Composition for preventing or treating parkinson's disease comprising novel compound

Abstract

The present invention relates to the use of a novel compound for preventing, alleviating or treating Parkinson's disease, said novel compound exhibiting an effect of inhibiting synuclein aggregation in a Parkinson's disease (PD) mouse model. As a result of histological analysis, it has been confirmed that the reduction of dopaminergic neurons is blocked by treatment with the novel compound. Therefore, the novel compound can be effectively used in the development of a Parkinson's disease therapeutic agent.

Description

Technical Field

[0001] This disclosure relates to compositions for the prevention or treatment of Parkinson's disease, said compositions comprising novel compounds. Background Technology

[0002] Parkinson's disease is a progressive neurodegenerative disorder that worsens with age and is caused by a reduction in dopamine neurons present in the dopamine pathway in the midbrain, which is responsible for motor function. Therefore, the main symptoms include motor disturbances such as muscle rigidity and tremors in the limbs.

[0003] The exact cause of Parkinson's disease is unknown, but it is believed to be related to environmental factors (e.g., neurotoxins such as pesticides), genetic factors, mitochondrial dysfunction, and aging. Genetic factors are known to be associated with mutations in genes such as α-synuclein, Parkin, PINK-1, UCH-L1, and DJ-1. Currently, many medications can alleviate the symptoms of Parkinson's disease, but no medication has been reported to stop the progression of the disease, and long-term use of these medications carries a high risk of debilitating side effects.

[0004] Known medications for treating Parkinson's disease include L-DOPA preparations, dopamine receptor agonists, anticholinergic drugs, and idopyryl (Eldepryl). However, most of these medications are not for treating the disease itself, but rather for controlling symptoms and require continuous use. Long-term use of these drugs leads to side effects. For example, anticholinergic drugs can cause autonomic nervous system disorders or mental dysfunction, which limits their continued use in older patients. Furthermore, in the case of L-DOPA preparations, efficacy gradually decreases with long-term intake, and side effects such as body contortions or abnormal movements of the hands and feet occur. In addition, while surgical treatments are also available, such as high-frequency nerve stimulation, high-frequency ablation, or deep brain stimulation, these are invasive procedures and expensive.

[0005] Therefore, since the exact cause of Parkinson's disease is still unknown, treatments mainly focus on relieving symptoms rather than treating the underlying cause, making it urgent to develop new and more effective therapies for the prevention and treatment of Parkinson's disease. Summary of the Invention

[0006] Technical goals

[0007] Therefore, the purpose of this disclosure is to provide uses for the prevention, improvement or treatment of Parkinson's disease, including novel compounds.

[0008] Technical solution

[0009] To achieve the above objectives, this disclosure provides pharmaceutical compositions for the prevention or treatment of Parkinson's disease, said pharmaceutical compositions comprising a compound represented by the following chemical formula 1 or a pharmaceutically acceptable salt thereof as an active ingredient:

[0010] [Chemical Formula 1]

[0011]

[0012] Wherein, in the chemical formula 1,

[0013] Is it a single or double bond?

[0014] n is an integer between 0 and 1.

[0015] X is CH or N, and

[0016] R 1 and R 2 They can be the same or different, and each is independently selected from hydrogen, (C1~C4)alkyl, (C1~C4)alkoxy, hydroxyl, halogen, nitro, cyano or (C1~C4)alkylcarbonyloxy.

[0017] In addition, this disclosure provides a health functional food composition for the prevention or improvement of Parkinson's disease, the food composition comprising a compound represented by chemical formula 1 or a pharmaceutically acceptable salt thereof as an active ingredient.

[0018] Beneficial effects

[0019] This disclosure relates to the use of a novel compound for the prevention, improvement, or treatment of Parkinson's disease, wherein the novel compound (PRG-A compound) exhibits an inhibitory effect on synuclein aggregation in a mouse model of Parkinson's disease (PD). Histological analysis revealed that treatment with the PRG-A compound blocked the reduction of dopaminergic neurons. In summary, the PRG-A compound can be used to develop as a therapeutic agent for Parkinson's disease. Attached Figure Description

[0020] Figure 1 The results of PRG-A compound reducing MT-synuclein aggregation are shown. A. The results of PRG-A compound reducing insoluble forms of MT-synuclein are shown. B. The results of PRG-A-04 dose-dependently inhibiting MT-synuclein aggregation are shown. C. The results of PRG-A compound eliminating synuclein aggregation in SK-N-SH cells overexpressing Syn-A30P are shown.

[0021] Figure 2 shows the results identifying the in vivo effects of PRG-A compounds in a mouse model of Parkinson's disease (PD). A. Schematic diagram of experiments using PRG-A compounds (PRG-A-03, PRG-A-04). B. Results measuring the reduction of dopaminergic neurons in the Syn-A53T Tg mouse model. C. Results showing that PRG-A-03 exhibits high expression of synuclein and inhibits Tau phosphorylation in the brain tissue of Syn-A53T Tg mice. D. Results showing that PRG-A-04 protects against the reduction of dopaminergic neurons in Syn-A53T Tg mice.

[0022] Figure 3 The toxic effects of the PRG-A compound are shown. Detailed Implementation

[0023] This disclosure will be described in detail below.

[0024] This disclosure provides pharmaceutical compositions for the prevention or treatment of Parkinson's disease, the pharmaceutical compositions comprising a compound represented by the following chemical formula 1, its hydrate or salt thereof.

[0025] [Chemical Formula 1]

[0026]

[0027] In the chemical formula 1,

[0028] Is it a single or double bond?

[0029] n is an integer between 0 and 1.

[0030] X is CH or N, and

[0031] R 1 and R 2 They can be the same or different, and each is independently selected from hydrogen, (C1~C4)alkyl, (C1~C4)alkoxy, hydroxyl, halogen, nitro, cyano or (C1~C4)alkylcarbonyloxy.

[0032] Preferably, in the compound, when When it is a single bond, n is 1, X is CH, and R is... 1 and R 2 They can be the same or different and each independently selected from (C1~C4)alkoxy, hydroxy or (C1~C4)alkylcarbonyloxy, but are not limited thereto.

[0033] Preferably, in the compound, when When it is a double bond, n is an integer from 0 to 1, X is CH or N, and R 1 and R2 They can be the same or different and each independently selected from hydrogen, (C1~C4)alkoxy, hydroxyl, halogen, nitro or (C1~C4)alkylcarbonyloxy, but are not limited thereto.

[0034] Preferably, the compound can be a compound represented by the following chemical formula 2.

[0035] [Chemical Formula 2]

[0036]

[0037] In the chemical formula 2,

[0038] n is an integer between 0 and 1, and

[0039] R 1 'and R 2 'They can be the same or different, and each is independently selected from hydrogen, (C1~C4)alkyl, (C1~C4)alkoxy, hydroxyl, halogen, nitro, cyano or (C1~C4)alkylcarbonyloxy.

[0040] Compounds represented by chemical formula 1 or chemical formula 2 can be named PRG-A compounds.

[0041] More preferably, the compound may be selected from, but is not limited to, the group consisting of: (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromen-7-yl(E)-3-(4-hydroxy-3-methoxyphenyl)acrylate (PRG-A-01), (S,E)-7-((3-(4-hydroxy-3-methoxyphenyl)allyl)oxy)-8,8-dimethyl-7,8-dihydro-2H,6H-pyrano[3,2-g]chromen-2-one (PRG-A-02), (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromen-7-yl3-(4-hydroxy-3-methoxy) (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromene-7-yl(E)-3-(3,4-dimethoxyphenyl)acrylate (SNU-C5), (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromene-7-yl3-(3,4-dimethoxyphenyl)propionate (SNU-C7), (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromene-7-yl(E)-3-(pyridin-4-yl)acrylate (SNU-C9), (S)-8,8-dimethyl- 2-Oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromeno-7-yl(E)-3-(3-hydroxyphenyl)acrylate (SNU-C10), (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromeno-7-yl(E)-3-(4-fluorophenyl)acrylate (PRG-A-03), (S,E)-7-((3-(4-fluorophenyl)allyl)oxy)-8,8-dimethyl-7,8-dihydro-2H,6H-pyrano[3,2-g]chromeno-2-one (SNU-C13), (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromeno-2-one (SNU-C13), (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g] Tinno-7-yl(E)-3-(3-acetoxyphenyl)acrylate (SNU-C14), (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]tinno-7-yl 3-(4-acetoxy-3-methoxyphenyl)propionate (SNU-C15), (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]tinno-7-yl(E)-3-(4-acetoxy-3-methoxyphenyl)acrylate (SNU-C17), (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]tinno-7-yl(E)-3-(3,4-Difluorophenyl)acrylate (SNU-C18) and (S,E)-7-((3-(3-methoxy-4-nitrophenyl)allyl)oxy)-8,8-dimethyl-7,8-dihydro-2H,6H-pyrano[3,2-g]chromone-2-one (PRG-A-04).

[0042] More preferably, the pharmaceutical composition can inhibit the aggregation of synuclein and the reduction of dopaminergic neurons.

[0043] In addition to the active ingredient, the pharmaceutical compositions of this disclosure can also be prepared using pharmaceutically suitable and physiologically acceptable adjuvants, which may include solubilizers such as excipients, disintegrants, sweeteners, binders, coating agents, swelling agents, lubricants, flow aids, or flavoring agents. The pharmaceutical compositions of this disclosure can preferably be formulated by including one or more pharmaceutically acceptable carriers in addition to the active ingredient for administration. In compositions formulated as liquid solutions, acceptable pharmaceutical carriers may be sterile and biocompatible carriers used by mixing saline solutions, sterile water, Ringer's solution, buffered saline, albumin injection solutions, dextran solutions, maltodextrin solutions, glycerol, ethanol, and one or more mixtures of these components, and other conventional additives, such as antioxidants, buffers, and antibacterial agents, may be added as needed. Furthermore, diluents, dispersants, surfactants, binders, and lubricants may be added to formulate injectable formulations (e.g., aqueous solutions, suspensions, and emulsions), pills, capsules, granules, or tablets.

[0044] The pharmaceutical compositions of this disclosure can be formulated as sustained-release dosage forms of the active compound, as well as granules, powders, coated tablets, tablets, capsules, suppositories, syrups, elixirs, suspensions, emulsions, drops, or injections. The pharmaceutical compositions of this disclosure can be administered conventionally via intravenous, intra-arterial, intra-peritoneal, intramuscular, intra-arterial, intra-sternal, percutaneous, intranasal, inhalation, topical, rectal, oral, intraocular, or intradermal routes. The effective amount of the active ingredient in the pharmaceutical compositions of this disclosure refers to the amount required for the prevention or treatment of a disease. Therefore, it can be adjusted according to various factors, including the type and severity of the disease, the type and content of the active ingredient and other components contained in the composition, the type of formulation, and the patient's age, weight, general health condition, sex, and diet, as well as the secretion rate of the composition, the time of administration, the route of administration, the treatment duration, and concurrent medications. Although not limited thereto, in the case of adults, for example, when administered once or several times a day, the compositions of this disclosure can be administered at doses from 0.01 ng / kg to 10 g / kg.

[0045] In addition, this disclosure provides a health functional food composition for the prevention or improvement of Parkinson's disease, the food composition comprising a compound represented by the following chemical formula 1, its hydrate or salt thereof.

[0046] [Chemical Formula 1]

[0047]

[0048] In the chemical formula 1,

[0049] Is it a single or double bond?

[0050] n is an integer between 0 and 1.

[0051] X is CH or N, and

[0052] R 1 and R 2 They can be the same or different, and each is independently selected from hydrogen, (C1~C4)alkyl, (C1~C4)alkoxy, hydroxyl, halogen, nitro, cyano or (C1~C4)alkylcarbonyloxy.

[0053] Preferably, in the compound, when When it is a single bond, n is 1, X is CH, and R is... 1 and R 2 They can be the same or different and each independently selected from (C1~C4)alkoxy, hydroxy or (C1~C4)alkylcarbonyloxy, but are not limited thereto.

[0054] Preferably, in the compound, when When it is a double bond, n is an integer from 0 to 1, X is CH or N, and R 1 and R 2 They can be the same or different and each independently selected from hydrogen, (C1~C4)alkoxy, hydroxyl, halogen, nitro or (C1~C4)alkylcarbonyloxy, but are not limited thereto.

[0055] Preferably, the compound can be a compound represented by the following chemical formula 2.

[0056] [Chemical Formula 2]

[0057]

[0058] In the chemical formula 2,

[0059] n is an integer between 0 and 1, and

[0060] R 1 'and R 2'They can be the same or different, and each is independently selected from hydrogen, (C1~C4)alkyl, (C1~C4)alkoxy, hydroxyl, halogen, nitro, cyano or (C1~C4)alkylcarbonyloxy.

[0061] Compounds represented by chemical formula 1 or chemical formula 2 can be named PRG-A compounds.

[0062] More preferably, the compound may be selected from, but is not limited to, the group consisting of: (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromen-7-yl(E)-3-(4-hydroxy-3-methoxyphenyl)acrylate (PRG-A-01), (S,E)-7-((3-(4-hydroxy-3-methoxyphenyl)allyl)oxy)-8,8-dimethyl-7,8-dihydro-2H,6H-pyrano[3,2-g]chromen-2-one (PRG-A-02), (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromen-7-yl3-(4-hydroxy-3-methoxy) (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromene-7-yl(E)-3-(3,4-dimethoxyphenyl)acrylate (SNU-C5), (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromene-7-yl3-(3,4-dimethoxyphenyl)propionate (SNU-C7), (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromene-7-yl(E)-3-(pyridin-4-yl)acrylate (SNU-C9), (S)-8,8-dimethyl- 2-Oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromeno-7-yl(E)-3-(3-hydroxyphenyl)acrylate (SNU-C10), (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromeno-7-yl(E)-3-(4-fluorophenyl)acrylate (PRG-A-03), (S,E)-7-((3-(4-fluorophenyl)allyl)oxy)-8,8-dimethyl-7,8-dihydro-2H,6H-pyrano[3,2-g]chromeno-2-one (SNU-C13), (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromeno-2-one (SNU-C13), (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g] Tinno-7-yl(E)-3-(3-acetoxyphenyl)acrylate (SNU-C14), (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]tinno-7-yl 3-(4-acetoxy-3-methoxyphenyl)propionate (SNU-C15), (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]tinno-7-yl(E)-3-(4-acetoxy-3-methoxyphenyl)acrylate (SNU-C17), (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]tinno-7-yl(E)-3-(3,4-Difluorophenyl)acrylate (SNU-C18) and (S,E)-7-((3-(3-methoxy-4-nitrophenyl)allyl)oxy)-8,8-dimethyl-7,8-dihydro-2H,6H-pyrano[3,2-g]chromone-2-one (PRG-A-04).

[0063] The health functional food compositions disclosed herein may further comprise one or more additives selected from the group consisting of: organic acids, phosphates, antioxidants, lactose casein, dextrin, glucose, sugars, and sorbitol. Organic acids may be, but are not limited to, citric acid, fumaric acid, adipic acid, lactic acid, or malic acid; phosphates may be, but are not limited to, sodium phosphate, potassium phosphate, acid pyrophosphate, or polyphosphate; and antioxidants may be, but are not limited to, natural antioxidants such as polyphenols, catechins, α-tocopherol, rosemary extract, licorice extract, chitosan, tannic acid, or phytic acid.

[0064] In another specific example of this disclosure, in addition to the active ingredients, the health functional food may also contain various nutrients, vitamins, minerals (electrolytes), flavoring agents (e.g., synthetic and natural flavoring agents), coloring agents and thickeners (cheese, chocolate, etc.), pectic acid and its salts, alginic acid and its salts, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohols, and carbonating agents used in carbonated beverages. Furthermore, a food composition according to one embodiment of this disclosure may contain fruit pulp used in the production of natural fruit juices, fruit juice beverages, and vegetable beverages.

[0065] According to one embodiment of this disclosure, the formulation of a health functional food may be in the form of a solid, powder, granules, tablet, capsule, liquid, or beverage, but is not limited thereto.

[0066] In addition, health functional foods can be used in the manufacture of foods, such as, but not limited to, sweets, sugar, ice cream products, dairy products, meat products, fish products, tofu or jelly, edible oils, noodles, tea, beverages, special nutritional foods, health products, condiments, frozen foods, ginseng products, pickled foods, dried foods, fruits, vegetables, dried fruits or vegetables, cut products, fruit juices, vegetable juices, their mixtures, potato chips, noodles, processed livestock and poultry foods, processed seafood foods, processed dairy foods, fermented dairy foods, legume foods, cereal foods, microbially fermented foods, pastries and baked goods, seasonings, processed meat products, acidic beverages, licorice and herbs.

[0067] Example

[0068] In the following description, embodiments will be presented in detail to aid in understanding this disclosure. However, the following embodiments are intended to illustrate the content of this disclosure only, and the scope of this disclosure is not limited to the following embodiments. The embodiments of this disclosure are provided to describe this disclosure more completely to those skilled in the art.

[0069] <Experimental Example>

[0070] The following experimental examples are intended to provide experimental examples that are typically applied to each embodiment of this disclosure.

[0071] 1. Preparation of the compound and mice

[0072] 1) Compounds

[0073] The chemical structures and NMR data of the compounds used in this disclosure are as follows.

[0074] (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromene-7-yl(E)-3-(4-hydroxy-3-methoxyphenyl)acrylate (PRG-A-01)

[0075]

[0076] 1 1H NMR (acetone-d6, 400MHz)

[0077] δ ppm 7.843(d, J=9.6Hz, 1H), 7.616(d, J=16.0Hz, 1H), 7.424(s, 1H), 7.357(s, 1H), 7.123(dd, J=2.0, 8.0Hz, 1H), 6.851(d, J=8.4Hz, 1H), 6.740(s,1H), 6.387(d, J=15.6Hz, 1H), 6.198(d, J=9.6Hz, 1H), 5.221(t, J=4.6Hz, 1H),3.895(s, 3H), 3.321(dd, J=4.6, 17.2Hz, 1H), 2.963(dd, J=4.4, 17.6Hz, 1H),1.421(s, 3H), 1.413(s, 3H); MS(m / z) 423 (M+H) + .

[0078] (S,E)-7-((3-(4-hydroxy-3-methoxyphenyl)allyl)oxy)-8,8-dimethyl-7,8-dihydro-2H,6H-pyrano[3,2-g]chromen-2-one (PRG-A-02)

[0079]

[0080] 1 H NMR: EW15731-164-P1D4 (400 MHz, CDCl3)

[0081] δ 7.58 (d, J = 12 Hz, 1H), 7.16 (s, 1H), 6.90 - 6.86 (m, 3H), 6.78(s, 1H), 6.51 (d, J =16 Hz, 1H), 6.22 (d, J = 8.0 Hz, 1H), 6.14 - 6.10 (m,1H), 5.65 (s, 1H), 4.32 - 4.30 (m, 1H), 4.22- 4.20 (m, 1H), 3.91 (s, 3H), 3.61 - 3.58 (m, 1H), 3.11 - 3.05 (m, 1H), 3.89 - 3.87 (m, 1H), 1.43 (s, 3H), 1.36 (s, 3H).

[0082] (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromene-7-yl(E)-3-(4-fluorophenyl)acrylate (PRG-A-03)

[0083]

[0084] 1 H NMR (CD2Cl2, 400MHz): δ ppm 7.66 - 7.55 (m, 2H), 7.55 - 7.46 (m,2H), 7.18 (s, 1H), 7.11 - 7.01 (m, 2H), 6.76 (s, 1H), 6.35 (d, J = 16.0 Hz,1H), 6.16 (d, J = 9.5 Hz, 1H), 5.17 (app.t, J = 4.7 Hz, 1H), 3.23 (ddd, J =17.3, 4.8, 1.2 Hz, 1H), 2.92 (dd, J = 17.3, 4.7 Hz, 1H), 1.40 (s, 3H), 1.36 (s, 3H).

[0085] (S,E)-7-((3-(3-methoxy-4-nitrophenyl)allyl)oxy)-8,8-dimethyl-7,8-dihydro-2H,6H-pyrano[3,2-g]chromene-2-one (PRG-A-04)

[0086]

[0087] 1 H NMR (CDCl3, 400MHz): δ ppm 7.85 (d, J = 8.4 Hz, 1H), 7.57 (d, J =9.5 Hz, 1H), 7.16 (s, 1H), 7.05 - 6.97 (m, 2H), 6.78 (s, 1H), 6.60 4.24 (ddd, J = 13.4, 5.5, 1.6Hz, 1H), 3.97 (s, 3H), 3.59 (dd, J = 7.1, 4.9 Hz, 1H), 3.11 (dd, J = 16.7,4.9 Hz, 1H), 2.87 (dd, J = 16.7, 7.2 Hz, 1H), 1.43 (s, 3H), 1.38 (s, 3H).

[0088] (S,E)-7-((3-(4-fluorophenyl)allyl)oxy)-8,8-dimethyl-7,8-dihydro-2H,6H-pyrano[3,2-g]chromene-2-one (SNU-C13)

[0089]

[0090] 1H NMR (CDCl3, 400MHz): δ ppm 7.57 (d, J = 9.5 Hz, 1H), 7.38 - 7.29(m, 2H), 7.16 (s, 1H), 7.06 - 6.93 (m, 2H), 6.77 (s, 1H), 6.55 (d, J = 15.8Hz, 1H), 6.25 - 6.13 (m, 2H), 4.33 (ddd, J = 12.8, 5.8, 1.5 Hz, 1H), 4.19 (ddd, J = 12.8, 6.3, 1.4 Hz, 1H), 3.58 (dd, J = 7.4, 5.0 Hz, 1H), 3.08 (dd, J= 16.4, 5.0 Hz, 1H), 2.85 (ddd, J = 16.4, 7.4, 1.1 Hz, 1H), 1.42 (s, 3H), 1.35 (s, 3H).

[0091] (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromene-7-yl(E)-3-(4-acetoxy-3-methoxyphenyl)acrylate (SNU-C17)

[0092]

[0093] 1 H NMR (CD2Cl2, 600MHz): δ ppm 7.67 - 7.56 (m, 2H), 7.21 (s, 1H), 7.14- 7.09 (m, 2H), 7.03 (d, J = 8.0 Hz, 1H), 6.78 (s, 1H), 6.41 (d, J = 16.0 Hz, 1H), 6.19 (d, J = 9.5 Hz, 1H), 5.20 (app.t, J = 4.7 Hz, 1H), 3.82 (s, 3H), 3.26 (ddd, J = 17.3, 4.8, 1.2 Hz, 1H), 2.95 (dd, J = 17.3, 4.5 Hz, 1H), 2.27(s, 3H), 1.43 (s, 3H), 1.38 (s, 3H).

[0094] (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromene-7-yl(E)-3-(3,4-difluorophenyl)acrylate (SNU-C18)

[0095]

[0096] 1 H NMR (CDCl3, 400MHz): δ ppm 7.62 - 7.53 (m, 2H), 7.32 (ddd, J =11.1, 7.6, 2.2 Hz, 1H), 7.24 - 7.11 (m, 3H), 6.83 (s, 1H), 6.33 (d, J = 16.0,1H), 6.24 (d, J = 9.4 Hz, 1H), 5.19 (app.t, J = 4.7 Hz, 1H), 3.25 (ddd, J =17.4, 4.8, 1.1 Hz, 1H), 2.93 (dd, J = 17.4, 4.5 Hz, 1H), 1.43 (s, 3H), 1.39 (s, 3H).

[0097] (7S)-(+)-3-(2-furanyl)-acrylic acid, 8,8-dimethyl-2-oxo-6,7-dihydro-2H,8H-pyrano[3,2-g]chromene-7-yl-ester (SLC-B050)

[0098]

[0099] 1 H NMR (400 MHz, CDCl3): δH 7.64(1H, s, H-6'), 7.58(1H, d, J = 9.6Hz, H-4), 7.56(1H, d, J = 16.0Hz, H-3'), 7.41(1H, d, J = 1.6Hz, H-7'), 7.17(1H,s, H-5), 6.82(1H, s, H-10), 6.55(1H, d, J = 1.6Hz, H-8'), 6.23(1H, d, J =9.6Hz, H-3), 6.13(1H, d, J = 16.0Hz, H-2'), 5.17(1H, t, J = 4.4Hz, H-7), 3.23(1H, dd, J = 4.4, 17.6Hz, H-6a), 2.92(1H, dd, J = 4.4, 17.6Hz, H-6b), 1.42(3H, s, CH3-8), 1.38(3H, s, CH3-8).

[0100] 2) Mice

[0101] Experiments were conducted in a facility approved by the International Committee for Assessment and Accreditation of Laboratory Animal Care (ICIAC) in accordance with the animal policy approved by Pusan ​​National University. B6.Cg-Tg (Prnp-SNCAA53T) / J mice were obtained from Jackson Laboratory (catalog number: 006823). All mice were maintained under controlled light and temperature conditions (20°C–23°C, 12h–12h light / dark cycle) and provided with autoclaved food and water.

[0102] 2. In vivo drug processing and histological analysis

[0103] Syn at 24 weeks of age A53T-Tg Mice were administered DMSO, PRG-A-03, and PRG-A-04 intraperitoneally twice weekly for 24 to 26 weeks. Control mice were treated under the same conditions. Mice were sacrificed at 48–52 weeks of age for histological analysis. After cardiac perfusion with 4% paraformaldehyde, the brain was dissected and embedded in paraffin blocks according to standard tissue processing procedures. The embedded tissue (cervical and lumbar regions of the spinal cord) was sectioned into 5 μm sections using a tissue sectioner (Leica microtome) and transferred to adhesive-coated slides (Marienfeld laboratory glassware, Germany). For Syn A53T-Tg Histopathological analysis of mice involved immunohistochemistry (IHC) using dopaminergic neuron markers (tyrosine hydroxylase) after dewaxing and rehydration. To monitor dopaminergic neurons by immunofluorescence (IF), dissected brain tissue was embedded in agarose scaffolds and sectioned using a vibratory microtome. Thin sections were treated with blocking solution (normal donkey serum diluted 1:500 in 0.1% Triton X-100 PBS) for 1 hour. Cells were incubated overnight at 4°C with anti-TH antibody (1:500 dilution). Finally, cells were incubated with FITC-conjugated secondary antibody at 4°C for 2 hours. Cells were washed three times with PBS and mounted on coverslips using mounting solution (H-5501, Vector Laboratories) for analysis under a fluorescence microscope (Zeiss).

[0104] 3. Cell Culture and Reagents

[0105] SK-N-SH cells were purchased from the Korean Cell Bank (KCLB, Seoul, South Korea) and maintained in MEM medium containing 10% fetal bovine serum, 1% antibiotics, 25 mM HEPES, and 300 mg / L L-Glu. Human fibroblasts (9-year-old female) were obtained from Coridll Cell Repositories (New Jersey, USA) and maintained in EMEM medium containing 15% FBS, 2 mM glutamine, and 26 mM HEPES, but without antibiotics.

[0106] 4. Western blot analysis

[0107] For SDS-PAGE, proteins were extracted from cells using RIPA buffer (50 mM Tris-Cl, pH 7.5, 150 mM NaCl, 1% NP-40, 0.1% SDS, and 10% sodium deoxycholate). Samples were separated by SDS-PAGE and transferred to PVDF membranes. The blotted membranes were blocked with 3% skim milk containing TBST buffer for 1 h and incubated with specific antibodies. The reacted antibodies were detected by ECL and X-ray film exposure. The antibody used in this experiment was pan-Syn (GTX112799), purchased from Genetex (California, USA). Actin antibody (66009-1-lg) and α-tubulin antibody (66240-1-lg) were obtained from Proteintech (Rosemont, IL, USA). Anti-FLAG (Sigma; F3165) from Sigma Aldrich (St. Louis, MO, USA) was used as the secondary antibody, along with HRP-conjugated goat anti-mouse, goat anti-rabbit, and mouse anti-goat antibodies (Pierce, Thermo Fisher Scientific, Inc., Rockford, IL, USA). FLAG-Syn-WT and MT vectors were transfected into SK-N-SH cells for 24 h. After incubation, cells were harvested with TNN buffer (50 mM Tris-Cl, pH 7.5, 150 mM NaCl, 0.3% NP-40) and centrifuged at 14,000 rpm for 30 min to separate the precipitate (insoluble) and supernatant (soluble). Input represents whole-cell lysate (WCL) harvested with RIPA buffer. Actin was used as a loading control.

[0108] 5. Immunofluorescence staining

[0109] Cells on coverslips were washed with PBS, fixed with 4% paraformaldehyde (PFA) at room temperature for 30 min, and then permeabilized in 0.1% Triton X-100 / PBS for 10 min. After treating cells with blocking solution (3% anti-human antibody diluted 1:500 in PBS) for 1 hour, cells were incubated overnight at 4°C with anti-FLAG (diluted 1:400). Finally, cells were incubated with FITC- and rhodamine-conjugated secondary antibodies at 4°C for 6 hours. Cell nuclei were stained with 4,6-diamidinyl-2-phenylindole (DAPI), and endoplasmic reticulum (ER) was stained with ER-Tracker Red dye for 10 min. Cells were washed three times with PBS, and then mounted with mounting solution (H-5501; Vector Laboratories, Burlingame, CA, USA) for analysis using a fluorescence microscope (Zeiss).

[0110] 6. Transfection of the vector

[0111] FLAG-Syn (WT, A30P, and A53T) vectors were purchased from Dr. Seol (Wonkwang University Korea). Transfection was performed using Jet-PEI reagent (JetPEI; Polyplus transfection, New York, NY, USA) according to the manufacturer's protocol. In short, the vectors were mixed with JetPEI reagent in 150 mM NaCl buffer, and the mixture was incubated for 15 minutes. The mixture was then added to cells in serum-free medium and incubated for 4 hours. After incubation, the cells were replaced with medium supplemented with 10% FBS.

[0112] 7. Measurement of cell viability

[0113] To study cell viability, cells were treated with a chemical (PRG-A compound) for 48 h. Then, the cells were incubated with 0.5 mg / mL 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) solution (475989; Merck, Darmstadt, Germany) at 37 °C for 4 h. After removing excess solution and washing with PBS, the precipitate was dissolved in 200 μL DMSO and quantified by measuring absorbance at 540 nm.

[0114] <Example 1>

[0115] To identify the effect of the PRG-A compound on α-synuclein aggregation, the inventors transfected SK-N-SH cells with the FLAG-Syn-A30P vector for 24 hours. After incubation, the cells were treated with the compound (5 μM) for 24 hours. As a result, most of the ectopically expressed mutant α-synuclein (Syn-A30P) was recovered as an insoluble fraction. Figure 1 A). However, treatment with PRG-A compounds significantly reduced synuclein aggregation ( Figure 1 A). Furthermore, SK-N-SH cells were transfected with the indicator vector (MT-Syn) for 24 h, followed by treatment with PRG-A-04 for 24 h. Cells were then harvested using TNN buffer (50 mM Tris-Cl, pH 7.5, 150 mM NaCl, 0.3% NP-40) and centrifuged at 14,000 rpm for 30 min to separate the precipitate (insoluble) and the supernatant (soluble). Actin was used as a loading control. Figure 1 B). Similar effects were observed in experiments using another synuclein mutant (Syn-A53T). Figure 1 B), in which PRG-A-04 can reduce insoluble synuclein in a dose-dependent manner. To observe this, immunofluorescence staining with Syn-Ab was performed. Cells were transfected with the Syn-A53T vector for 24 hours. After culture, cells were treated with PRG-A compound (5 μM) for 24 hours and fixed with 4% PFA. By treating with PRG-A compound, aggregated synuclein (white arrow) was dissolved ( Figure 1 C). Synuclein expression observed under a fluorescence microscope using pan-Syn antibody (white arrow; high intensity of Syn). Chem-001 (decursinol) shows marginal effects in inhibiting synuclein aggregation.

[0116] <Example 2>

[0117] To investigate the in vivo effects of the PRG-A compounds, 24-week-old mice were intraperitoneally injected with 20 mg / kg or 40 mg / kg of the PRG-A compounds (PRG-A-03, PRG-A-04) for 24 to 26 weeks. Mice were then sacrificed for histological analysis, and brain tissue was dissected at 48 to 49 weeks of age (Fig. 2A). Dopaminergic neurons in the substantia nigra (SN) were observed using tyrosine hydroxylase (TH) staining (Fig. 2B). In mice injected with PRG-A-03, the dopaminergic neuronal marker (tyrosine hydroxylase, TH) was maintained. Representative images are shown at 10x magnification and 20x inset (yellow box) (Fig. 2B). Compound treatment blocked the reduction of dopaminergic neurons (Fig. 2B). Furthermore, the expression of the mutant Syn was reduced in brain tissue (Fig. 2C). In addition, the dissected brain was embedded in an agarose scaffold and sectioned using a vibratory microtome. Thin sections of tissue were stained with TH antibody (Fig. 2D). As a result, similar results were obtained with PRG-A-04 treatment using IF staining with TH, which prevented the reduction of dopaminergic neurons in SN (Figure 2D). These results indicate that the PRG-A compound not only blocks synuclein (Syn) aggregation, but also protects against neuronal reduction in Parkinson's disease (PD).

[0118] <Example 3>

[0119] Finally, to determine the toxicity of the PRG-A compounds, normal human fibroblasts and neuroblastoma cells (SK-N-SH) were cultured with each compound for 48 hours, and cell viability was measured by MTT assay. The effects on the survival of neurons and normal fibroblasts were identified. Figure 3 Compounds PRG-A-01, PRG-A-02, and PRG-A-04 did not exhibit cytotoxic effects in normal human fibroblasts. Compound PRG-A-03 showed low effects on cell viability at high doses (50 μM and 100 μM). In summary, the novel compounds did not show toxicity in normal cells at concentrations up to 50 μM, therefore, the side effects of these new compounds are not expected to be severe.

[0120] In summary, this novel compound can be used to treat Parkinson's disease (PD) without serious side effects.

[0121] Although specific portions of this disclosure have been described in detail above, it will be apparent to those skilled in the art that such detailed description is merely a preferred exemplary embodiment, and the scope of this disclosure is not limited thereto. In other words, the essential scope of this disclosure will be defined by the appended claims and their equivalents.

Claims

1. A pharmaceutical composition for the prevention or treatment of Parkinson's disease, said pharmaceutical composition comprising a compound represented by the following chemical formula 1, its hydrate, or a salt thereof: [Chemical Formula 1] in, In the chemical formula 1, Is it a single or double bond? n is an integer between 0 and 1. X is CH or N, and R 1 and R 2 They may be the same or different, and each is independently selected from hydrogen, (C1~C4)alkyl, (C1~C4)alkoxy, hydroxyl, halogen, nitro, cyano or (C1~C4)alkylcarbonyloxy.

2. The pharmaceutical composition according to claim 1, wherein, In the compound, when When it is a single bond, n is 1, X is CH, and R is... 1 and R 2 They are the same or different and are each independently selected from (C1~C4)alkoxy, hydroxy or (C1~C4)alkylcarbonyloxy.

3. The pharmaceutical composition according to claim 1, wherein, In the compound, when When it is a double bond, n is an integer from 0 to 1, X is CH or N, and R 1 and R 2 They are the same or different and are each independently selected from hydrogen, (C1~C4)alkoxy, hydroxyl, halogen, nitro or (C1~C4)alkylcarbonyloxy.

4. The pharmaceutical composition according to claim 1, wherein, The compounds are selected from the group consisting of: (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromen-7-yl(E)-3-(4-hydroxy-3-methoxyphenyl)acrylate, (S,E)-7-((3-(4-hydroxy-3-methoxyphenyl)allyl)oxy)-8,8-dimethyl-7,8-dihydro-2H,6H-pyrano[3,2-g]chromen-2-one, (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromen-7-yl 3-(4-hydroxy-3-methoxyphenyl)propionate, (S)-8,8-dimethyl-2-oxo 7,8-Dihydro-2H,6H-pyrano[3,2-g]chromene-7-yl(E)-3-(3,4-dimethoxyphenyl)acrylate, (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromene-7-yl 3-(3,4-dimethoxyphenyl)propionate, (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromene-7-yl(E)-3-(pyridin-4-yl)acrylate, (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromene-7-yl(E)-3-(3-hydroxyphenyl) Acrylates, (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromene-7-yl(E)-3-(4-fluorophenyl)acrylates, (S,E)-7-((3-(4-fluorophenyl)allyl)oxy)-8,8-dimethyl-7,8-dihydro-2H,6H-pyrano[3,2-g]chromene-2-one, (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromene-7-yl(E)-3-(3-acetoxyphenyl)acrylates, (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromene-7-yl(E)-3-(3-acetoxyphenyl)acrylates, (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g] Tinmen-7-yl 3-(4-acetoxy-3-methoxyphenyl)propionate, (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]tinmen-7-yl(E)-3-(4-acetoxy-3-methoxyphenyl)acrylate, (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]tinmen-7-yl(E)-3-(3,4-difluorophenyl)acrylate, and (S,E)-7-((3-(3-methoxy-4-nitrophenyl)allyl)oxy)-8,8-dimethyl-7,8-dihydro-2H,6H-pyrano[3,2-g]tinmen-2-one.

5. The pharmaceutical composition according to claim 1, wherein, The compound is represented by the following chemical formula 2: [Chemical Formula 2] In the chemical formula 2, n is an integer between 0 and 1, and R 1 'and R 2 'They are the same or different, and each is independently selected from hydrogen, (C1~C4)alkyl, (C1~C4)alkoxy, hydroxyl, halogen, nitro, cyano or (C1~C4)alkylcarbonyloxy.

6. The pharmaceutical composition according to claim 1, wherein, The pharmaceutical composition inhibits the aggregation of synuclein and the reduction of dopaminergic neurons.

7. A health functional food composition for the prevention or improvement of Parkinson's disease, said health functional food composition comprising a compound represented by the following chemical formula 1, its hydrate, or a salt thereof: [Chemical Formula 1] in, In the chemical formula 1, Is it a single or double bond? n is an integer between 0 and 1. X is CH or N, and R 1 and R 2 They may be the same or different, and each is independently selected from hydrogen, (C1~C4)alkyl, (C1~C4)alkoxy, hydroxyl, halogen, nitro, cyano or (C1~C4)alkylcarbonyloxy.

8. The health functional food composition according to claim 7, wherein, In the compound, when When it is a single bond, n is 1, X is CH, and R is... 1 and R 2 They are the same or different and are each independently selected from (C1~C4)alkoxy, hydroxy or (C1~C4)alkylcarbonyloxy.

9. The health functional food composition according to claim 7, wherein, In the compound, when When it is a double bond, n is an integer from 0 to 1, X is CH or N, and R 1 and R 2 They are the same or different and are each independently selected from hydrogen, (C1~C4)alkoxy, hydroxyl, halogen, nitro or (C1~C4)alkylcarbonyloxy.

10. The health functional food composition according to claim 7, wherein, The compounds are selected from the group consisting of: (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromen-7-yl(E)-3-(4-hydroxy-3-methoxyphenyl)acrylate, (S,E)-7-((3-(4-hydroxy-3-methoxyphenyl)allyl)oxy)-8,8-dimethyl-7,8-dihydro-2H,6H-pyrano[3,2-g]chromen-2-one, (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromen-7-yl 3-(4-hydroxy-3-methoxyphenyl)propionate, (S)-8,8-dimethyl-2-oxo 7,8-Dihydro-2H,6H-pyrano[3,2-g]chromene-7-yl(E)-3-(3,4-dimethoxyphenyl)acrylate, (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromene-7-yl 3-(3,4-dimethoxyphenyl)propionate, (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromene-7-yl(E)-3-(pyridin-4-yl)acrylate, (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromene-7-yl(E)-3-(3-hydroxyphenyl) Acrylates, (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromene-7-yl(E)-3-(4-fluorophenyl)acrylates, (S,E)-7-((3-(4-fluorophenyl)allyl)oxy)-8,8-dimethyl-7,8-dihydro-2H,6H-pyrano[3,2-g]chromene-2-one, (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromene-7-yl(E)-3-(3-acetoxyphenyl)acrylates, (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]chromene-7-yl(E)-3-(3-acetoxyphenyl)acrylates, (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g] Tinmen-7-yl 3-(4-acetoxy-3-methoxyphenyl)propionate, (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]tinmen-7-yl(E)-3-(4-acetoxy-3-methoxyphenyl)acrylate, (S)-8,8-dimethyl-2-oxo-7,8-dihydro-2H,6H-pyrano[3,2-g]tinmen-7-yl(E)-3-(3,4-difluorophenyl)acrylate, and (S,E)-7-((3-(3-methoxy-4-nitrophenyl)allyl)oxy)-8,8-dimethyl-7,8-dihydro-2H,6H-pyrano[3,2-g]tinmen-2-one.

11. The health functional food composition according to claim 7, wherein, The compound is represented by the following chemical formula 2: [Chemical Formula 2] In the chemical formula 2, n is an integer between 0 and 1, and R 1 'and R 2 'They are the same or different, and each is independently selected from hydrogen, (C1~C4)alkyl, (C1~C4)alkoxy, hydroxyl, halogen, nitro, cyano or (C1~C4)alkylcarbonyloxy.

Images