KR20200083134A - Novel isoquinoline derivative, preparing method thereof, and pharmaceutical composition for preventing or treating autophagy related diseases containing the same as an active ingredient - Google Patents

Abstract

The present invention relates to a novel isoquinoline derivative, a method for manufacturing the same, and a pharmaceutical composition for preventing or treating autophagy-related diseases containing the same as an active component. A novel isoquinoline furanone derivative according to the present invention can control autophagy activity. Therefore, by containing the novel isoquinoline furanone derivative as the active component, there is a useful effect of being provided as the pharmaceutical composition for preventing or treating autophagy-related diseases, such as neurodegenerative diseases, cancer, metabolic diseases, inflammatory diseases, or melanogenesis-related diseases, a health functional food composition for alleviating autophagy-related diseases, or a cosmetic composition having a whitening function.

Description

Novel isoquinoline derivative, preparing method thereof, and pharmaceutical composition for preventing or treating autophagy related diseases containing the same as an active ingredient}

The present invention relates to a novel isoquinoline derivative, a method for manufacturing the same, and a pharmaceutical composition for preventing or treating autophagy related diseases containing the same as an active ingredient.

Autophagy (autophagy) plays an important role in regulating cell functions such as survival in starvation, protection from infectious bacteria, and regulation of neurodecay. Autophagy is an evolutionarily conserved process known to occur in all eukaryotic cells, from yeast to mammals.

When the autophagy is activated, the autophagosome membrane structure is formed by the atg12-atg5 complex and LC3 aggregation. The cytosolic form of LC3 (LC3-I) is split into a membrane bound form (LC3-II) and the membrane matures into an autophagosome surrounding the component to be degraded. The autopagosome is then fused with the lysosome to form an autolysosome, which causes lysosomal degradation of intracellular components. Various studies are currently underway to identify the complex molecular mechanisms involved in this process.

On the other hand, autophages are associated with various pathological processes such as neurodegenerative diseases, cancer and melanogenesis. In cancer, autophages can be involved in a variety of stages, and because oxygen and nutrients are limited in growing tumor cells, autophages are used to survive until the necessary components are provided by angiogenesis. In this case, autophagy should be suppressed to stop tumor cell survival.

Melanogenesis refers to the formation of melanin, a pigment found in the eyes, skin and hair. An increase in autophagosomes is observed in patients with pigmented disease, and more recent studies have reported that the lack of becklin 1 or LC3-I significantly attenuates pigment accumulation. In the in vivo assay, lack of Becklin 1 heterozygosity caused a mouse coat pigment defect. As an external factor of melanin production, it is activated by stimulation such as UV. Pigmentation diseases (freckles, freckles, blotch, etc.) may occur due to melanin production. In this case, autophagy should be inhibited to stop melanin production.

As described above, in neurodegenerative diseases related to autophages, cancer and melanin-producing diseases, and the like, it is possible to develop drugs having the ability to control autophages as potential pharmaceutical candidates.

Meanwhile, as a patent related to Beclin 1 involved in an autophagy (autophagy) mechanism, EP 2383294 discloses a technique for treating or diagnosing neurodegenerative diseases using an antibody that specifically binds Thr119 phosphorylation of Beclin 1, US 6432914 discloses a technique for treating cancer using Beclin 1 itself as a protein drug, and US 5858669 discloses a technique for diagnosing cancer using a mutation in Beclin 1, but still uses autophagy-regulating drugs. Development of disease treatment is required.

EP 2383294 US 6432914 US 5858669

An object of the present invention is to provide a novel isoquinoline derivative as an active ingredient for providing an autophagy activity modulating drug.

Another object of the present invention is to provide a method for preparing the isoquinoline derivative.

Another object of the present invention is to provide a pharmaceutical composition for the prevention or treatment of autophagy-related diseases containing the isoquinoline derivative as an active ingredient.

Another object of the present invention is to provide a pharmaceutical composition for preventing or treating inflammatory diseases or metabolic diseases containing the isoquinoline derivative as an active ingredient.

Another object of the present invention is to provide a pharmaceutical composition for preventing or treating skin hyperpigmentation disease, which contains the isoquinoline derivative as an active ingredient.

In order to achieve the above object,

In one aspect of the invention,

A compound represented by Formula 1 below, or a pharmaceutically acceptable salt thereof is provided.

[Formula 1]

(In the above formula 1,

R ¹ is substituted or unsubstituted phenyl,

The substituted phenyl is substituted with one or more substituents selected from the group consisting of hydroxy, cyano, nitro, amino, halogen and substituted or unsubstituted C _1-5 alkyl of straight or branched chain,

Here, the substituted alkyl is substituted with halogen;

R ² is hydroxy, amino, cyano, halogen, C _1-3 straight or branched chain alkyl, or C _1-3 straight or branched alkoxy;

n is 0 to 8;

X and Y are each independently CH or N; And

Z is S, sulfinyl or sulfonyl).

In addition, in another aspect of the invention,

As shown in Scheme 1 below,

A method for preparing a compound represented by Formula 1, including a step of preparing a compound represented by Formula 1 from a compound represented by Formula 4 is provided.

[Scheme 1]

(In Scheme 1,

R ¹ , R ² , n, X, Y and Z are as defined in Formula 1).

Furthermore, in another aspect of the invention,

Provided is a pharmaceutical composition for preventing or treating autophagy-related diseases containing the compound represented by Formula 1 or a pharmaceutically acceptable salt thereof as an active ingredient.

In addition, in another aspect of the invention,

Provided is a pharmaceutical composition for preventing or treating an inflammatory disease or metabolic disease, which contains the compound represented by Formula 1 or a pharmaceutically acceptable salt thereof as an active ingredient.

Furthermore, in another aspect of the invention,

Provided is a pharmaceutical composition for preventing or treating skin hyperpigmentation disease, which contains the compound represented by Formula 1 or an acceptable salt thereof as an active ingredient.

The novel isoquinoline furanone derivative according to the present invention is capable of regulating autophagy activity, and as an active ingredient, autophagy related diseases, for example, neurodegenerative diseases, cancer, metabolic diseases, inflammatory diseases, or melanin formation There is a useful effect that can be provided as a pharmaceutical composition for preventing or treating a disease, or a health functional food composition for improvement, or a cosmetic composition having a whitening function.

1 is a Western blot graph confirming the conversion of LC3-I and LC3-II according to Example 1 Compounds 1, 5, 10, and 20 μM treatment according to the present invention to Raw 264.7 mouse macrophages.
Figure 2 after adjusting the inflammatory response to Raw 264.7 mouse macrophages (LPS treatment), 5, 10, 20, 50 μM treatment of Example Compound 1 of the present invention, fluorescence micrograph confirming the change in the observed auto-pagosome to be.
Figure 3 (a): After the production of NO by LPS (control of inflammatory reaction), Example 1 compounds of the present invention were treated with 0.1, 0.5, 1, 10, and 100 μM, respectively, and absorbance was measured using a grease reagent. Graph showing the measured NO production rate as a percentage in terms of LPS, and (b): control the cell viability according to 0.1, 0.5, 1, 10, and 100 μM treatment of LPS and Example 1 compounds, respectively, respectively. Control).
Figure 4 is a graph showing the measurement of the gene expression level of the cytokine IL-6 related to the inflammatory response by the compound treatment of Example 1 according to the present invention.
Figure 5 is a graph showing the protein measurement of the protein IL-6 and IL-1β related to the inflammatory response by Example 1 of the present invention compound 0.1, 1, and 10 μM.
Figure 6 (a): a photograph of a zebrafish that confirmed the inhibitory effect of melanocyte formation at 32 hours after fertilization after 22 hours of treatment with the compound of Example 1 of the present invention, targeting the zebrafish fertilized egg at 10 hours after fertilization. , And (b) is a photograph of a zebrafish, which was confirmed for the melanin synthesis inhibitory effect at 72 hours after fertilization after 12 hours of treatment with the compound of Example 1 of the present invention, targeting the zebrafish pom at 60 hours after fertilization. .
FIG. 7 is a view of a normal microscope image, a fluorescence microscope image, and a heat map conversion image observed after treating DMSO, tamoxifen, and Example 1 of the present invention for a zebrafish cheer 5 days after fertilization.
FIG. 8 shows the liver area (a) and fluorescence intensity (b) on the observed zebrafish cheer image after treating DMSO, tamoxifen, and Example 1 of the present invention for the zebrafish cheer 5 days after fertilization, respectively. It is a graph showing the results quantified using the software (National Institutes of Health, USA) (p value is the t-test result for each experimental group compared to the tamoxifen treated group. *p ≤ 0.01, **p ≤ 0.001).
9 is a graph showing the cell viability according to the concentration-dependent treatment of the compound of Example 1 of the present invention for VERO, HFL-1, L929, NIH 3T3, CHO-K1, and HMV-II cell lines.
10 is a graph showing the binding and inhibitory activity of the hug channel protein by the compound of Example 1 of the present invention by ligand binding and patch clamp curves, respectively.
11 shows the survival rate of vehicle, ARB, SDS, Example 1 compounds 10, 20, and 50 μM treatment group based on the measured absorbance after eluting MTT stained in artificial skin tissue with isopropanol. It is a graph.
FIG. 12 shows the survival rate of vehicle, ARB, acetic acid, Example 1 compound 10, 20, and 50 μM treated group based on the measured absorbance after eluting MTT stained in artificial corneal tissue with isopropanol. It is a graph.
13 is a graph showing the measurement of ALT (aminotransferase) activity as an indicator of liver damage after administration of a vehicle, a positive control (10 mpk Rosi), and a compound of Example 1 (10 mpk and 100 mpk) of the present invention, respectively, in an obese-controlled rodent model. to be.
Figure 14 in the obesity-controlled rodent model, respectively, after administration of the vehicle, the positive control (10 mpk Rosi), the present invention Example 1 compound (10 mpk and 100 mpk), the activity of the oxidative stress indicator TBARS (Thiobarbituric acid reactive substance) It is a graph that is measured and shown.

Hereinafter, the present invention will be described in detail.

The following description is provided to help the understanding of the invention, and the present invention is not limited to the contents of the following description.

In one aspect of the invention,

A compound represented by Formula 1 below, or a pharmaceutically acceptable salt thereof is provided.

[Formula 1]

(In the above formula 1,

R ¹ is substituted or unsubstituted phenyl,

The substituted phenyl is substituted with one or more substituents selected from the group consisting of hydroxy, cyano, nitro, amino, halogen and substituted or unsubstituted C _1-5 alkyl of straight or branched chain,

Here, the substituted alkyl is substituted with halogen;

R ² is hydroxy, amino, cyano, halogen, C _1-3 straight or branched chain alkyl, or C _1-3 straight or branched alkoxy;

n is 0 to 8;

X and Y are each independently CH or N; And

Z is S, sulfinyl or sulfonyl).

In one embodiment of the invention,

R ¹ may be phenyl substituted with one or more substituents selected from the group consisting of F, Cl, Br, I, and CF ₃ .

In another embodiment of the invention,

When any one of X and Y is CH, the other may be N.

In another embodiment of the invention,

When X is CH, Y is N.

In another embodiment of the invention,

The compound represented by Formula 1 may be a compound, or a pharmaceutically acceptable salt thereof, characterized in that it is any one selected from the following group of compounds:

(1) 1-((4-fluorophenyl)sulfinyl)isoquinoline;

(2) 1-((4-fluorophenyl)thio)isoquinoline;

(3) 1-((4-fluorophenyl)sulfonyl)isoquinoline;

(4) 4-((4-fluorophenyl)sulfinyl)quinoline;

(5) 1-((2,4-difluorophenyl)sulfinyl)isoquinoline;

(6) 1-((4-chlorophenyl)sulfinyl)isoquinoline; And

(7) 1-((4-(trifluoromethyl)phenyl)sulfinyl)isoquinoline.

The compound represented by Formula 1 of the present invention can be used in the form of a pharmaceutically acceptable salt, and as the salt, an acid addition salt formed by a pharmaceutically acceptable free acid is useful. Acid addition salts include inorganic acids such as hydrochloric acid, nitric acid, phosphoric acid, sulfuric acid, hydrobromic acid, hydroiodic acid, nitrous acid and phosphorous acid, aliphatic mono and dicarboxylates, phenyl-substituted alkanoates, hydroxy alkanoates and alkanes It is obtained from non-toxic organic acids such as dioate, aromatic acids, aliphatic and aromatic sulfonic acids, acetic acid, benzoic acid, citric acid, lactic acid, maleic acid, gluconic acid, methanesulfonic acid, 4-toluenesulfonic acid, tartaric acid, fumaric acid and the like. The types of pharmaceutically non-toxic salts include sulfate, pyrosulfate, bisulfate, sulfite, bisulfite, nitrate, phosphate, monohydrogen phosphate, dihydrogen phosphate, metaphosphate, pyrophosphate chloride, bromide, and eye Odide, fluoride, acetate, propionate, decanoate, caprylate, acrylate, formate, isobutyrate, caprate, heptanoate, propiolate, oxalate, malonate, succinate, sube Late, sebacate, fumarate, maleate, butyne-1,4-dioate, hexane-1,6-dioate, benzoate, chlorobenzoate, methylbenzoate, dinitro benzoate, hydroxybenzoate, Methoxybenzoate, phthalate, terephthalate, benzenesulfonate, toluenesulfonate, chlorobenzenesulfonate, xylenesulfonate, phenylacetate, phenylpropionate, phenylbutyrate, citrate, lactate, β-hydroxybutyrate, Glycolate, malate, tartrate, methanesulfonate, propanesulfonate, naphthalene-1-sulfonate, naphthalene-2-sulfonate, mandelate and the like.

The acid addition salt according to the present invention can be prepared by a conventional method, for example, a derivative of Formula 1 is dissolved in an organic solvent such as methanol, ethanol, acetone, dichloromethane, acetonitrile, etc. and a precipitate formed by adding an organic acid or inorganic acid It can be prepared by filtration and drying, or by distilling the solvent and excess acid under reduced pressure and drying to crystallize under an organic solvent.

In addition, bases can be used to make pharmaceutically acceptable metal salts. The alkali metal or alkaline earth metal salt is obtained, for example, by dissolving the compound in an excess of an alkali metal hydroxide or alkaline earth metal hydroxide solution, filtering the inexpensive compound salt, and evaporating and drying the filtrate. At this time, it is suitable to manufacture sodium, potassium or calcium salts as metal salts. Further, the corresponding salt is obtained by reacting an alkali metal or alkaline earth metal salt with a suitable negative salt (eg, silver nitrate).

Furthermore, the present invention includes all of the compounds represented by Formula 1 and pharmaceutically acceptable salts thereof, as well as solvates, optical isomers, and hydrates that can be prepared therefrom.

In addition, in another aspect of the invention,

As shown in Scheme 1 below,

A method for preparing a compound represented by Formula 1, including a step of preparing a compound represented by Formula 1 from a compound represented by Formula 4 is provided.

[Scheme 1]

(In Scheme 1,

R ¹ , R ² , n, X, Y and Z are as defined in Formula 1).

In one embodiment of the invention,

When Z is S, the compound represented by Chemical Formula 1 may be understood to be a compound represented by Chemical Formula 1',

The preparation method represented by the scheme 1, as shown in scheme 2 below,

It may be a method for preparing a compound represented by Formula 1'by reacting a compound represented by Formula 4 with a compound represented by Formula 3.

[Scheme 2]

(In the above reaction formula 2,

R ¹ , R ² , n, X and Y are as defined in Formula 1).

In the step of preparing a compound represented by Formula 1'by reacting the compound represented by Formula 4 and the compound represented by Formula 3,

The above step is not limited thereto, and H ₂ O, ethanol, tetrahydrofuran (THF), dichloromethane, toluene, acetonitrile, dimethylformamide or the like can be used as a usable solvent, and ethanol is preferably used. Can. In addition, the reaction time is not particularly limited, it is preferable to react for 5 to 30 hours, 10 to 20 hours, or about 12 hours. Furthermore, the reaction temperature is not particularly limited, but is preferably performed at room temperature or 20 to 30°C.

In the most preferred aspect, it can be carried out as in the following examples of the present invention, from which methods that can be modified, such as aspects of the experiment, conditions are also included in the scope of the present invention.

In another embodiment of the invention,

When Z is sulfinyl or sulfonyl, the compound represented by Formula 1 may be understood to be a compound represented by Formula 1'' below,

The manufacturing method represented by the scheme 1, as shown in scheme 3 below,

Reacting the compound represented by Formula 4 with the compound represented by Formula 3 to prepare a compound represented by Formula 1'; And

The step of preparing a compound represented by the formula (1) from the compound represented by the formula (1') prepared in the above step; may be a manufacturing method comprising a.

[Scheme 3]

(In the above reaction formula 3,

R ¹ , R ² , n, X and Y are as defined in Formula 1 above; And

W is sulfinyl, or sulfonyl).

In the step of preparing a compound represented by Formula 1'by reacting the compound represented by Formula 4 and the compound represented by Formula 3,

The above step is not limited thereto, and H ₂ O, ethanol, tetrahydrofuran (THF), dichloromethane, toluene, acetonitrile, dimethylformamide or the like can be used as a usable solvent, and ethanol is preferably used. Can. In addition, the reaction time is not particularly limited, it is preferable to react for 5 to 30 hours, 10 to 20 hours, or about 12 hours. Furthermore, the reaction temperature is not particularly limited, but is preferably performed at room temperature or 20 to 30°C.

In the most preferred aspect, it can be carried out as in the following examples of the present invention, from which methods that can be modified, such as aspects of the experiment, conditions are also included in the scope of the present invention.

Furthermore, in the step of preparing a compound represented by Formula 1'' from the compound represented by Formula 1'prepared in the above step,

The above step is not limited thereto, and may be considered as a reaction for introducing a sulfinyl group or a sulfonyl group, and may be reacted using, for example, m-chloroperoxybenzoic acid. Usable solvents include alcohols of C ₁ -C ₄ , acetone, acetonitrile, benzene, 2-butanone, chlorobenzene, chloroform, cyclonucleic acid, toluene, 1,2-dichloromethane (DCM), heptane, hexane, etc. Can be used, preferably chloroform. In addition, the reaction time is not particularly limited, it is preferable to react for 5 to 30 hours, 10 to 20 hours, or about 12 hours. Furthermore, the reaction temperature is not particularly limited, but is preferably performed at room temperature or 20 to 30°C.

In the most preferred aspect, it can be carried out as in the following examples of the present invention, from which methods that can be modified, such as aspects of the experiment, conditions are also included in the scope of the present invention.

Furthermore, in another aspect of the invention,

Provided is a pharmaceutical composition for preventing or treating autophagy-related diseases containing the compound represented by Formula 1 or a pharmaceutically acceptable salt thereof as an active ingredient.

Here, the compound represented by Chemical Formula 1, or a pharmaceutically acceptable salt thereof, may be understood as an active ingredient that modulates autophagy activity.

In one embodiment of the present invention, the compound represented by Chemical Formula 1 of the present invention showed that autophagy activity is regulated, as shown in Experimental Example 1 below.

Specifically, autophagy (autophagy) is a cell exposed to stress, for example, blocking of nutrient supply, exposure to stress such as invasion of bacteria, etc., may be a mediated step. The compound is an anti-inflammatory agent capable of inhibiting the inflammatory response through the inhibition of inflammation-related factors such as NO production inhibition, IL-6, and IL-β, as demonstrated in the following experimental example, by controlling such autophagy activity. It has an inflammatory effect, alternatively it has a melanin-producing inhibitory effect, and alternatively it has an inhibitory effect on metabolic syndrome such as fatty liver suppression and obesity suppression, and is understood as an effective component showing a significant effect on other autophagy-related diseases.

Furthermore, the compound represented by Chemical Formula 1 of the present invention has been found to be a safe compound in general cytotoxicity experiments, cardiac toxicity evaluation, skin toxicity evaluation, eye toxicity evaluation, and liver damage evaluation, and will be provided as an active ingredient that is safer for the living body. It could be suggested.

In another embodiment of the present invention, the autophagy-related diseases include all diseases that have been identified, studied, or reported as being related to, or related to, autophagy activity, to date, for example, cancer. , Atherosclerosis, neurodegenerative disease, Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, Huntington's disease, spinocerebellar ataxia, otolopharyngeal muscular dystrophy, prion disease , Fatal familial insomnia, alpha-1 antitrypsin deficiency, dentatorubral pallidoluysian atrophy, frontal lobe dementia, progressive supranuclear palsy, x-linked spinobulbar muscular atrophy and neuronucleus My intravitreal hyaline inclusion disease.

In another embodiment of the present invention, the autophage-related disease may be a disease associated with metabolic syndrome (differently metabolic disease), for example, obesity, diabetes, dyslipidemia, alcoholic fatty liver, non-alcoholic fatty liver, Hypertension, arteriosclerosis, hyperlipidemia and hyperinsulinemia.

In another embodiment of the present invention, the autophage-related disease may be an inflammatory disease, for example, a disease that can be improved, prevented, or treated from inhibition of NO production, inhibition of IL-6 or IL-1β expression, etc. There are, for example, the inflammatory disease may be intestinal disease, ulcerative colitis, Crohn's disease, arthritis, dermatitis, atopic dermatitis, ophthalmitis, keratitis, hepatitis, non-alcoholic hepatitis.

In another embodiment of the present invention, the autophage-related disease may be a disease related to melanin production, and may be a skin disease resulting from melanogenesis, for example, skin hyperpigmentation disease.

Alternatively, the skin disease caused by melanin production may be understood as a disease including lesions due to pigmentation such as blemishes, freckles, surpluses, and senile pigment spots.

In the above-described autophagy disease, the compound represented by Formula 1 of the present invention, or a pharmaceutically acceptable salt thereof, may exhibit a medically useful effect, such as prevention, improvement, treatment, etc. And as supported by the experimental examples, it is possible to provide a pharmaceutical composition containing it as an active ingredient.

On the other hand, beyond the effects and explanations related to autophages described herein, the present invention is not limited to the autophagy-related diseases, that is, regardless of the above-described autophagy-related theory or mechanism, purely as shown in the following experimental examples of the present invention On the basis of this, a pharmaceutical composition for preventing or treating the following diseases may be provided.

In another aspect of the invention,

Provided is a pharmaceutical composition for preventing or treating an inflammatory disease or metabolic disease, which contains the compound represented by Formula 1 or a pharmaceutically acceptable salt thereof as an active ingredient.

Here, the inflammatory disease may be, for example, a disease that can be improved, prevented, or treated from NO production inhibition, IL-6 or IL-1β expression inhibition, for example, the inflammatory disease is bowel disease, Ulcerative colitis, Crohn's disease, arthritis, dermatitis, atopic dermatitis, ophthalmitis, keratitis, hepatitis, non-alcoholic hepatitis, and the like.

As shown in the following experimental examples and FIGS. 3, 4, and 5 herein, the compound represented by Formula 1 confirmed the NO production inhibition, IL-6 or IL-1β expression inhibitory effect, which is related to the autophagy control effect. It may or may not be described as an independent effect, but is not limited to these descriptions, rather, irrespective of this, from the inflammatory response inhibitory effect demonstrated herein, a pharmaceutical composition for the prevention or treatment of inflammatory diseases will be provided Can.

In addition, the metabolic disease, can be understood as a large number of syndromes, non-limiting examples, obesity, diabetes, dyslipidemia, alcoholic fatty liver, non-alcoholic fatty liver, hypertension, arteriosclerosis, hyperlipidemia, hyperinsulinemia , Etc.

As shown in the following experimental examples and FIGS. 7 and 8 herein, the compound represented by Formula 1 is confirmed to have a fatty liver inhibitory effect, which is related to the autophagy control effect, or may be described as an independent effect. , It is not limited to this description, but rather, a pharmaceutical composition for preventing or treating metabolic disease may be provided based on the fatty liver inhibitory effect, liver damage inhibitory effect, and the like, which are demonstrated herein irrespective of this.

Furthermore, in another aspect of the invention,

Provided is a pharmaceutical composition for preventing or treating skin hyperpigmentation disease, which contains the compound represented by Formula 1 or an acceptable salt thereof as an active ingredient.

Here, the skin hyperpigmentation disease may be, for example, a skin disease caused by melanin hyperplasia, or a disease including lesions due to pigmentation such as blemishes, freckles, surpluses, and senile plaques.

As shown in the following experimental examples and FIG. 9, the compound represented by Chemical Formula 1 of the present invention at all time points before and after the formation of melanocytes is confirmed to excellently inhibit melanin formation, and contains it as an active ingredient, thereby hyperpigmenting the skin. Pharmaceutical compositions for the prevention or treatment of deposition diseases may be provided.

The compound represented by the formula (1) according to the present invention may be administered in various dosage forms, oral and parenteral, during clinical administration, and when formulated, usually used as fillers, extenders, binders, wetting agents, disintegrants, surfactants, etc. It is prepared using diluents or excipients.

Solid preparations for oral administration include tablets, patients, powders, granules, capsules, troches, etc. These solid preparations include at least one excipient such as starch, calcium carbonate, water in one or more compounds of the present invention. It is prepared by mixing sucrose or lactose or gelatin. In addition, lubricants such as magnesium stearate talc are used in addition to simple excipients. Liquid preparations for oral administration include suspending agents, intravenous solutions, emulsions or syrups, etc. In addition to the commonly used simple diluents, water and liquid paraffin, various excipients such as wetting agents, sweeteners, flavoring agents, preservatives, etc. are included. Can be.

Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solvents, suspension solutions, emulsions, lyophilized preparations, suppositories, and the like.

As non-aqueous solvents and suspension solvents, propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate may be used. As a base for suppositories, witepsol, macrogol, tween 61, cacao butter, laurin, glycerol, gelatin, etc. may be used.

In addition to this, transdermal preparations such as creams, ointments, gels, lotions, liquids or patches can be mentioned.

In addition, the effective dosage of the compound of the present invention to the human body may vary depending on the patient's age, weight, sex, dosage form, health status and disease level, and is generally about 0.001-100 mg/kg/day, preferably It is 0.01-35 mg/kg/day. Based on an adult patient weighing 70 kg, it is generally 0.07-7000 mg/day, preferably 0.7-2500 mg/day, and once a day at regular time intervals according to the judgment of a doctor or pharmacist It can also be administered in multiple doses.

In another aspect of the invention,

Provided is a method for treating an autophagy related disease, comprising administering a compound represented by Formula 1 or a pharmaceutically acceptable salt thereof to a subject in a therapeutically effective amount.

At this time, the autophage-related disease is as described herein.

The therapeutically effective amount refers to an amount sufficient to treat, prevent or improve the symptoms or condition of a subject when administered into the body, depending on the administration method. In addition, the amount may vary depending on the weight, age, sex, condition, and family history of the subject to be administered, and in the present invention, the treatment method may determine a different amount of dose according to different conditions for each subject.

The "effective amount" is an autophagy related disease, such as cancer, atherosclerosis, neurodegenerative disease, Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, Huntington's disease, spinocerebellar ataxia ataxia), oculopharyngeal muscular dystrophy, prion disease, fatal familial insomnia, alpha-1 antitrypsin deficiency, biliary tract hypothalamus (dentatorubral pallidoluysian atrophy), frontotemporal dementia, progressive supranuclear palsy ), an effective amount to treat diseases such as x-linked spinobulbar muscular atrophy or neuronal intranuclear hyaline inclusion disease. In other embodiments, “effective amount” of a compound refers to a minimum amount that is capable of modulating autophagy activity.

The compounds and compositions according to the methods of the present invention can be administered using any amount and any route of administration effective for treating a disease. The exact amount required will vary from subject to subject, depending on the subject's species, age, and general condition, severity of infection, specific agent, mode of administration, and the like. The compounds of the present invention are frequently formulated in dosage unit form for ease of administration and uniformity of dosage. The expression “dosage unit form” as used herein means a physically discrete unit of agent suitable for the subject to be treated. It will also be understood that the total daily use of the compounds and compositions of the present invention will be determined by the attending physician within the scope of sound medical judgment. The specific effective dosage level for any particular subject or organism depends on various factors, including: the disease to be treated and the severity of the disease; The activity of the specific compound employed; Specific composition used; Age, weight, general health, subject's gender and diet; The time of administration, route of administration, and rate of excretion of the specific compound employed; Duration of treatment; The specific compound employed alone or co-administered drugs, and other factors well known in medical technology.

The term “subject” may refer to a subject in need of a compound, a pharmaceutically acceptable salt, or pharmaceutical composition thereof provided in the present invention, or alternatively, prevention, amelioration, improvement of an autophagy related disease. , Or can be understood to mean a subject in need of treatment, the subject being, for example, an animal, preferably a non-mammalian, or a mammal, more preferably a livestock, a human, and most preferably a human. it means.

In another aspect, the following health functional food composition, or cosmetic composition may be provided.

First, in one aspect of the present invention,

Provided is a health functional food composition for preventing or improving autophagy-related diseases containing the compound represented by Chemical Formula 1 or a pharmaceutically acceptable salt thereof as an active ingredient.

Herein, the autophagy-related diseases include all diseases that have been identified, studied, or reported as being related to autophagy activity or by regulating autophagy activity to date, for example, cancer, atherosclerosis, nerve Degenerative diseases, Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, Huntington's disease, spinocerebellar ataxia, oculopharyngeal muscular dystrophy, prion disease, fatal familial insomnia, Alpha-1 antitrypsin deficiency, dentatorubral pallidoluysian atrophy, frontal lobe dementia, progressive supranuclear palsy, x-linked spinobulbar muscular atrophy, and intraneuronal vaginal insufficiency (neuronal) intranuclear hyaline inclusion disease).

In addition, in another aspect of the invention,

Provided is a health functional food composition for preventing or improving an inflammatory disease or metabolic disease containing the compound represented by Formula 1 or a pharmaceutically acceptable salt thereof as an active ingredient.

Here, the inflammatory disease may be, for example, a disease that can be improved, prevented, and treated from NO production inhibition, IL-6 or IL-1β expression inhibition, for example, the inflammatory disease is bowel disease, ulcer Sexual colitis, Crohn's disease, arthritis, dermatitis, atopic dermatitis, ophthalmitis, keratitis, hepatitis, non-alcoholic hepatitis, and the like.

In addition, the metabolic disease, can be understood as a large number of syndromes, non-limiting examples, obesity, diabetes, dyslipidemia, alcoholic fatty liver, non-alcoholic fatty liver, hypertension, arteriosclerosis, hyperlipidemia, hyperinsulinemia , Etc.

Furthermore, in another aspect of the invention,

Provided is a cosmetic composition for whitening containing the compound represented by Formula 1 or a salt thereof.

Here, since the cosmetic composition contains the compound represented by the formula (1) as an active ingredient, it is possible to suppress melanin production, and from this, even when aiming at inhibiting melanin formation in the skin, hair, and especially the eye, It can be usefully used as a cosmetic composition for whitening.

Here, the compound represented by Chemical Formula 1, which is a component of the cosmetic composition for whitening, is found in toxicological aspects, as shown in the Experimental Example, and is confirmed to be a safe component for cells, skin, and eyes, forming melanocytes. , There is an advantage that can be applied, without special restrictions, to the area applied to inhibit melanin synthesis.

In one embodiment of the invention,

As shown in the following experimental examples and FIG. 6, the compound represented by Chemical Formula 1 of the present invention can excellently inhibit melanin cell formation, and can inhibit melanin synthesis of already produced melanin cells. Silver may be provided as a cosmetic composition for whitening.

In particular, it may be useful for subjects suffering from skin hyperpigmentation disorders, and may be useful for lesions caused by pigmentation such as spots, freckles, surpluses, and senile plaques.

Alternatively, even if the subject does not suffer from the disease, it can be easily used for purposes such as skin whitening.

On the other hand, the formulation of the cosmetic composition of the present invention is not particularly limited and can be arbitrarily selected and applied. Conventional cosmetic formulations such as external ointment for skin, essence, whitening cream, lotion, emulsion, pack, general lotion, skin milk, cream, serum , Beauty soap, flexible cosmetic, medicinal cosmetic, hair tonic, systemic detergent, oil gel, etc. can be prepared and used in various formulations.

Hereinafter, the present invention will be described in detail by examples and experimental examples.

However, the following examples and experimental examples are merely illustrative of the present invention, and the contents of the present invention are not limited thereto.

<Example 1> Preparation of 1-((4-fluorophenyl)sulfinyl)isoquinoline

Step 1: Preparation of 1-((4-fluorophenyl)thio)isoquinoline

To a solution of 1-chloroisoquinoline (2 g, 12.225 mmol) in ethanol, 4-fluorobenzenethiol (1.436 mL, 13.447 mmol) was added. The reaction was stirred at room temperature for 12 hours. The mixture was concentrated in vacuo. The crude product was purified by silica gel column chromatography using ethyl acetate/hexane to give the target compound (686 mg, 22%).

¹ H NMR (400MHz, DMSO-d6) δ 8.28 (d, J = 8.24 Hz, 1H), 8.18 (d, J = 5.49 Hz, 1H), 7.99 (d, J = 8.24 Hz, 1H), 7.81-7.85 (m, 1H), 7.72-7.77 (m, 1H), 7.61-7.66 (m, 3H), 7.29-7.35 (m, 2H).

Step 2: Preparation of 1-((4-fluorophenyl)sulfinyl)isoquinoline

The 1-((4-fluorophenyl)thio)isoquinoline (850 mg, 3.329 mmol) prepared in Step 1 was dissolved in THF/water. To this mixture, Oxone (574.53 mg, 3.329 mmol) was added and the mixture was stirred at room temperature for 12 hours. The mixture was diluted with water, extracted with ethyl acetate, washed with brine, dried over anhydrous sodium sulfate and concentrated in vacuo. The crude product was purified by silica gel column chromatography using ethyl acetate/hexane to obtain the desired pure compound (218 mg. 24%).

¹ H NMR (400MHz, CDCl ₃ ) δ 8.79 (d, J = 8.54 Hz, 1H), 8.47 (d, J = 5.49 Hz, 1H), 7.52-7.76 (m, 6H), 6.99 (t, J = 8.54 Hz, 2H).

<Example 2> Preparation of 1-((4-fluorophenyl)thio)isoquinoline

Step 1: Preparation of 1-((4-fluorophenyl)thio)isoquinoline

To a solution of 1-chloroisoquinoline (2 g, 12.225 mmol) in ethanol, 4-fluorobenzenethiol (1.436 mL, 13.447 mmol) was added. The reaction was stirred at room temperature for 12 hours. The mixture was concentrated in vacuo. The crude product was purified by silica gel column chromatography using ethyl acetate/hexane to give the target compound (686 mg, 22%).

¹ H NMR (400MHz, DMSO-d6) δ 8.28 (d, J = 8.24 Hz, 1H), 8.18 (d, J = 5.49 Hz, 1H), 7.99 (d, J = 8.24 Hz, 1H), 7.81-7.85 (m, 1H), 7.72-7.77 (m, 1H), 7.61-7.66 (m, 3H), 7.29-7.35 (m, 2H).

<Example 3> Preparation of 1-((4-fluorophenyl)sulfonyl)isoquinoline

Step 1: Preparation of 1-((4-fluorophenyl)thio)isoquinoline

To a solution of 1-chloroisoquinoline (2 g, 12.225 mmol) in ethanol, 4-fluorobenzenethiol (1.436 mL, 13.447 mmol) was added. The reaction was stirred at room temperature for 12 hours. The mixture was concentrated in vacuo. The crude product was purified by silica gel column chromatography using ethyl acetate/hexane to give the target compound (686 mg, 22%).

¹ H NMR (400MHz, DMSO-d6) δ 8.28 (d, J = 8.24 Hz, 1H), 8.18 (d, J = 5.49 Hz, 1H), 7.99 (d, J = 8.24 Hz, 1H), 7.81-7.85 (m, 1H), 7.72-7.77 (m, 1H), 7.61-7.66 (m, 3H), 7.29-7.35 (m, 2H).

Step 2: Preparation of 1-((4-fluorophenyl)sulfonyl)isoquinoline

1-((4-fluorophenyl)thio)isoquinoline (850 mg, 3.329 mmol) was dissolved in THF/water. To this mixture, oxone (2.2 eq, 7.3238 mmol) was added and the mixture was stirred at room temperature for 12 hours. The mixture was diluted with water, extracted with ethyl acetate, washed with brine, dried over anhydrous sodium sulfate and concentrated in vacuo. The crude product was purified by silica gel column chromatography using ethyl acetate/hexane to give the pure target compound (4c, 66%).

¹ HNMR (400 MHz, CDCl ₃ ) δ 8.79 (d, J = 8.54 Hz, 1H), 8.47 (d, J = 5.49 Hz, 1H), 7.52-7.76 (m, 6H), 6.99 (t, J = 8.54 Hz, 2H).

<Example 4> Preparation of 4-((4-fluorophenyl)sulfinyl)quinoline

Step 1: Preparation of 4-((4-fluorophenyl)thio)quinolone

It was carried out as in step 1 of Example 1, 4-chloroquinoline (250 mg, 1.528 mmol) was used instead of 1-chloroisoquinoline.

To a solution of 4-chloroquinoline (250 mg, 1.528 mmol) in ethanol, 4-fluorobenzenethiol (250 mg, 1.528 mmol) was added. The reaction was stirred at room temperature for 12 hours. The mixture was concentrated in vacuo. The crude product was purified by silica gel column chromatography using ethyl acetate/hexane to obtain the target compound (185.0 mg, 47.4%).

¹ HNMR (400 MHz, CDCl ₃ ) δ 8.565 (d, J = 4.88 Hz, 1H), 8.178 (d, J = 9.16 Hz, 1H), 8.069 (d, J = 8.24 Hz, 1H), 7.730 (m, 1H), 7.583 (m, 3H), 7.181 (t, J = 17.40, 2H), 6.673 (d, J = 4.58, 1H).

Step 2: Preparation of 4-((4-fluorophenyl)sulfinyl)quinoline

Performed as in Step 2 of Example 1, but instead of 1-((4-fluorophenyl)thio)isoquinoline, 4-((4-fluorophenyl)thio)quinolone prepared in Step 1 (280mg) , 1.097 mmol) to give the target compound (47%).

¹ HNMR (400 MHz, CDCl ₃ ) δ 9.124 (d, J = 4.58 Hz, 1H), 8.156 (m, 2H), 7.974 (d, J = 8.85 Hz, 1H), 7.711 (m, 3H), 7.572 ( m, 1H), 7.096 (t, J=17.09 Hz, 2H).

<Example 5> Preparation of 1-((2,4-difluorophenyl)sulfinyl)isoquinoline

Step 1: Preparation of 1-((2,4-difluorophenyl)thio)isoquinoline

It was carried out as in step 1 of Example 1, but instead of 4-fluorobenzenethiol, 2,4-difluorobenzenethiol (1.527 mmol) was used.

To a solution of 1-chloroquinoline (0.25 g, 1.527 mmol) in ethanol, 2,4-difluorobenzenethiol (1.527 mmol) was added. The reaction was stirred at room temperature for 12 hours. The mixture was concentrated in vacuo. The crude product was purified by silica gel column chromatography using ethyl acetate/hexane to give the target compound (42%).

¹ HNMR (400 MHz, CDCl ₃ ) δ 8.308 (d, J = 8.24 Hz, 1H), 8.159 (d, J = 5.95 Hz, 1H), 7.778 (d, J = 8.24 Hz, 1H), 7.690 (t, J = 14.95 Hz, 1H), 7.602 (m, 2H), 7.367 (d, J = 5.49 Hz, 1H), 6.961 (m, 2H).

Step 2: Preparation of 1-((2,4-difluorophenyl)sulfinyl)isoquinoline

Except that the 1-((2,4-difluorophenyl)thio)isoquinoline (200 mg) prepared in Step 1 was used, the same procedure as in Step 2 of Example 1 was carried out to obtain the final target compound. Obtained (57%).

¹ HNMR (400 MHz, CDCl ₃ ) δ 8.685 (d, J = 8.24 Hz, 1H), 8.579 (d, J = 5.49 Hz, 1H), 8.085 (m, 1H), 7.898 (d, J = 8.24 Hz, 1H), 7.742 (m, 3H), 7.074 (m, 1H), 6.747 (m, 1H).

<Example 6> Preparation of 1-((4-chlorophenyl)sulfinyl)isoquinoline

Step 1: Preparation of 1-((4-chlorophenyl)thio)isoquinoline

Performed as in Step 1 of Example 1 above, 4-chlorobenzenethiol (1.527 mmol) was used instead of 4-fluorobenzenethiol to obtain the target compound (48%).

¹ HNMR (400 MHz, CDCl ₃ ) δ 8.313 (d, J = 8.24, 1H), 8.220 (d, J = 5.80 Hz, 1H), 7.782 (d, J = 8.24 Hz, 1H), 7.690 (m, 1H ), 7.607 (m, 1H), 7.510 (m, 2H), 7.379 (m, 3H).

Step 2: Preparation of 1-((4-chlorophenyl)sulfinyl)isoquinoline

Except for using the 1-((4-chlorophenyl)thio)isoquinoline (200 mg) prepared in Step 1, it was carried out as in Step 2 of Example 1 to obtain the final target compound (25 %).

¹ HNMR (400 MHz, CDCl ₃ ) δ 8.903 (d, J = 8.54 Hz, 1H), 8.558 (d, J = 5.49 Hz, 1H), 7.871 (d, J = 8.24 Hz, 1H), 7.750 (m, 3H), 7.706 (d, J = 5.49 Hz, 1H), 7.648 (m, 1H), 7.374 (d, J = 8.54 Hz, 2H).

<Example 7> Preparation of 1-((4-(trifluoromethyl)phenyl)sulfinyl)isoquinoline

Step 1: Preparation of 1-((4-(trifluoromethyl)phenyl)thio)isoquinoline

Performed as in Step 1 of Example 1, 4-(trifluoromethyl)benzenethiol (1.527 mmol) was used instead of 4-fluorobenzenethiol to obtain the target compound (59%).

¹ HNMR (400 MHz, CDCl ₃ ) δ 8.335 (d, J = 8.24 Hz, 1H), 8.269 (d, J = 5.49 Hz, 1H), 7.811 (d, J = 7.93 Hz, 1H), 7.710 (m, 1H), 7.630 (m, 5H), 7.465 (d, J = 5.49 Hz, 1H)

Step 2: Preparation of 1-((4-(trifluoromethyl)phenyl)sulfinyl)isoquinoline

Except that the 1-((4-(trifluoromethyl)phenyl)thio)isoquinoline (250 mg) prepared in Step 1 was used, and was carried out as in Step 2 of Example 1, the final target compound Was obtained (62%).

¹ HNMR (400 MHz, CDCl ₃ ) δ 8.972 (d, J = 8.54 Hz, 1H), 8.558 (d, J = 5.49 Hz, 1H), 7.946 (d, J = 8.24 Hz, 2H), 7.880 (d, J = 8.24 Hz, 1H), 7.714 (m, 5H).

The chemical structural formula of the compound prepared in Example 1-7 is shown in Table 1 below.

Example Chemical structural formula Example Chemical structural formula One

5

2

6

3

7

4

<Experiment 1> Evaluation of autophagy control activity

In order to evaluate the autophagy control activity of the novel isoquinoline derivatives according to the present invention, experiments were conducted as follows.

<1-1> Autophagy index analysis

The conversion of the autophagy indicator LC3-I to LC3-II was evaluated by Western blot analysis.

Specifically, Western blots were divided into 20 × 10 ⁵ mouse macrophage RAW264.7 cells in a 60 mm cell culture plate and cultured for one day. After treating 1, 5, 10, and 20 μM of the compound of Example 1 of the present invention using a DMSO solution, it was cultured in a 37° C. CO ₂ incubator for one day. Cells were separated using 0.1% Trypsin-EDTA, and then centrifuged to collect cells. Cells were lysis buffer (RIPA lysis buffer) (50 mM Tris, pH 7.4, 0.1% SDS, 1% NP-40, 150 mM NaCl, 1 mM PMSF, 10 mM NaF, 10 ug/ml aprotinin, 10 ug/ml leupeptin, After lysis using 10 mM Na ₃ VO ₄ ), centrifugation was performed to obtain cell lysate. The obtained cell lysate was quantified by protein, and the cell lysate containing the same amount (50 ug) of protein was separated by electrophoresis on a 4-12% Bis-Tris acrylamide gel. The isolated proteins were transferred to the PVDF membrane, followed by western blotting. Blocking of the PVDF membrane was carried out for 1 hour in TBST (25 mM Tris, pH 7.5, 150 mM NaCl, 0.1% Tween-200) containing 5% skim milk. As a primary antibody, an anti-LC3 antibody was used. Anti-GAPDH (anti-GAPDH) purchased from Santacruz was used as an internal standard protein. The primary antibody was reacted at room temperature for 3 hours. Then, washed three times with TBST buffer solution, and reacted with a secondary antibody (horseradish peroxidase-conjugated anti-goat IgG) for 2 hours at room temperature. After the reaction was completed, the PVDF membrane was washed 3 times with TBST buffer and reacted with the ECL solution to obtain chemiluminescence using an imaging device, and the results confirmed by Western blotting are shown in FIG. 1.

1 is a Western blot graph confirming the conversion of LC3-I and LC3-II according to Example 1 Compounds 1, 5, 10, and 20 μM treatment according to the present invention to Raw 264.7 mouse macrophages.

<1-2> Intracellular autopagosome visualization analysis

In order to detect autophagosomes by autophagy control, observation analysis was performed using acridine orange, which selectively stains an acidic antifoaming agent in the cytoplasm.

Specifically, autophagosomes undergo a change in fluorescence properties by protonation of acridine orange accumulated due to acidification in the cytoplasm, and experiments as follows using these fluorescence properties.

Frequency divider such that the 5 × 10 ⁴ number / well for Raw 264.7 mouse macrophages in 96-well plates and allowed to stabilize at 37 ℃, 5% CO ₂ incubator for 24 hours. Raw 264.7 cells were treated with Example 1 compounds at 5, 10, 20, and 50 μM concentrations, and after 1 hour, LPS 1 ug/ml was treated, and then cultured for 24 hours. The medium was removed, replaced with a medium containing 5 μM acridine orange, and stained for 15 minutes in a 37° C., 5% CO ₂ incubator. After staining was completed, washing was performed with PBS buffer, and intracellular autopagosome formation was observed using a fluorescence microscope. The results visualized by acridine orange fluorescence are shown in FIG. 2.

Figure 2 after adjusting the inflammatory response to Raw 264.7 mouse macrophages (LPS treatment), 5, 10, 20, 50 μM treatment of Example Compound 1 of the present invention, fluorescence micrograph confirming the change in the observed auto-pagosome to be.

<Experiment 2> Evaluation of disease-related efficacy

<2-1> Measurement of anti-inflammatory (NO) activity

Macrophages are cells that induce an inflammatory reaction, and when stress or stimulation occurs, they produce NO and cytokines and play a biodefensive role. NO plays an essential role in innate immune response in response to pathogens such as viruses and bacteria, and L-arginine is produced by NO synthase (NOS), and NOS is divided into cNOS (constitutive NOS) and iNOS (inducible NOS). Of these, iNOS produces a large amount of NO when stimulating stress or cytokines, causing an inflammatory reaction.

In order to confirm the effect on the NO production of the compound of Example 1 of the present invention in Raw 264.7, a macrophage in which the inflammatory response by LPS is regulated, the experiment was conducted as follows.

Macrophage Raw 264.7 cells were prepared in 96-well plates at 1×10 ⁵ cells/well and stabilized in a 37° C., 5% CO ₂ incubator for 24 hours. Raw 264.7 cells were treated with the compound of Example 1 by concentration, and after 1 hour, LPS 1 μg/ml was treated and cultured for 24 hours. To measure NO production, prepare 40 mg/ml grease reagent, mix with the culture supernatant in the same ratio, react for 15 minutes at room temperature, measure absorbance at 540 nm, and convert it to LPS based on the results obtained therefrom. The inhibitory effect of NO production was expressed as a percentage (Fig. 3(a)).

On the other hand, according to the treatment according to the concentration of Example 1 of the present invention, cell viability was measured and expressed as a percentage compared to a control (FIG. 3(b)).

Figure 3 (a): After the production of NO by LPS (control of inflammatory reaction), Example 1 compounds of the present invention were treated with 0.1, 0.5, 1, 10, and 100 μM, respectively, and absorbance was measured using a grease reagent. Graph showing the measured NO production rate as a percentage in terms of LPS, and (b): control the cell viability according to 0.1, 0.5, 1, 10, and 100 μM treatment of LPS and Example 1 compounds, respectively, respectively. Control).

Referring to (a) of FIG. 3, it can be seen that NO production is inhibited from the treatment of the compound of Example 1 of the present invention, and it is confirmed that the NO production inhibitory effect is also large as the treatment concentration is increased. On the other hand, looking at (b) of Figure 3, the present invention Example 1 compound was not found significant cytotoxicity, in the case of 10 μM treatment, 60% NO production inhibitory effect is confirmed, cytotoxicity is 12% As a result, it was found that the inhibitory effect of NO production was not caused by cytotoxicity.

<2-2> Measurement of cytokine gene expression related to inflammatory response

LPS is an endotoxin in the extracellular membrane of Gram-negative bacteria that stimulates macrophages to stimulate the secretion of inflammatory mediator cytokines, and these pro-inflammatory factors activate immune cells to help effectively defend against bacterial invasion. In addition, since cytokines released from LPS-stimulated macrophages can degrade normal cell function, it is very important to find a drug that can suppress the immune function of LPS-stimulated macrophages.

Raw 264.7 cells were prepared in 96-well plates at 1×10 ⁵ cells/well and stabilized in a 37° C., 5% CO ₂ incubator for 24 hours. Raw 264.7 cells were treated with Example 1 Compound-1 at 1 μM or 10 μM, and after 1 hour treatment with LPS 1 μg/ml was incubated for 24 hours. Cells from which the culture was removed were washed with PBS buffer, separated using 0.1% Trypsin-EDTA and centrifuged to collect. RNA was isolated using a trizol solution (Trizole reagent) for cell precipitation. The amount of isolated RNA was quantified by measuring absorbance at 260 nm. For real-time polymerase chain reaction experiment, after mixing about 100 ng of RNA with Verso SYBR Green 1-Step qRT-PCR Low ROX Mix according to the manufacturer's manual, Real-time PCR machine (ABI 7500 fast-real time PCR, Applied biosystem , USA).

Genes used for real-time polymerase chain reaction were ordered from Bioneer. The polymerase reaction was performed at 50°C for 15 minutes, and then at 95°C for 15 minutes, and then a cycle of denaturation temperature of 95°C for 15 seconds, annealing temperature of 60°C for 15 seconds, and an enzyme reaction of 72°C was repeated 40 times. The results are shown in FIG. 4.

Figure 4 is a graph showing the measurement of the gene expression level of the cytokine IL-6 related to the inflammatory response by the compound treatment of Example 1 according to the present invention.

<2-3> Measurement of protein expression related to inflammatory response

In order to measure the anti-inflammatory effect of the novel isoquinoline derivatives of the present invention, experiments were conducted as follows.

From the control of the inflammatory response by LPS treatment in macrophage Raw 264.7 cells, inflammatory proteins such as interleukin-6 (IL-6) and interleukin-1 beta (IL-1β) are secreted outside the cell. This experiment is a method of evaluating anti-inflammatory activity by measuring the amount of inflammatory proteins secreted after controlling the inflammatory response by LPS using an enzyme-linked immunosorbent assay (ELISA).

Specifically, Raw 264.7 cells were prepared in a 96-well plate at 1×10 ⁵ cells/well and stabilized in a 37° C. incubator maintaining 5% CO ₂ for 24 hours. Raw 264.7 cells were treated with 0.1, 1, and 10 μM of the compound of the present invention Example 1, and cultured for 24 hours by treating LPS 1 μg/ml after 1 hour. After taking only the cell culture solution, the supernatant was collected by centrifugation at 13,000 rpm for 5 minutes. For the determination of secreted interleukin, a commercially available kit from Elabscience was used, and the test procedure was performed according to the manufacturer's manual. 100 μl of the culture solution obtained from the cultured cells was dispensed into the provided wells, and then cultured at 37°C for 90 minutes. The culture solution was removed and 100 μl of biotin-labeled selective antibody was reacted at 37° C. for 1 hour, and then washed 3 times with a washing solution. After the HRP-labeled 100 μl solution was reacted at 37° C. for 30 minutes, it was washed 5 times with a washing solution. After adding 90 μl of the solution containing the specific substrate of HRP, incubated for 15 minutes at 37° C., and after adding 50 μl of the termination solution, absorbance was measured at 450 nm, and the experimental results are shown in FIG. 5. .

Figure 5 is a graph showing the protein measurement of the protein IL-6 and IL-1β related to the inflammatory response by Example 1 of the present invention compound 0.1, 1, and 10 μM.

<2-4> Whitening efficacy evaluation

To evaluate the whitening efficacy of the novel isoquinoline derivatives of the present invention, the following experiment was performed.

The black melanin cells of zebrafish did not migrate to keratinocytes, but had the characteristic of being limited to melanin cells. This can be investigated by limiting the formation of melanin pigments when observing the regeneration of melanin pigments, then confining to the melanin cell production pathway, and it is easy to observe the melanin synthesized in the early stages of development. In comparison, it is more useful for melanin pigment change studies (Logan DW, Burn SF, Jackson IJ. (2006) Regulation of pigmentation in zebrafish melanophores.Pigment Cell Res. 2006 Jun; 19(3):206-13.).

Specifically, 10 individual zebrafish fertilized eggs at a time of 10 hours after fertilization are added to each 24-well plate, and 1 ml of breeding egg water is added. Dilute 0.1 μM and 1 μM of the Example 1 compound in each well and add the final volume of each well to 2 ml. Subsequently, each 24-well plate is wrapped with foil to block light, and then incubated for 22 hours in a 28°C incubator. At 32 hours after fertilization, in order to photograph the formation of black melanocytes, the membranes of the embryos were removed using Forcep (Fine Science Tools), and the embryos were developed with Tricaine (4 g/L). After anesthesia, the embryo was transferred onto 3% methyl cellulose and photographed. Genetic embryo imaging equipment was used LEICA MZ10F fluorescence microscope, LEICA DFC425 camera, Leica Application Suite software (v4.5). In all experiments, the solution used for culturing the embryos was dissolved in 60 mg/L sea salt (Sigma-Aldrich, S9883) in tertiary distilled water, and sterilized egg water was used. As a control group, phenylthiourea (200 μM PTU) known as a solvent control group (0.4% DMSO) and a tyrosinase inhibitor was used (FIG. 6(a)).

On the other hand, in order to confirm the ability to inhibit melanin synthesis of already produced melanocytes, an experiment was conducted at a 60 hour time point after fertilization. Specifically, 10 fertilizers are placed in 24-well plates for 60 hours after fertilization, and 1 ml of egg water, a breeding water, is added. Subsequently, 10 μM and 20 μM of Example 1 compound are diluted and added to each well in a final volume of 2 ml. The 24-well plate containing the compound is wrapped with foil to block light, and then incubated for 12 hours in a 28°C incubator. To photograph the formation of black melanocytes, the embryos were anesthetized with Tricaine (4 g/L), and then the embryos were transferred onto 3% methyl cellulose and photographed. The equipment described above was used for imaging (Fig. 6(b)).

Figure 6 (a): a photograph of a zebrafish that confirmed the inhibitory effect of melanocyte formation at 32 hours after fertilization after 22 hours of treatment with the compound of Example 1 of the present invention, targeting the zebrafish fertilized egg at 10 hours after fertilization. , And (b) is a photograph of a zebrafish, which was confirmed for the melanin synthesis inhibitory effect at 72 hours after fertilization after 12 hours of treatment with the compound of Example 1 of the present invention, targeting the zebrafish pom at 60 hours after fertilization. .

6, excellent melanin synthesis inhibitory effect of the novel isoquinoline derivative of the present invention is confirmed. Particularly, at a time point of 32 hours after fertilization, it was confirmed that the synthesis of melanin was inhibited at a level similar to that of the PTU treated group in the Example 1 compound treatment group of the present invention, compared to the PTU 0.2 mM treated group. In addition, at 72 hours after fertilization, compared to the 0.2 mM PTU treated group, the Example 1 compound treated group of the present invention was found to have superior melanin synthesis inhibitory effect compared to PTU, especially at 20 μM treatment.

<2-5> Fat liver inhibition efficacy evaluation

In order to evaluate the effect of inhibiting fatty liver of the novel isoquinoline derivatives of the present invention, the following experiment was performed using LipidGreen2 dye which can dye lipid droplets.

In the case of normal liver, the proportion of fat is about 5%, and the state in which more fat is accumulated is called fat liver. It can be largely divided into alcoholic fatty liver due to excessive drinking and non-alcoholic fatty liver due to obesity, diabetes, hyperlipidemia, and drugs. The main causes are drinking and obesity, and diseases such as hyperlipidemia and diabetes, which have high levels of fat in the blood, may be accompanied, and drugs such as adrenal cortical hormones (steroids) or female hormones may also be the cause.

Tamoxifen is a nonsteroidal anti-estrogen agent that competes and binds estrogen receptor α/β with estrogen, developed by the British ICI Company, and has endocrine therapy for female hormone-dependent cancer since the 1980s. Zero was used most. In particular, it was known that necrosis of hepatocytes occurred in the zebrafish pomace treated with tamoxifen, and the findings of fatosis in adult liver were confirmed (Nam et al., (2016) Expression of miRNA-122 induced by liver toxicants in zebrafish.BioMed Research International. 2016: 1-7.).

Specifically, the zebrafish used in this experimental example is a cheer at 5 days after fertilization. 10 healthy cheers are placed in 24-well plates and 1 ml of egg water is added. DMSO was diluted to 0.4% with a negative control of non-fatty liver and exposed to the pom. To control alcoholic fatty liver, tamoxifen (Sigma-Aldrich, T5648) was diluted with 5 μM and exposed to the pom. In order to examine the effect of inhibiting fatty liver of the Example 1 compound of the present invention, 0.5 μM, 1 μM, and 5 μM of the Example 1 compound was mixed with 5 μM tamoxifen and exposed to the pom (the final volume of each well was 2 ml). The 24-well plate containing the compound is wrapped with foil to block light, and then incubated for 24 hours in a 28°C incubator. To remove the compound in each well, wash three times with egg water for 5 minutes. For fat-specific staining, LipidGreen2 was diluted with 5 μM and exposed to the pom for 30 minutes, and then washed three times with egg water for 5 minutes. To evaluate the fatty liver of zebrafish pom, anesthetized embryos were taken with Tricaine (4 g/L), and then the embryos were transferred onto 3% methyl cellulose and photographed. Genetic embryo imaging equipment was used LEICA MZ10F fluorescence microscope, LEICA DFC425 camera, Leica Application Suite software (v4.5). The fluorescence wavelength used was a GFP Plants filter set (Excitation 470/40 nm, Emmision 525/50 nm). In order to quantify the liver area and fat staining degree of the pom on the image, the liver portion of each image was quantified using the ImageJ 1.50i software (National Institutes of Health, USA), and the fluorescence intensity was converted to a heat map image. The experimental results are shown in FIGS. 7 and 8.

FIG. 7 is a view of a normal microscope image, a fluorescence microscope image, and a heat map conversion image observed after treating DMSO, tamoxifen, and Example 1 of the present invention for a zebrafish cheer 5 days after fertilization.

FIG. 8 shows the liver area (a) and fluorescence intensity (b) on the observed zebrafish cheer image after treating DMSO, tamoxifen, and Example 1 of the present invention for the zebrafish cheer 5 days after fertilization, respectively. It is a graph showing the results quantified using the software (National Institutes of Health, USA) (p value is the t-test result for each experimental group compared to the tamoxifen treated group. *p ≤ 0.01, **p ≤ 0.001).

In FIG. 7, the white dotted line on the general microscope image and the fluorescence microscope image and the black dotted line on the heat map conversion image indicate liver areas of the same cheer. In particular, the stronger the fluorescence intensity is, the more red color is displayed on the heat map conversion image. The blue color indicates that the fluorescence intensity is weaker.

In FIG. 7, LipidGreen2 fat staining from the Example 1 compound treatment group of the present invention confirms the effect that the fatty liver regulated by tamoxifen is restored to a normal level.

In addition, in FIG. 8, it can be confirmed that the liver area of the normal cheer level was maintained from the Example 1 compound treatment group of the present invention, and it was confirmed that the intensity of fluorescence was reduced to a normal level compared to the tamoxifen-controlled fatty liver cheer as a result of fat staining. have.

<Experiment 3> Toxicity evaluation

<3-1> Non-selective general cytotoxicity evaluation

In order to evaluate non-selective cytotoxicity in various cell lines by the compound of Example 1 of the present invention, the following experiment was performed.

These experiments are cell lines originating from different species: African green monkey kidney cell line (VERO), human embryonic lung cell line (HFL-1), mouse fibroblast cell line (L929), mouse embryonic fibroblast cell line (NIH 3T3), CHO-K1 (Chinese hamster ovary cell line) and HMV-II (human malignant melanoma cell line) were used.

Specifically, VERO, HFL-1, L929, NIH 3T3 cell line was DMEM medium, CHO-K1 was RPMI1640 medium, HMV-II cells were DME/F-12 medium, and 10% FBS (Fetal Bovine Serum) was added to each medium. And 100 μg/mL of antibiotic was added. Cells were cultured by adaptation in a 37°C, 5% CO ₂ humidified incubator, and washed with PBS (Phosphate buffer solution) at about 70-80% growth point of the culture dish every 2-3 days. The cells were passaged by treatment with Trypsin-EDTA (Gibco, USA).

Cells used in this example were dispensed into 96-wells so that the number was 15,000-25,000, and then cultured by adapting in a humidified incubator at 37°C and 5% CO ₂ for 24 hours. The Example 1 compound of the present invention was prepared to be 10 mM using DMSO, diluted sequentially, treated to each cell to a final concentration, and cultured for 24 hours. Cell viability was measured according to the standard method of Cyto XTM cell viability assay kit (LPS solution). After adding 10 μl of the coloring reagent to each well and incubating for 2 hours in a 37°C, 5% CO ₂ wet incubator, absorbance was measured at 450 nm. Survival was calculated by calculating the ratio of absorbance changed compared to absorbance measured in the DMSO-only treatment group. The results are shown in FIG. 9.

9 is a graph showing the cell viability according to the concentration-dependent treatment of the compound of Example 1 of the present invention for VERO, HFL-1, L929, NIH 3T3, CHO-K1, and HMV-II cell lines.

Referring to FIG. 9, it was found that the compound of Example 1 of the present invention had no significant toxicity in all cell lines up to 10 μM.

<3-2> Evaluation of cardiac toxicity

In order to evaluate the possibility of inducing cardiotoxicity when the compound of Example 1 of the present invention is introduced into the body, the possibility of inhibition against the H ⁺ (HERG, human ether-a-go-go related gene) K ⁺ channel protein known as the main cause of arrhythmia Experimented.

First, a ligand binding force test was performed to evaluate the binding force of the compound of Example 1 that binds to a hug channel protein under in vitro conditions. After dispensing 10 μl of the biofilm containing the Hug channel protein into the 384-black well plate at room temperature, prepare DMSO, the positive control E-4031, and the compound of Example 1 to be 4 times the final test concentration, and then 5 μl each Was added. After dispensing 5 μl of the tracer material in which the fluorescent substance was attached to the astigizole (astemizole), which is known as the selective binding substance of the hug channel protein, to the mixed solution of the two solutions, the cells were incubated for 2 hours at room temperature with light blocked. In order to excite the tracer material contained in the reaction solution, light of 530 nm was irradiated, and then light of 585 nm coming back was measured by polarization. In this experiment, a high polarization value by the tracer material means a lot of binding between the hug protein and the tracer, whereas a low polarization value means a lot of binding between the hug protein and the test material. The binding capacity of the hug protein by the test substance was expressed as the inhibition rate (%) compared to the DMSO treatment group.

Next, in order to evaluate the activity of the hug channel protein using an electrophysiological method, a patch clamp test was performed. To measure the activity of the Hug channel protein, a HEK-293 cell line overexpressing the Hug channel protein was used. Cells were cultured by adaptation in a 37°C, 5% CO ₂ humidified incubator, and after 2-3 days of growth, about 70-80% of the culture dish was washed with PBS (Phosphate buffer solution), Trypsin-EDTA treatment was followed by passage culture. For the test, 2 x 10 ⁵ cells were dispensed into a 60 mm cell culture dish, and then cultured in a 5% CO ₂ wetted incubator for 48 hours. The cultured cells were separated by treatment with Trypsin-EDTA (Gibco, USA), and then applied to an automated patch clamp device (PatchXpress, Molecular device) to measure the flow of K ⁺ by the Hug channel protein.

The experimental results by the above two experimental methods are shown in FIG. 10.

10 is a graph showing the binding and inhibitory activity of the hug channel protein by the compound of Example 1 of the present invention by ligand binding and patch clamp curves, respectively.

As can be seen in Figure 10, the Example 1 compound of the present invention was found to have about 10% binding and inhibitory activity in the Hug channel protein at 10 μM. These results are significantly below the 50% inhibition rate at 10 μM, which is a standard for judging toxicity, and it is determined that the Example 1 compound of the present invention has a low possibility of inducing cardiac toxicity by inhibition of the Hug channel protein in vivo.

<3-3> Skin toxicity evaluation

In order to evaluate the possibility of skin toxicity that may be caused upon skin contact of the Example 1 compound of the present invention, the following experiment was performed using artificial skin made of a structure similar to that of a living body.

Specifically, the experiment was conducted by OECD Guideline for the Testing of Chemicals No. 439, OECD (2010) and the manufacturer (Tego Science) was performed in accordance with the recommended method.

First, the artificial skin cultured in agarose medium was transferred to a 12-well cell culture plate, and a culture medium was added, followed by incubation for 1 day in a 37°C incubator wetted with 5% CO ₂ . After removing the medium of the artificial skin culture surface, 90 μl of the diluted Example 1 compound of the present invention was evenly applied to the skin surface using PBS buffer, and cultured at room temperature for 15 minutes. The cell surface was washed twice using 10 ml PBS buffer, and a new culture medium was added, followed by incubation for 42 hours in a 5% CO ₂ wet 37° C. incubator. In order to measure the survival rate of the skin, the artificial skin was transferred to a 12-well plate containing 2 ml MTT solution (0.3 mg/ml) and incubated for 3 hours. The stained artificial skin tissue was removed and transferred to a tube containing 500 μl of 0.04N-isopropanol, followed by shaking for 4 hours to discolor. The solution was centrifuged at 13,000 rpm for 5 minutes, and 100 μl of the supernatant was transferred to a 96-well plate, and absorbance was measured at 570 nm. The results of the experiment are shown in FIG. 11.

11 shows the survival rate of vehicle, ARB, SDS, Example 1 compounds 10, 20, and 50 μM treatment group based on the measured absorbance after eluting MTT stained in artificial skin tissue with isopropanol. It is a graph.

In FIG. 11, a 5% SDS treatment as a positive control group showed a significant decrease in MTT development due to necrosis of skin tissue, whereas the Example 1 compound of the present invention did not have a significant decrease in survival rate up to 20 μM, only a maximum concentration of 50 A slight decrease in survival was observed only in μM.

<3-4> Eye toxicity evaluation

In order to evaluate the possibility of ocular toxicity that may be caused when the compound is exposed to the eye of Example 1 of the present invention, the cultured artificial cornea was used as follows.

The artificial corneal tissue cultured in agarose medium was transferred to a 12-well cell culture plate, and a culture medium was added, followed by incubation for 1 day in a 37°C incubator wetted with 5% CO ₂ . After removing the culture medium of the artificial cornea, 30 μl of the present invention Example 1 compound diluted with PBS buffer was evenly applied to the surface of the artificial cornea and incubated at room temperature for 30 minutes. The cell surface was washed twice using 10 ml PBS buffer, and a new culture medium was added, followed by incubation for 30 minutes in a 5% CO ₂ wet 37°C incubator. To measure the survival rate of the cornea, the artificial cornea was transferred to a 12-well plate containing 2 ml MTT solution (1 mg/ml), and then cultured for 3 hours. The dyed artificial corneal tissue was removed and transferred to a tube containing 1.5 ml of isopropanol, followed by shaking culture to discolor. The solution was centrifuged at 13,000 rpm for 5 minutes, and 100 μl of the supernatant was transferred to a 96-well plate, and absorbance was measured at 570 nm. The results of the experiment are shown in FIG. 12.

FIG. 12 shows the survival rate of vehicle, ARB, acetic acid, Example 1 compound 10, 20, and 50 μM treated group based on the measured absorbance after eluting MTT stained in artificial corneal tissue with isopropanol. It is a graph.

In FIG. 12, in the treatment of 10% acetic acid, which is a positive control group, a significant decrease in MTT development was observed due to necrosis of corneal tissue, whereas a decrease in survival was not observed until the maximum concentration of 50 μM in the Example 1 compound of the present invention.

<3-5> ALT (Aminotransferase) activity evaluation of liver damage index

In order to evaluate the liver damage index activity of the compound of Example 1 of the present invention, the liver damage index ALT (aminotransferase) activity was analyzed in ob/ob mice, an obese-controlled rodent model.

ALT is an enzyme that helps exchange amino groups between keto acid and amino acids. When a cell is damaged, an enzyme leaked into the blood is measured, and when the liver is damaged, such as viral hepatitis, it is elevated.

Specifically, the experimental animals used ob/ob mice lacking leptin in C57B/6 mice, and were administered 10 mpk or 100 mpk of Example 1 compound of the present invention for 3 weeks. After dosing was completed, CO ₂ gas was stunned and the liver was collected. The liver tissue was washed with a saline solution, and then 50 mg was removed and homogenized. Only the lysate was collected by centrifugation. The substrate for the enzyme reaction with the lysate was added and the fluorescence value was measured at 535/587 nm, and the results are shown in the graph of FIG. 13.

13 is a graph showing the measurement of ALT (aminotransferase) activity as an indicator of liver damage after administration of a vehicle, a positive control (10 mpk Rosi), and a compound of Example 1 (10 mpk and 100 mpk) of the present invention, respectively, in an obese-controlled rodent model. to be.

<3-6> Oxidative stress evaluation

In order to evaluate the oxidative stress of the compound of Example 1 of the present invention, the activity of thiobarbituric acid reactive substance (TBARS), an index of oxidative stress, was analyzed in ob/ob mice, an obese-controlled rodent model.

Lipid peroxidation is a well-known mechanism of cell damage and is used as an indicator of oxidative stress. Measurement of TBARS is a lipid peroxidation confirmation method, and MDA and thiobarburic acid (TBA) are combined to form an MDA-TBA product. By measuring this, the degree of lipid peroxidation can be determined.

Specifically, the experimental animals used ob/ob mice lacking leptin in C57B/6 mice, and were administered 10 mpk or 100 mpk of Example 1 compound of the present invention for 3 weeks. After completion of the administration, they were stunned with CO ₂ gas and liver was collected. The liver tissue was washed with a saline solution and then homogenized by removing 50 mg. Only the lysate was collected by centrifugation. The MDA-TBA product formed as a result of the reaction of MDA and TBA at an acidic condition of 95° C. was measured at an OD of 532 nm, and the results are shown in FIG.

Figure 14 in the obesity-controlled rodent model, respectively, after administration of the vehicle, the positive control (10 mpk Rosi), the present invention Example 1 compound (10 mpk and 100 mpk), the activity of the oxidative stress indicator TBARS (Thiobarbituric acid reactive substance) It is a graph that is measured and shown.

Claims (11)

A compound represented by Formula 1, or a pharmaceutically acceptable salt thereof:
[Formula 1]

(In the above formula 1,
R ¹ is substituted or unsubstituted phenyl,
The substituted phenyl is substituted with one or more substituents selected from the group consisting of hydroxy, cyano, nitro, amino, halogen and substituted or unsubstituted C _1-5 alkyl of straight or branched chain,
Here, the substituted alkyl is substituted with halogen;
R ² is hydroxy, amino, cyano, halogen, C _1-3 straight or branched chain alkyl, or C _1-3 straight or branched alkoxy;
n is 0 to 8;
X and Y are each independently CH or N; And
Z is S, sulfinyl or sulfonyl).
According to claim 1,
The compound represented by Formula 1 is a compound, or a pharmaceutically acceptable salt thereof, characterized in that any one selected from the following group of compounds:
(1) 1-((4-fluorophenyl)sulfinyl)isoquinoline;
(2) 1-((4-fluorophenyl)thio)isoquinoline;
(3) 1-((4-fluorophenyl)sulfonyl)isoquinoline;
(4) 4-((4-fluorophenyl)sulfinyl)quinoline;
(5) 1-((2,4-difluorophenyl)sulfinyl)isoquinoline;
(6) 1-((4-chlorophenyl)sulfinyl)isoquinoline; And
(7) 1-((4-(trifluoromethyl)phenyl)sulfinyl)isoquinoline.
As shown in Scheme 1 below,
Preparing a compound represented by the formula (1) from the compound represented by the formula (4) Method for producing a compound represented by the formula (1) comprising:
[Scheme 1]

(In Scheme 1,
R ¹ , R ² , n, X, Y and Z are as defined in Formula 1 of claim 1).
A pharmaceutical composition for the prevention or treatment of autophagy-related diseases containing the compound represented by Formula 1 of claim 1 or a pharmaceutically acceptable salt thereof as an active ingredient.
According to claim 4,
The autophagy-related diseases include cancer, atherosclerosis, neurodegenerative disease, Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, Huntington's disease, spinocerebellar ataxia, ophthalmology dystrophy muscular dystrophy, prion's disease, fatal familial insomnia, alpha-1 antitrypsin deficiency, dentatorubral pallidoluysian atrophy, frontal lobe dementia, progressive supranuclear palsy, spinal muscular dystrophy (x- A pharmaceutical composition characterized by being selected from the group consisting of linked spinobulbar muscular atrophy and neuronal intranuclear hyaline inclusion disease.
A pharmaceutical composition for the prevention or treatment of an inflammatory disease or metabolic disease, comprising the compound represented by Formula 1 of claim 1 or a pharmaceutically acceptable salt thereof as an active ingredient.
The method of claim 6,
The metabolic disease is characterized in that it is selected from the group consisting of obesity, diabetes, dyslipidemia, alcoholic fatty liver, non-alcoholic fatty liver, hypertension, arteriosclerosis, hyperlipidemia and hyperinsulinemia, pharmaceutical composition.
The method of claim 6,
The inflammatory disease is selected from the group consisting of inflammatory bowel disease, ulcerative colitis, Crohn's disease, arthritis, dermatitis, atopic dermatitis, ophthalmitis, keratitis, hepatitis, non-alcoholic hepatitis, pharmaceutical composition.
A pharmaceutical composition for the prevention or treatment of skin hyperpigmentation disease, which contains the compound represented by Formula 1 of claim 1 or a pharmaceutically acceptable salt thereof as an active ingredient. A cosmetic composition for whitening containing the compound represented by Formula 1 of claim 1 or a salt thereof.
A health functional food composition for preventing or improving autophagy related diseases containing the compound represented by Formula 1 of claim 1 or a pharmaceutically acceptable salt thereof as an active ingredient.

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