화합물들은 10 μM 까지도 큰 독성이 나타나지 않는 것을 확인하였다. 특히 KS104의 경우는 40 μM 농도에서도 세포독성이 높지 않은 것을 확인하였다 (도 6).To evaluate the cytotoxicity of the KS compounds (KS101, KS102, KS103, and KS104 compounds) of Example 1, they were compared using compound H, which was the basis for the development of these compounds. Cytotoxicity was measured using a reagent called EZ-CYTOX, a product of Dugen Biotechnology Research Institute. Using this reagent, the formation of an orange-colored water-soluble substance through dehydrogenase, which is active only in living cells, was quantified by measuring the absorbance at 405 nm. As a result of the experiment, it was confirmed that compound H showed clear cytotoxicity even at a concentration of about 5 μM, while KS compounds did not show significant toxicity even at a concentration of 10 μM. In particular, in the case of KS104, it was confirmed that cytotoxicity was not high even at a concentration of 40 μM (Figure 6).

실험예 7. BSO로 유발된 산화스트레스로부터 KS화합물들의 세포 보호 효과 확인Experimental Example 7. Confirmation of the cell protection effect of KS compounds from BSO-induced oxidative stress

세포의 산화스트레스 유발물질인 BSO는 세포 내 활성산소 제거 물질인 glutathione(GSH)의 생산을 억제한다. BSO에 의한 세포독성으로부터 세포를 보호하는 효과를 확인하기 위해 배양 중인 망막색소상피세포에 실시예 1의 KS화합물 (KS101, KS102, KS103, KS104 화합물)을 각각 2.5 μM 씩 첨가한 후 3시간 배양하였다. 이 세포들에 예비 실험을 통해 확인된 세포 독성을 유발하는 농도의 BSO(50mM)를 추가적으로 처리하였다. 이후 6시간 배양 뒤 세포 독성을 EZ-CYTOX 시약으로 측정하였다. 그 결과 KS화합물 모두 세포보호 효과가 DMSO처리한 대조군에 비해 통계적으로 높은 것(p <0.05)을 확인하였다 (도 7).BSO, a substance that causes cellular oxidative stress, inhibits the production of glutathione (GSH), an intracellular oxygen radical scavenging substance. To confirm the effect of protecting cells from cytotoxicity caused by BSO, 2.5 μM each of the KS compounds (KS101, KS102, KS103, KS104 compounds) of Example 1 were added to retinal pigment epithelial cells in culture, and cultured for 3 hours. . These cells were additionally treated with BSO (50mM) at a concentration that induces cytotoxicity, which was confirmed through preliminary experiments. After 6 hours of incubation, cytotoxicity was measured using EZ-CYTOX reagent. As a result, it was confirmed that the cytoprotective effect of all KS compounds was statistically higher ( p <0.05) than the DMSO-treated control group (Figure 7).

실험예 8. NaIOExperimental Example 8. NaIO 33 로 유발된 산화스트레스로부터 KS화합물들의 세포 보호 효과 확인Confirmation of the cell protection effect of KS compounds from oxidative stress induced by

세포 내 산화스트레스 유발물질인 NaIO3는 망막색소상피세포에서 노인성황반변성(age-related macular degeneration, AMD)을 일으키는 물질로 알려져 있다. 산화성 세포독성을 일으키는 농도의 NaIO3로부터 세포를 보호하는 KS화합물들의 효과를 확인하였다. 이를 위해 망막색소상피세포에 실시예 1의 KS화합물(KS101, KS102, KS103, KS104 화합물)을 각각 2.5 μM 씩 첨가한 후 3시간 배양하고 이 세포들에 5 mM 농도로 NaIO3를 첨가하고 24시간 추가 배양하였다. 이후 세포들에서의 독성보호 효과를 EZ-CYTOX 시약으로 측정하였다. 그 결과 KS화합물 모두 세포보호 효과가 DMSO처리한 대조군에 비해 통계적으로 높은 것(p <0.05)을 확인하였다 (도 8). NaIO 3 , a substance that causes intracellular oxidative stress, is known to be a substance that causes age-related macular degeneration (AMD) in retinal pigment epithelial cells. The effectiveness of KS compounds in protecting cells from NaIO 3 at concentrations that cause oxidative cytotoxicity was confirmed. For this purpose, 2.5 μM each of the KS compounds of Example 1 (KS101, KS102, KS103, and KS104 compounds) were added to the retinal pigment epithelial cells and cultured for 3 hours. NaIO 3 was added at a concentration of 5 mM to the cells and incubated for 24 hours. Additional culture was performed. Afterwards, the toxicity protection effect in cells was measured using EZ-CYTOX reagent. As a result, it was confirmed that the cytoprotective effect of all KS compounds was statistically higher ( p <0.05) than the DMSO-treated control group (FIG. 8).

Claims (12)

하기 화학식 1로 표시되는 화합물 또는 이의 약학적으로 허용가능한 염을 유효성분으로 포함하는, 황반변성 예방 또는 치료용 약학적 조성물:A pharmaceutical composition for preventing or treating macular degeneration, comprising a compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient: [화학식 1][Formula 1]
Figure PCTKR2023003592-appb-img-000019
Figure PCTKR2023003592-appb-img-000019
 (상기 화학식 1에서 (In Formula 1 above, R1 및 R2는 각각 -H, 할로겐 또는 C1-6의 직쇄 또는 측쇄 알킬이고,R 1 and R 2 are each -H, halogen or C 1-6 straight or branched alkyl, R3 및 R4는 각각 -H 또는 할로겐이다).R 3 and R 4 are each -H or halogen). 제1항에 있어서, According to paragraph 1, 상기 화학식 1로 표시되는 화합물은 하기 화학식 1-1로 표시되는 화합물인 것을 특징으로 하는, 황반변성 예방 또는 치료용 약학적 조성물: A pharmaceutical composition for preventing or treating macular degeneration, characterized in that the compound represented by Formula 1 is a compound represented by the following Formula 1-1: [화학식 1-1][Formula 1-1]
Figure PCTKR2023003592-appb-img-000020
Figure PCTKR2023003592-appb-img-000020
 (상기 화학식 1-1에서,(In Formula 1-1 above, R1 및 R2는 각각 -H, 할로겐 또는 C1-6의 직쇄 또는 측쇄 알킬이다).R 1 and R 2 are each -H, halogen or C 1-6 straight or branched alkyl). 제2항에 있어서, According to paragraph 2, 상기 화학식 1-1로 표시되는 화합물은, The compound represented by Formula 1-1 is, 하기 화학식 1-1A로 표시되는 화합물;A compound represented by the following formula 1-1A; [화학식 1-1A][Formula 1-1A]
Figure PCTKR2023003592-appb-img-000021
Figure PCTKR2023003592-appb-img-000021
 하기 화학식 1-1B로 표시되는 화합물;A compound represented by the following formula 1-1B; [화학식 1-1B][Formula 1-1B]
Figure PCTKR2023003592-appb-img-000022
Figure PCTKR2023003592-appb-img-000022
 하기 화학식 1-1C로 표시되는 화합물; 및A compound represented by the following formula 1-1C; and [화학식 1-1C][Formula 1-1C]
Figure PCTKR2023003592-appb-img-000023
Figure PCTKR2023003592-appb-img-000023
 하기 화학식 1-1D로 표시되는 화합물;Compounds represented by the following formula 1-1D; [화학식 1-1D][Formula 1-1D]
Figure PCTKR2023003592-appb-img-000024
Figure PCTKR2023003592-appb-img-000024
 로 구성된 군에서 선택된 어느 하나인 것을 특징으로 하는, 황반변성 예방 또는 치료용 약학적 조성물.A pharmaceutical composition for preventing or treating macular degeneration, characterized in that it is any one selected from the group consisting of. 제1항에 있어서, According to paragraph 1, 상기 조성물은 클로소(Klotho) 유전자의 발현을 증가시키는 것을 특징으로 하는, 황반변성 예방 또는 치료용 약학적 조성물.The composition is a pharmaceutical composition for preventing or treating macular degeneration, characterized in that it increases the expression of the Klotho gene. 제1항에 있어서, According to paragraph 1, 상기 조성물은 산화스트레스로부터 망막색소상피세포를 보호하는 것을 특징으로 하는, 황반변성 예방 또는 치료용 약학적 조성물.The composition is a pharmaceutical composition for preventing or treating macular degeneration, characterized in that it protects retinal pigment epithelial cells from oxidative stress. 하기 화학식 1로 표시되는 화합물 또는 이의 약학적으로 허용가능한 염을 유효성분으로 포함하는, 황반변성 예방 또는 개선용 식품 조성물:A food composition for preventing or improving macular degeneration, comprising a compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient: [화학식 1][Formula 1]
Figure PCTKR2023003592-appb-img-000025
Figure PCTKR2023003592-appb-img-000025
 (상기 화학식 1에서 (In Formula 1 above, R1 및 R2는 각각 -H, 할로겐 또는 C1-6의 직쇄 또는 측쇄 알킬이고,R 1 and R 2 are each -H, halogen or C 1-6 straight or branched alkyl, R3 및 R4는 각각 -H 또는 할로겐이다).R 3 and R 4 are each -H or halogen). 제6항에 있어서, According to clause 6, 상기 화학식 1로 표시되는 화합물은 하기 화학식 1-1로 표시되는 화합물인 것을 특징으로 하는, 황반변성 예방 또는 개선용 식품 조성물: A food composition for preventing or improving macular degeneration, characterized in that the compound represented by Formula 1 is a compound represented by the following Formula 1-1: [화학식 1-1][Formula 1-1]
Figure PCTKR2023003592-appb-img-000026
Figure PCTKR2023003592-appb-img-000026
 (상기 화학식 1-1에서,(In Formula 1-1 above, R1 및 R2는 각각 -H, 할로겐 또는 C1-6의 직쇄 또는 측쇄 알킬이다).R 1 and R 2 are each -H, halogen or C 1-6 straight or branched alkyl). 제7항에 있어서, In clause 7, 상기 화학식 1-1로 표시되는 화합물은, The compound represented by Formula 1-1 is, 하기 화학식 1-1A로 표시되는 화합물;A compound represented by the following formula 1-1A; [화학식 1-1A][Formula 1-1A]
Figure PCTKR2023003592-appb-img-000027
Figure PCTKR2023003592-appb-img-000027
 하기 화학식 1-1B로 표시되는 화합물;A compound represented by the following formula 1-1B; [화학식 1-1B][Formula 1-1B]
Figure PCTKR2023003592-appb-img-000028
Figure PCTKR2023003592-appb-img-000028
 하기 화학식 1-1C로 표시되는 화합물; 및A compound represented by the following formula 1-1C; and [화학식 1-1C][Formula 1-1C]
Figure PCTKR2023003592-appb-img-000029
Figure PCTKR2023003592-appb-img-000029
 하기 화학식 1-1D로 표시되는 화합물;Compounds represented by the following formula 1-1D; [화학식 1-1D][Formula 1-1D]
Figure PCTKR2023003592-appb-img-000030
Figure PCTKR2023003592-appb-img-000030
 로 구성된 군에서 선택된 어느 하나인 것을 특징으로 하는, 황반변성 예방 또는 개선용 식품 조성물.A food composition for preventing or improving macular degeneration, characterized in that it is any one selected from the group consisting of. 제6항에 있어서, According to clause 6, 상기 조성물 클로소(Klotho) 유전자의 발현정도를 증가시키는 것을 특징으로 하는, 황반변성 예방 또는 개선용 식품 조성물.A food composition for preventing or improving macular degeneration, characterized in that the composition increases the expression level of the Klotho gene. 제6항에 있어서, According to clause 6, 상기 조성물은 산화스트레스로부터 망막색소상피세포를 보호하는 것을 특징으로 하는, 황반변성 예방 또는 개선용 식품 조성물.The composition is a food composition for preventing or improving macular degeneration, characterized in that it protects retinal pigment epithelial cells from oxidative stress. 하기 화학식 1로 표시되는 화합물 또는 이의 약학적으로 허용가능한 염을 유효성분으로 포함하는 조성물을 개체에 투여하는 단계를 포함하는 황반변성 예방 또는 치료 방법. A method for preventing or treating macular degeneration, comprising administering to an individual a composition comprising a compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient. [화학식 1][Formula 1]
Figure PCTKR2023003592-appb-img-000031
Figure PCTKR2023003592-appb-img-000031
 (상기 화학식 1에서 (In Formula 1 above, R1 및 R2는 각각 -H, 할로겐 또는 C1-6의 직쇄 또는 측쇄 알킬이고,R 1 and R 2 are each -H, halogen or C 1-6 straight or branched alkyl, R3 및 R4는 각각 -H 또는 할로겐이다).R 3 and R 4 are each -H or halogen). 하기 화학식 1로 표시되는 화합물 또는 이의 약학적으로 허용가능한 염을 유효성분으로 포함하는 조성물의 황반변성 예방 또는 치료 용도. Use for the prevention or treatment of macular degeneration of a composition comprising a compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient. [화학식 1][Formula 1]
Figure PCTKR2023003592-appb-img-000032
Figure PCTKR2023003592-appb-img-000032
 (상기 화학식 1에서 (In Formula 1 above, R1 및 R2는 각각 -H, 할로겐 또는 C1-6의 직쇄 또는 측쇄 알킬이고,R 1 and R 2 are each -H, halogen or C 1-6 straight or branched alkyl, R3 및 R4는 각각 -H 또는 할로겐이다).R 3 and R 4 are each -H or halogen).
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