Α-synuclein phagocytosis promoter, oral composition for promoting α -synuclein phagocytosis, and use of compound

Abstract

Provided is an α -synuclein phagocytosis promoter comprising plasmalogen or a compound represented by formula (I), a racemic body thereof, or a salt thereof. [In formula (I): X represents a carbon atom or an oxygen atom; R1 represents a saturated aliphatic hydrocarbon group that optionally includes a ring structure; R2 represents a saturated or unsaturated aliphatic hydrocarbon group that has 10-30 carbon atoms and that optionally has a substituent which is an alkyl group having 1-4 carbon atoms or an alkoxy group having 1-4 carbon atoms; carbon atoms of the glycerol skeleton optionally have substituents; and R3 is represented by formula (A) that optionally has a substituent, where * represents the oxygen atom bound to R3.]

Description

Alpha-synuclein phagocytosis promoter, oral composition for promoting alpha-synuclein phagocytosis, and use of the compound

[0001] The present invention relates to the use of an agent for promoting alpha-synuclein phagocytosis, an oral composition and a compound for promoting alpha-synuclein phagocytosis.

[0002] α-Synuclein is a 14-19 kDa phosphorylated protein that is primarily localized in the presynaptic terminals of mature neurons and is involved in the regulation of synaptic function and plasticity. Under normal conditions, α-synuclein exists as a monomer in a naturally unfolded state. On the other hand, under pathological conditions, α-synuclein undergoes structural changes and folds upon binding and interaction with liquid droplets, phospholipid bilayers, lipid membranes, etc., forming an α-helical secondary structure and forming aggregates containing dimers, oligomers, or fibrous molecules.

[0003] α-Synuclein aggregates are known to induce toxicity in cells. These aggregates are the main component of Lewy bodies, which are abnormal protein aggregates found in neurons in various diseases such as Parkinson's disease, multiple system atrophy, and dementia with Lewy bodies. Post-translational modifications of α-synuclein, such as phosphorylation and ubiquitination, are also known to be associated with α-synuclein aggregation and neurotoxicity. α-Synuclein aggregates are known to be associated with a group of neurodegenerative diseases called synucleinopathies (α-synucleinopathies), including Parkinson's disease, multiple system atrophy, dementia with Lewy bodies, and other axonal diseases.

[0004] Patent Document 1 discloses that, based on the finding that Fas-associated factor 1 (FAF1), a protein that binds to the apoptosis receptor Fas, increases the accumulation and aggregation of α-synuclein in neuronal cells, aminopyrazole derivatives, which are FAF1 inhibitors, reduce the accumulation and aggregation of α-synuclein in neuronal cells. Furthermore, Patent Document 2 discloses that anti-α-synuclein antibodies inhibit the aggregation of α-synuclein induced by α-synuclein filaments in mouse primary neurons, and discloses the use of anti-α-synuclein antibodies in the treatment of synucleinopathy.

[0005] JP 2023-118923 A JP 2023-127585 A

[0006] If the clearance of α-synuclein and its aggregates in the nervous system can be promoted, diseases caused by the accumulation of α-synuclein can be treated.

[0007] The present invention has been made in view of the above-mentioned circumstances, and aims to provide an α-synuclein phagocytosis promoter that can promote the clearance of α-synuclein, an oral composition for promoting α-synuclein phagocytosis, and use of the compound.

[0008] An α-synuclein phagocytosis promoter according to a first aspect of the present invention comprises a plasmalogen, or a compound represented by formula (I), its racemate, or a salt thereof. [In formula (I), X represents a carbon atom or an oxygen atom, and R １ represents a saturated aliphatic hydrocarbon group which may contain a cyclic structure, and R ２ represents a saturated or unsaturated aliphatic hydrocarbon group having 10 to 30 carbon atoms which may have a substituent, which is an alkyl group having 1 to 4 carbon atoms or an alkoxy group having 1 to 4 carbon atoms, the carbon atoms of the glycerol skeleton may have a substituent, R ３ The oxygen atom bonded to is *, and R ３ may have a substituent It is.]

[0009] An oral composition for promoting α-synuclein phagocytosis according to a second aspect of the present invention comprises a plasmalogen, or a compound represented by formula (I), its racemate, or a salt thereof. [In formula (I), X represents a carbon atom or an oxygen atom, and R １ represents a saturated aliphatic hydrocarbon group which may contain a cyclic structure, and R ２ represents a saturated or unsaturated aliphatic hydrocarbon group having 10 to 30 carbon atoms which may have a substituent, which is an alkyl group having 1 to 4 carbon atoms or an alkoxy group having 1 to 4 carbon atoms, the carbon atoms of the glycerol skeleton may have a substituent, R ３ The oxygen atom bonded to is *, and R ３ may have a substituent It is.]

[0010] X is an oxygen atom, and R １ is an alkyl group having 18 carbon atoms.

[0011] R ２ is R ２ It may be a monovalent group which, when expressed as COOH, represents arachidonic acid, oleic acid, or palmitic acid.

[0012] Use of a compound according to a third aspect of the present invention is use of a plasmalogen, or a compound represented by formula (I), its racemate, or a salt thereof, for the production of an agent for promoting α-synuclein phagocytosis or an oral composition for promoting α-synuclein phagocytosis. [In formula (I), X represents a carbon atom or an oxygen atom, and R １ represents a saturated aliphatic hydrocarbon group which may contain a cyclic structure, and R ２ represents a saturated or unsaturated aliphatic hydrocarbon group having 10 to 30 carbon atoms which may have a substituent, which is an alkyl group having 1 to 4 carbon atoms or an alkoxy group having 1 to 4 carbon atoms, the carbon atoms of the glycerol skeleton may have a substituent, R ３ The oxygen atom bonded to is *, and R ３ may have a substituent It is.]

[0013] According to the present invention, the clearance of α-synuclein can be promoted.

[0014] FIG. 1 is a diagram showing a separation method by high performance liquid chromatography (HPLC) of plasmalogen used in Test Example 1. FIG. 2 is a diagram showing a chromatogram of a lipid fraction extracted by HPLC. FIG. 3 is a diagram showing a chromatogram of ethanolamine-type plasmalogen. FIG. 4 is a diagram showing a fluorescence microscope image according to Test Example 1. FIG. 5 is a diagram showing the proportion of α-synuclein-positive cells according to Test Example 1. FIG. 6 is a diagram showing a fluorescence microscope image according to Test Example 2. FIG. 7 is a diagram showing the proportion of α-synuclein-positive cells according to Test Example 2. FIG. 8 is a diagram showing a fluorescence microscope image according to Test Example 3. FIG. 9 is a diagram showing the proportion of α-synuclein-positive cells according to Test Example 3.

[0015] Embodiments of the present invention will be described with reference to the drawings. Note that the present invention is not limited to the following embodiments and drawings. Note that in the following embodiments, the expressions "have," "include," or "contain" also include the meaning of "consisting of" or "consisting of."

[0016] The α-synuclein phagocytosis promoter according to this embodiment comprises a plasmalogen, a compound represented by formula (I), its racemate, or a salt thereof. Plasmalogens belong to a subclass of glycerophospholipids characterized by having a vinyl ether bond at the sn-1 position of the glycerol backbone and an ester bond at the sn-2 position. Plasmalogens are present at high concentrations in the cell membranes of many mammalian tissues.

[0017] The plasmalogen contained in the α-synuclein phagocytosis promoter is not particularly limited as long as it is generally classified as a plasmalogen. Examples of plasmalogens include those containing a —CH group at the oxygen atom bonded to phosphorus at the sn-3 position of the glycerol backbone. ２ CH ２ NH ２ an ethanolamine-type plasmalogen having a structure in which -CH is bonded to the oxygen atom bonded to phosphorus at the sn-3 position of the glycerol skeleton; ２ CH ２ N (CH ３ ) ３ Examples of such plasmalogens include choline-type plasmalogens, inositol-type plasmalogens, and serine-type plasmalogens, which have a structure in which the following are bound:

[0018] Preferably, plasmalogen is extracted from animal tissue. The animal tissue is not particularly limited as long as it contains plasmalogen. For example, the animal tissue is exemplified by tissue of an animal selected from the group consisting of shellfish, sea squirts, and birds. More specifically, tissue of aquatic animals such as shellfish, sea squirts, sea cucumbers, salmon, saury, and bonito is preferred as the animal tissue. Preferably, the animal tissue is shellfish tissue. Among animal tissues, edible parts are preferred. The animal tissue may be cut tissue, or may be pulverized tissue to more efficiently extract plasmalogen.

[0019] Examples of shellfish include bivalves or gastropods such as scallops, mussels, and abalone. Scallops are particularly preferred as shellfish. Scallops are edible bivalves belonging to the Pectenidae family, and examples thereof include scallops belonging to the Mizuhopecten or Pecten genus. Specific examples of shellfish include scallops (Mizuhopectenyessoensis) harvested in Japan and European scallops (Pectenmaximus (Linnaeus)) harvested in Europe. Examples of edible parts of scallops include the adductor muscle and string.

[0020] Sea squirts are edible chordates belonging to the family Halocynthia, including those belonging to the genus Halocynthia or Halocynthia. Specific examples of sea squirts include the Halocynthia loretzi and the Halocynthia aurantium. The edible part of the sea squirt is the flesh (fascia).

[0021] Birds are, for example, edible birds. Examples of birds include chickens, silkie chickens, ducks, etc. As for bird tissue, breast meat is preferred because it is rich in plasmalogens.

[0022] Plasmalogen can be extracted using an organic solvent or a water-containing organic solvent, preferably in combination with an enzyme treatment. Examples of methods for extracting plasmalogen include ethanol extraction and hexane extraction.

[0023] In the ethanol extraction method, animal tissues are exposed to an extracting solution containing ethanol. The extracting solution containing ethanol used in the extraction step may be aqueous ethanol or a mixture of ethanol and another organic solvent. The ethanol concentration in the extracting solution is 50% by mass or more, preferably 80% by mass or more, more preferably 95% by mass or more, and may be 100% by mass. The amount of ethanol used is, for example, preferably 100 to 3000 ml, more preferably 200 to 2000 ml, and even more preferably 250 to 1000 ml per 100 g of animal tissue. Extraction using ethanol may be performed multiple times.

[0024] Preferably, the animal tissue exposed to the ethanol-containing extract is animal tissue treated with a protease. Examples of proteases include proteases derived from filamentous fungi such as Aspergillus and Rhizopus; proteases derived from bacteria such as Bacillus; and proteases extracted from plants such as papaya and pineapple. Commercially available proteases may also be used. Proteases derived from filamentous fungi are particularly preferred, with proteases derived from koji mold being even more preferred. Furthermore, since plasmalogens are unstable in acidic and basic conditions, neutral proteases are preferred. The amount of protease used is, for example, 0.1 to 10.0 g, preferably 0.2 to 8.0 g, and more preferably 0.3 to 5.0 g per 100 g of animal tissue.

[0025] After extraction, the solid content is removed, the extract is recovered, and if necessary, dried and solidified to obtain an extract containing plasmalogen. The extract may also be subjected to a concentration treatment to concentrate the plasmalogen.

[0026] In the case of the hexane extraction method, the total lipids contained in the extract obtained by the ethanol extraction method are extracted with a mixed solvent of n-hexane and a water-soluble ketone solvent to separate them into an insoluble portion and a soluble portion. The water-soluble ketone solvent is, for example, acetone, methyl ethyl ketone, or a combination thereof, with acetone being preferred. When a mixture of n-hexane and acetone is used as the mixed solvent, the volume ratio is, for example, 4:6 to 6:4, preferably 4.5:5.5 to 5.5:4.5. The amount of mixed solvent used is, for example, 10 to 30 ml per 1 g (dry mass) of total lipids. The soluble portion obtained by extraction with the mixed solvent is dried and then extracted with a water-soluble ketone solvent, allowing plasmalogen to be separated and recovered.

[0027] Formula (I) representing the above compound is as follows:

[0028]

[0029] In formula (I), X represents a carbon atom or an oxygen atom. Preferably, X is an oxygen atom. １represents a saturated aliphatic hydrocarbon group which may contain a cyclic structure. The number of carbon atoms in the saturated aliphatic hydrocarbon group is not particularly limited, but is, for example, 10 to 30, 10 to 25, or 10 to 20. The number of carbon atoms constituting the cyclic structure is, for example, 3 to 6. The cyclic structure is formed by the cycloalkylene group R １ The cycloalkyl group may be included in R １ The number of carbon atoms constituting the cyclic structure is preferably 3. The saturated aliphatic hydrocarbon group may be a cyclic structure represented by R １ Preferably, R １ is an alkyl group having 18 carbon atoms.

[0030] R ２ represents a saturated or unsaturated aliphatic hydrocarbon group having 10 to 30 carbon atoms which may have a substituent which is an alkyl group having 1 to 4 carbon atoms or an alkoxy group having 1 to 4 carbon atoms. For example, R ２ is R ２ It is a monovalent group that, when expressed as COOH, represents an ω-3 fatty acid, an ω-6 fatty acid, an ω-7 fatty acid, an ω-9 fatty acid, or an ω-10 fatty acid. Preferably, R ２ is R ２ When COOH is used, it is a monovalent group representing arachidonic acid, a monovalent group representing docosahexaenoic acid, a monovalent group representing oleic acid, a monovalent group representing palmitic acid, a monovalent group representing eicosapentaenoic acid, or a monovalent group representing linoleic acid. ２ is R ２ It is a monovalent group which, when expressed as COOH, represents arachidonic acid, oleic acid, or palmitic acid.

[0031] The carbon atoms of the glycerol skeleton in formula (I) may have a substituent. Examples of the substituent that the carbon atoms of the glycerol skeleton may have include a hydroxy group, a halogen, an aryl group, an alkyl group having 1 to 4 carbon atoms, and an alkoxy group having 1 to 4 carbon atoms. R ３ The oxygen atom bonded to is *, and R ３ may have a substituent It is. ３Examples of the substituent that may be contained in the alkyl group include a hydroxy group, a halogen atom, an aryl group, an alkyl group having 1 to 4 carbon atoms, and an alkoxy group having 1 to 4 carbon atoms.

[0032] The salts of the compound represented by formula (I) and the racemic salts of the compound are not particularly limited as long as they are pharmacologically acceptable salts, and may be either acidic salts or basic salts. Examples of the salts include alkali metal salts such as lithium salt, sodium salt, and potassium salt; alkaline earth metal salts such as magnesium salt and calcium salt; inorganic acid salts such as hydrochloride, hydrobromide, sulfate, nitrate, oxalate, and phosphate; acetate, propionate, hexanoate, cyclopentanepropionate, glycolate, pyruvate, lactate, malonate, succinate, malate, fumarate, tartrate, citrate, benzoate, o-(4-hydroxybenzoyl)benzoate, cinnamate, mandelate, methanesulfonate, ethoxylated ... and organic acid salts such as benzenesulfonate, 1,2-ethanedisulfonate, 2-hydroxyethanesulfonate, benzenesulfonate, p-chlorobenzenesulfonate, 2-naphthalenesulfonate, p-toluenesulfonate, camphorsulfonate, glucoheptanoate, 3-phenylpropionate, trimethyl acetate, tert-butyl acetate, lauryl sulfate, gluconate, glutamate, hydroxynaphthoate, salicylate, stearate, trifluoroacetate (TFA) salt, maleate, and muconate.

[0033] For example, the α-synuclein phagocytosis promoter according to this embodiment includes a compound represented by formula (II), its racemate, or a salt thereof. １ and R ２ respectively represent X and R in the above formula (I). １ and R ２ is the same as

[0034]

[0035] Preferably, the α-synuclein phagocytosis promoter according to this embodiment comprises a compound represented by the following formula (i), its racemate, or a salt thereof:

[0036]

[0037] The compound according to this embodiment can be synthesized by a known method. For example, tert-butyl(2-((tert-butoxy((R)-2-hydroxy-3-(octadecyloxy)propoxy)phosphoryl)oxy)ethyl)carbamate is converted to R ２ COOH is reacted. The resulting product is deprotected by the action of a strong acid such as trifluoroacetic acid, to obtain the compound according to this embodiment. The condensing agent is not particularly limited, and examples that can be used include 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC) hydrochloride, dicyclohexylcarbodiimide (DCC), HATU, HBTU, TATU, TBTU, and diphenylphosphoryl azide (DPPA). A nucleophile such as dimethylaminopyridine (DMAP) may be used to promote the dehydration condensation reaction using the condensing agent.

[0038] The α-synuclein phagocytosis promoter according to this embodiment contains, as an active ingredient, a plasmalogen or a compound represented by formula (I), its racemate, or a salt thereof (hereinafter also referred to as a "compound, etc."), and may also contain other pharmacologically acceptable ingredients. Examples of other pharmacologically acceptable ingredients include excipients, lubricants, binders, disintegrants, solvents, solubilizers, suspending agents, isotonicity agents, buffers, soothing agents, etc. Furthermore, additives such as preservatives, antioxidants, colorants, and sweeteners may be incorporated into the α-synuclein phagocytosis promoter as needed.

[0039] Examples of excipients include lactose, sucrose, D-mannitol, starch, crystalline cellulose, light anhydrous silicic acid, etc. Examples of lubricants include magnesium stearate, calcium stearate, talc, colloidal silica, etc. Examples of binders include crystalline cellulose, sucrose, D-mannitol, dextrin, hydroxypropyl cellulose, hydroxypropylmethylcellulose, polyvinylpyrrolidone, etc. Examples of disintegrants include starch, carboxymethylcellulose, carboxymethylcellulose calcium, croscarmellose sodium, carboxymethylstarch sodium, etc.

[0040] Examples of solvents include water for injection, alcohol, propylene glycol, macrogol, etc. Examples of solubilizing agents include polyethylene glycol, propylene glycol, D-mannitol, benzyl benzoate, ethanol, trisaminomethane, cholesterol, triethanolamine, sodium carbonate, sodium citrate, etc. Examples of suspending agents include surfactants, hydrophilic polymers, etc., such as stearyltriethanolamine, sodium lauryl sulfate, laurylaminopropionic acid, lecithin, benzalkonium chloride, benzethonium chloride, glycerin monostearate, polyvinyl alcohol, polyvinylpyrrolidone, sodium carboxymethylcellulose, methylcellulose, hydroxymethylcellulose, hydroxyethylcellulose, hydroxypropylcellulose, etc.

[0041] Examples of isotonic agents include sodium chloride, glycerin, D-mannitol, etc. Examples of buffers include phosphate, acetate, carbonate, citrate buffer solutions, etc. Examples of soothing agents include benzyl alcohol, etc. Examples of preservatives include paraoxybenzoic acid esters, chlorobutanol, benzyl alcohol, phenethyl alcohol, dehydroacetic acid, sorbic acid, etc. Examples of antioxidants include sulfites, ascorbic acid, etc.

[0042] As shown in the examples below, the α-synuclein phagocytosis promoter according to this embodiment enhances the endocytosis of α-synuclein by microglial cells, thereby promoting the clearance of α-synuclein in the nervous system.

[0043] The α-synuclein phagocytosis promoter according to this embodiment promotes the clearance of α-synuclein and is therefore useful for the prevention and treatment of symptoms or diseases caused by the accumulation of α-synuclein and its aggregates, such as α-synucleinopathies, Parkinson's disease, multiple system atrophy, dementia with Lewy bodies, and pure autonomic neuropathy.

[0044] The α-synuclein phagocytosis promoter is produced by a known method and contains the above-mentioned compound, etc. as an active ingredient. The α-synuclein phagocytosis promoter contains, for example, 0.1 to 99 wt %, 1 to 50 wt %, and preferably 1 to 20 wt % of the compound, etc. as an active ingredient.

[0045] The α-synuclein phagocytosis promoter according to this embodiment is administered to humans or animals other than humans. The animal is preferably a mammal, and more specifically, examples include dogs, cats, cows, pigs, horses, sheep, and deer. The route of administration of the α-synuclein phagocytosis promoter to humans is not particularly limited. The α-synuclein phagocytosis promoter is preferably used as an external preparation, an injection, or an oral preparation.

[0046] The α-synuclein phagocytosis promoter is provided in the form of, for example, a liquid, tablet, granule, fine granule, powder, tablet, capsule, or the like.

[0047] The dosage of the α-synuclein phagocytosis promoter is determined appropriately depending on the age, weight, symptoms, etc. of the human subject. The α-synuclein phagocytosis promoter is administered so that the compound or the like is in an effective amount. The effective amount is the amount of the compound or the like necessary to promote the clearance of α-synuclein.

[0048] As explained in detail above, the α-synuclein phagocytosis promoter according to this embodiment can promote the clearance of α-synuclein.

[0049] In another embodiment, there is provided an oral composition for promoting α-synuclein phagocytosis, which comprises the above-mentioned compound, etc. Specific examples of oral compositions for promoting α-synuclein phagocytosis include supplements, food compositions, foods and beverages, functional foods, and food additives. The α-synuclein phagocytosis promoter may be used as a supplement, or may be added to foods and beverages and functional foods.

[0050] The form of the supplement is not particularly limited and may be any form such as tablets, powders, granules, capsules, sugar-coated tablets, films, lozenges, chewable tablets, solutions, emulsions, suspensions, etc. The supplement may contain any ingredient typically used as a supplement.

[0051] The term "functional food" refers to food or beverages taken for the purpose of maintaining health, and includes foods with health claims, foods with specified health uses, foods with functional claims, foods with nutrient functions, health foods, and nutritional supplements. Functional foods are preferably foods with health claims or foods with nutrient functions. When commercializing a functional food, various additives used in foods, specifically colorants, preservatives, thickening agents, antioxidants, bleaching agents, antibacterial and antifungal agents, acidulants, sweeteners, seasonings, emulsifiers, strengthening agents, manufacturing agents, flavorings, and the like, may be added to the oral composition for promoting α-synuclein phagocytosis.

[0052] Functional foods may be either foods or beverages, and are not particularly limited as long as they can be taken orally. Examples of functional foods include beverages, confectioneries, processed grain products, paste products, dairy products, seasonings, etc. Examples of beverages include nutritional drinks, soft drinks, black tea, green tea, etc. Examples of confectioneries include candy, cookies, tablet candy, chewing gum, jelly, etc. Examples of processed grain products include noodles, bread, cooked rice, biscuits, etc. Examples of paste products include sausages, ham, kamaboko, etc. Examples of dairy products include butter, yogurt, etc.

[0053] The oral composition for promoting α-synuclein phagocytosis may be added to food as a food additive. In this case, the food additive may be in the form of a paste, gel, powder, liquid, suspension, emulsion, granules, or the like, so as to be easily added to food.

[0054] The oral composition for promoting α-synuclein phagocytosis may contain water, vitamins, minerals, organic acids, organic bases, fruit juice, flavors, functional ingredients, food additives, etc., within the range that maintains the effect of promoting α-synuclein clearance. The oral composition for promoting α-synuclein phagocytosis can be produced by known methods, adding ingredients other than the above-mentioned compounds, etc., as necessary.

[0055] The oral composition for promoting α-synuclein phagocytosis may be divided and stored in one or more containers so that the daily intake amount is the above-mentioned intake amount. In this case, preferably, one container contains a one-day amount of the oral composition for promoting α-synuclein phagocytosis.

[0056] The oral composition for promoting α-synuclein phagocytosis is provided in a form that can be distinguished from other products in that it is used to promote the clearance of α-synuclein. For example, at least one of the packaging, instructions, and promotional materials for the product relating to the oral composition for promoting α-synuclein phagocytosis indicates that the composition has the effect of promoting the clearance of α-synuclein.

[0057] When an oral composition for promoting α-synuclein phagocytosis is provided as a food or beverage composition, it may be provided or sold as a food or beverage labeled with the use of promoting α-synuclein phagocytosis (including health uses). "Labeling" includes all acts of informing consumers of the above-mentioned use. Any expression that can recall or infer the above-mentioned use is considered to be "labeling," regardless of the purpose, content, object, or medium of the labeling.

[0058] It is preferable that the "labeling" be done in an expression that allows consumers to directly recognize the intended use. Specifically, this includes acts of transferring, delivering, displaying for the purpose of transferring or delivering, or importing food and beverage products or product packaging that lists the intended use, displaying or distributing advertisements, price lists, or transaction documents that list the intended use, or providing information containing the above-mentioned uses by electromagnetic means (such as the Internet).

[0059] "Labeling" includes labeling as health food, functional food, enteral nutritional food, special dietary food, health functional food, food for specified health uses, food with nutrient functions, food with functional claims, quasi-drugs, etc. Among these, labeling approved under systems related to foods for specified health uses, foods with nutrient functions, or foods with functional claims, or similar systems, is particularly included. Specific examples include labeling as a food for specified health uses, labeling as a conditional food for specified health uses, labeling that indicates an effect on the structure or function of the body, labeling that reduces disease risk, and labeling of functionality based on scientific evidence. More specifically, labeling as a food for specified health uses (especially labeling of health uses) and similar labeling are typical examples.

[0060] In another aspect of the present embodiment, there is provided use of the above-mentioned compound, etc. for the manufacture of an agent for promoting α-synuclein phagocytosis or an oral composition for promoting α-synuclein phagocytosis. In another aspect of the present embodiment, there is provided a method for promoting α-synuclein phagocytosis, comprising the step of administering the above-mentioned compound, etc. to a subject. In another aspect of the present embodiment, there is provided a method for treating a symptom or disease caused by the accumulation of α-synuclein and its aggregates in a subject, or a method for regulating the concentration of α-synuclein and its aggregates in the subject, comprising the step of administering the above-mentioned compound, etc. to a subject having a symptom or disease caused by the accumulation of α-synuclein and its aggregates. In another aspect of the present embodiment, there is provided the above-mentioned compound, etc. for use in promoting α-synuclein phagocytosis.

[0061] The present invention will be explained in more detail with reference to the following examples, but the present invention is not limited to these examples.

[0062] Example 1: Preparation of Plasmalogen and Synthesis of Compound KIT-13 (Preparation of Plasmalogen) Plasmalogen (sPls) consisting solely of ethanolamine type was prepared from scallops (Mizuhopecten yessoensis) by the following method. sPls was extracted from raw scallops using hexane, as described in Japanese Patent No. 7304643. The steps for extracting plasmalogen from scallops are as follows: (1) Cocurase P (manufactured by Mitsubishi Chemical Foods Corporation) and phospholipase A1 (PLA1, manufactured by Mitsubishi Chemical Foods Corporation) were added to the scallops and decomposed for 1 hour. (2) Hexane / isopropanol was added and stirred, and after allowing to stand, the supernatant was filtered by suction. (3) Aqueous sodium sulfate solution was added to the filtrate and mixed thoroughly. (4) The upper layer was evaporated to dryness using a rotary evaporator. (5) 40 ml of acetone cooled to 4°C was added to this and mixed. (6) Separation was carried out using a centrifuge at 3000 rpm for 10 minutes at 4°C. (7) The supernatant was discarded and the precipitate was collected. (8) It was dried overnight in a desiccator.

[0063] Following the specification of Japanese Patent No. 5430566, separation was performed by HPLC using the mobile phase solvent shown in Figure 1A. Figure 1B shows the lipid composition of the fractions extracted in steps (1) to (8) above. As shown in Figure 1C, only ethanolamine-type plasmalogen was separated and designated as sPls.

[0064] (Synthesis of KIT-13) KIT-13 was synthesized as a compound according to formula (I) as follows.

[0065]

[0066] To a dichloromethane solution (3 ml) of compound 1 (tert-butyl(2-((tert-butoxy((R)-2-hydroxy-3-(octadecyloxy)propoxy)phosphoryl)oxy)ethyl)carbamate, 187 mg, 0.30 mmol), EDC hydrochloride (115 mg, 0.60 mmol), DMAP (37 mg, 0.30 mmol), and arachidonic acid (100 mg, 0.33 mmol) were added at room temperature, and the mixture was stirred for 20 hours. The reaction solution was diluted with ethyl acetate and then washed with water. The ethyl acetate layer was dried over anhydrous sodium sulfate and then concentrated under reduced pressure. The residue was purified by column chromatography to give the desired (2R)-1-((tert-butoxy(2-((tert-butoxycarbonyl)amino)ethoxy)phosphoryl)oxy)-3-(octadecyloxy)propan-2-yl(5Z,8Z,11Z,14Z)-icosa-5,8,11,14-tetraenoate as a colorless liquid (260 mg, 95%).

[0067] 1 ​H NMR (500 MHz, CDCl3) δ 5.43-5.31 (m, 8H), 5.16 (quin, J = 4.9 Hz, 1H), 5.16 (br, 1H), 4.21 (m, 1H), 4.14-4.09 (m, 1H), 4.07-4.02 (m, 2H), 3.55 (d, J = 5.3 Hz, 2H), 3.47-3.38 (m, 5.3 Hz), 2.85-2.79 (m, 6H), 2.35 (t, J = 7.6 Hz), 2.12 (q, J = 7.0 Hz, 2H), 2.06 (q, J = 7.1 Hz, 2H), 1.74-1.67 (m, 2H), 1.56-1.54 (m, 2H), 1.502 (s, 9H, one diastereomer), 1.496 (s, 9H, the other diastereomer), 1.39-1.25 (m, 38H), 0.90-0.87 (m, 6H).

[0068]

[0069] To a dichloromethane solution (2 ml) of (2R)-1-((tert-butoxy(2-((tert-butoxycarbonyl)amino)ethoxy)phosphoryl)oxy)-3-(octadecyloxy)propan-2-yl(5Z,8Z,11Z,14Z)-icosa-5,8,11,14-tetraenoate (260 mg, 0.29 mmol), 2 ml of trifluoroacetic acid was added at room temperature, and the mixture was stirred for 3 hours. The reaction mixture was concentrated under reduced pressure to give the desired 2-(hydroxy((R)-2-(((5Z,8Z,11Z,14Z)-icosa-5,8,11,14-tetraenoyl)oxy)-3-(octadedecyloxy)propoxy)phosphoryl)oxy)ethane-1-ammonium 2,2,2-trifluoroacetate (KIT-13) as a colorless liquid (249 mg, quant.).

[0070] KIT-13: 1H NMR (500 MHz, CDCl3) δ 11.31 (br, 1H), 5.41-5.31 (m, 8H), 5.16-5.15 (m, 1H), 4.17 (br, 2H), 4.07-4.00 (m, 2H), 3.54 (d, J = 4.9 Hz, 2H), 3.45-3.39 (m, 2H), 3.27 (br), 2.84-2.78 (m, 6H), 2.35 (t, J = 7.4 Hz, 2H), 2.10 (q. J = 7.1 Hz, 2H), 2.05 (q. J = 7.2 Hz, 2H), 1.69-1.66 (m, 2H), 1.54-1.51 (m, 2H), 1.39-1.25 (m, 38H), 0.90-0.86 (m, 6H).

[0071] Test Example 1 MG6 microglial cells (RCB2403, obtained from Riken Cell Bank) were cultured in DMEM medium (Nissui Pharmaceutical Co., Ltd., 05919) containing 10% FBS (Gibco, lot number 2319479). The positive control group was treated with recombinant α-synuclein (SPR-322, StressMarq Biosciences) at a concentration of 0.5 μM for 16 hours. MG6 microglial cells untreated with recombinant α-synuclein served as a negative control. In the sPls group, MG6 microglial cells were simultaneously treated with recombinant α-synuclein and sPls (5 μg / ml) for 16 hours.

[0072] Subsequently, cells were fixed with 4% paraformaldehyde solution and immunostained using an anti-α-synuclein antibody (Cell Signaling, #23706) and a secondary antibody (TRITC-conjugated anti-rabbit antibody, Invitrogen, #A16101). Endocytosed α-synuclein detected intracellularly was used as a marker for the uptake of reactive α-synuclein fragments in microglial cells. DAPI was used to stain cell nuclei. Images were acquired using a fluorescence microscope (Zeiss, Axioskop 2). The number of α-synuclein-positive cells was quantified using Image-J software. Data were plotted as the mean of three independent experiments (n = 3). P values ​​were calculated by ANOVA with post-hoc correction by the Bonferroni method.

[0073] (Results) Figure 2A shows a fluorescence microscope image. Figure 2B shows the percentage of α-synuclein-positive cells. No α-synuclein droplets were detected in the negative control group. In the positive control group, α-synuclein droplets were detected intracellularly and around the cells. In the sPls group, endocytosed α-synuclein increased in MG6 microglial cells (uptake of harmful α-synuclein). In this test example, plasmalogen was shown to improve α-synuclein endocytosis by microglial cells, preventing the accumulation of α-synuclein, which is highly toxic to neurons. In other words, plasmalogen suppresses the accumulation of excessive α-synuclein in the brain.

[0074] Test Example 2 MG6 microglial cells were cultured on glass coverslips in the same manner as in Test Example 1. The cells were treated with 50 μM of the autophagy inhibitor hydroxychloroquine (HCQ, Sigma, 747-36-4) for 1 hour, followed by 0.5 μM of recombinant α-synuclein for 16 hours (KIT-13 + HCQ group). In the KIT-13 group, cells were cultured for 16 hours in the presence of 5 μg / ml KIT-13 together with 0.5 μM of recombinant α-synuclein. In the control group, cells were cultured for 16 hours in the presence of 0.5 μM of recombinant α-synuclein. Each cell was immunostained and images were acquired in the same manner as in Test Example 1. The number of α-synuclein-positive cells was quantified using Image-J software. Data were plotted as the average of three independent experiments (n = 3). P values ​​were calculated by ANOVA and post-hoc corrected by the Bonferroni method.

[0075] (Results) Figure 3A shows fluorescence microscopy images. Figure 3B shows the percentage of α-synuclein-positive cells. In the KIT-13 group, α-synuclein endocytosis in cells was promoted. In the KIT-13 + HCQ group, KIT-13-dependent α-synuclein endocytosis was reduced. This suggests that KIT-13 enhanced autophagy-induced α-synuclein endocytosis in MG6 microglial cells.

[0076] Test Example 3 MG6 microglial cells were prepared and cultured in the same manner as in Test Example 1. In the control group, cells were treated with 0.5 μM recombinant α-synuclein for 16 hours. In the KIT-13 group, cells were cultured for 16 hours in the presence of 0.5 μM recombinant α-synuclein and 5 μg / ml KIT-13. Then, cells were fixed and immunostained in the same manner as in Test Example 1, and endocytosed α-synuclein was quantified. Data were plotted as the average of three independent experiments (n = 3). P values ​​were calculated by ANOVA and post-hoc corrected by the Bonferroni method.

[0077] (Results) Figure 4A shows a fluorescent microscope image. Figure 4B shows the percentage of α-synuclein-positive cells. KIT-13 enhanced autophagy-mediated endocytosis of α-synuclein in MG6 microglial cells.

[0078] The present invention allows various embodiments and modifications without departing from the broad spirit and scope of the present invention. Furthermore, the above-described embodiments are intended to illustrate the present invention and do not limit the scope of the present invention. That is, the scope of the present invention is defined by the claims, not the embodiments. Various modifications made within the scope of the claims and the meaning of the invention equivalent thereto are considered to be within the scope of the present invention.

[0079] This application is based on Japanese Patent Application No. 2024-032659, filed on March 5, 2024. The entire specification, claims, and drawings of Japanese Patent Application No. 2024-032659 are incorporated herein by reference.

[0080] The present invention is useful in medicines and foods.

Claims

1. An α-synuclein phagocytosis promoter comprising a plasmalogen, or a compound represented by formula (I), its racemate, or a salt thereof. [In formula (I), X represents a carbon atom or an oxygen atom, and R １ represents a saturated aliphatic hydrocarbon group which may contain a cyclic structure, and R ２ represents a saturated or unsaturated aliphatic hydrocarbon group having 10 to 30 carbon atoms which may have a substituent, which is an alkyl group having 1 to 4 carbon atoms or an alkoxy group having 1 to 4 carbon atoms, the carbon atoms of the glycerol skeleton may have a substituent, R ３ The oxygen atom bonded to is *, and R ３ may have a substituent It is.] 2. An oral composition for promoting α-synuclein phagocytosis, comprising a plasmalogen, or a compound represented by formula (I), its racemate, or a salt thereof. [In formula (I), X represents a carbon atom or an oxygen atom, and R １ represents a saturated aliphatic hydrocarbon group which may contain a cyclic structure, and R ２ represents a saturated or unsaturated aliphatic hydrocarbon group having 10 to 30 carbon atoms which may have a substituent, which is an alkyl group having 1 to 4 carbon atoms or an alkoxy group having 1 to 4 carbon atoms, the carbon atoms of the glycerol skeleton may have a substituent, R ３ The oxygen atom bonded to is *, and R ３ may have a substituent It is.] 3. X is an oxygen atom and R １ The agent for promoting α-synuclein phagocytosis according to claim 1 or the oral composition for promoting α-synuclein phagocytosis according to claim 2, wherein is an alkyl group having 18 carbon atoms.

4. R ２ is R ２ The agent for promoting α-synuclein phagocytosis or the oral composition for promoting α-synuclein phagocytosis according to claim 3, wherein COOH is a monovalent group that represents arachidonic acid, oleic acid, or palmitic acid.

5. Use of a plasmalogen, or a compound represented by formula (I), its racemate, or a salt thereof, for the manufacture of an agent for promoting α-synuclein phagocytosis or an oral composition for promoting α-synuclein phagocytosis. [In formula (I), X represents a carbon atom or an oxygen atom, and R １ represents a saturated aliphatic hydrocarbon group which may contain a cyclic structure, and R ２ represents a saturated or unsaturated aliphatic hydrocarbon group having 10 to 30 carbon atoms which may have a substituent, which is an alkyl group having 1 to 4 carbon atoms or an alkoxy group having 1 to 4 carbon atoms, the carbon atoms of the glycerol skeleton may have a substituent, R ３ The oxygen atom bonded to is *, and R ３ may have a substituent It is.]

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