Classifications

• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K31/00—Medicinal preparations containing organic active ingredients
  • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
  • A61K31/7028—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages
  • A61K31/7034—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages attached to a carbocyclic compound, e.g. phloridzin
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
  • A61P1/04—Drugs for disorders of the alimentary tract or the digestive system for ulcers, gastritis or reflux esophagitis, e.g. antacids, inhibitors of acid secretion, mucosal protectants
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K31/00—Medicinal preparations containing organic active ingredients
  • A61K31/045—Hydroxy compounds, e.g. alcohols; Salts thereof, e.g. alcoholates
  • A61K31/047—Hydroxy compounds, e.g. alcohols; Salts thereof, e.g. alcoholates having two or more hydroxy groups, e.g. sorbitol
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K31/00—Medicinal preparations containing organic active ingredients
  • A61K31/185—Acids; Anhydrides, halides or salts thereof, e.g. sulfur acids, imidic, hydrazonic or hydroximic acids
  • A61K31/19—Carboxylic acids, e.g. valproic acid
  • A61K31/195—Carboxylic acids, e.g. valproic acid having an amino group
  • A61K31/197—Carboxylic acids, e.g. valproic acid having an amino group the amino and the carboxyl groups being attached to the same acyclic carbon chain, e.g. gamma-aminobutyric acid [GABA], beta-alanine, epsilon-aminocaproic acid or pantothenic acid
  • A61K31/198—Alpha-amino acids, e.g. alanine or edetic acid [EDTA]
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K31/00—Medicinal preparations containing organic active ingredients
  • A61K31/33—Heterocyclic compounds
  • A61K31/335—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin
  • A61K31/35—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having six-membered rings with one oxygen as the only ring hetero atom
  • A61K31/352—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having six-membered rings with one oxygen as the only ring hetero atom condensed with carbocyclic rings, e.g. methantheline 
  • A61K31/353—3,4-Dihydrobenzopyrans, e.g. chroman, catechin
  • A61K31/355—Tocopherols, e.g. vitamin E
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K31/00—Medicinal preparations containing organic active ingredients
  • A61K31/33—Heterocyclic compounds
  • A61K31/335—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin
  • A61K31/365—Lactones
  • A61K31/375—Ascorbic acid, i.e. vitamin C; Salts thereof
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K31/00—Medicinal preparations containing organic active ingredients
  • A61K31/33—Heterocyclic compounds
  • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
  • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
  • A61K31/439—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom the ring forming part of a bridged ring system, e.g. quinuclidine
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K31/00—Medicinal preparations containing organic active ingredients
  • A61K31/66—Phosphorus compounds
  • A61K31/665—Phosphorus compounds having oxygen as a ring hetero atom, e.g. fosfomycin
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K31/00—Medicinal preparations containing organic active ingredients
  • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
  • A61K31/7016—Disaccharides, e.g. lactose, lactulose
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K31/00—Medicinal preparations containing organic active ingredients
  • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
  • A61K31/702—Oligosaccharides, i.e. having three to five saccharide radicals attached to each other by glycosidic linkages
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K31/00—Medicinal preparations containing organic active ingredients
  • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
  • A61K31/7028—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K31/00—Medicinal preparations containing organic active ingredients
  • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
  • A61K31/7028—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages
  • A61K31/7032—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages attached to a polyol, i.e. compounds having two or more free or esterified hydroxy groups, including the hydroxy group involved in the glycosidic linkage, e.g. monoglucosyldiacylglycerides, lactobionic acid, gangliosides
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K31/00—Medicinal preparations containing organic active ingredients
  • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
  • A61K31/7042—Compounds having saccharide radicals and heterocyclic rings
  • A61K31/7048—Compounds having saccharide radicals and heterocyclic rings having oxygen as a ring hetero atom, e.g. leucoglucosan, hesperidin, erythromycin, nystatin, digitoxin or digoxin
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K38/00—Medicinal preparations containing peptides
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
  • A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P25/00—Drugs for disorders of the nervous system
  • A61P25/28—Drugs for disorders of the nervous system for treating neurodegenerative disorders of the central nervous system, e.g. nootropic agents, cognition enhancers, drugs for treating Alzheimer's disease or other forms of dementia
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P3/00—Drugs for disorders of the metabolism
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D471/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, at least one ring being a six-membered ring with one nitrogen atom, not provided for by groups C07D451/00 - C07D463/00
  • C07D471/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, at least one ring being a six-membered ring with one nitrogen atom, not provided for by groups C07D451/00 - C07D463/00 in which the condensed system contains two hetero rings
  • C07D471/04—Ortho-condensed systems

Definitions

• the present invention relates to an autophagy activator and a composition for autophagy activation.
• Autophagy responds to extracellular or intracellular stress and signals such as starvation, growth factor deficiency, and pathogen infections by breaking down aging or damaged intracellular substances and organelles to regenerate energy. It is a mechanism for production and removal of damaging substances, and is important for maintaining the homeostasis of normal cells. From past studies, it has been reported that the intracellular autophagy activity decreases sharply as aging progresses (Non-Patent Document 1). In addition, when autophagy is suppressed, aged mitochondria and accidentally folded proteins are excessively accumulated in the cells, and the oxidative stress in the cells is increased to induce cell death, resulting in cell aging. It will be. Therefore, by decomposing the aged substances and organelles in the cells and activating autophagy that recycles the decomposition products, it is possible to promptly remove unnecessary substances in the cells and increase the homeostasis of the cells. ..
• Non-Patent Documents 2 and 3 Especially in Alzheimer's disease, since the function of autophagy is inhibited, aggregated protein called amyloid ⁇ is accumulated in the living body, and it is said that this is involved in the onset (Non-Patent Document 4).
• SENDA disease SENDA: static encephalopathy of childhood with neurodegeneration in adulthood
• SENDA a neurodegenerative disease associated with iron deposition in the melanoma and paleosphere of the brain and atrophy of the cerebral tract
• clone disease which is an inflammatory bowel disease that causes it, and cancer
• the process of autophagy has been studied in both yeast and mammals, with up to 36 proteins utilized. Among them, the formation of autophagosomes and the differentiation of their contents are controlled by the Atg protein encoded by the autophagy-related gene (ATG), and the Atg12-Atg5 binding system and the LC3-Phosphatidyl Ethanolamine (PE) binding system are used. It can be divided into 6 groups including, each of which acts stepwise in each process.
• ATG autophagy-related gene
• PE LC3-Phosphatidyl Ethanolamine
• Autophagy activators include compounds that increase LC3-related factors that are markers of autophagy activity and activate autophagy, and promote autophagy flux (including fusion of autophagosomes to lysosomes). Compounds have been reported (Patent Documents 1 to 4). Further, hesperidin, which is a kind of polyphenol extracted from citrus fruits, is known to have an action of activating autophagy (Non-Patent Document 7).
• an object of the present invention is to provide an autophagy activator and an autophagy activation composition containing the autophagy activator, which can effectively activate autophagy. ..
• the present invention includes the following aspects.
• the methyl hesperidin is one or more selected from the group consisting of chalcone methyl hesperidin represented by the following general formula (1) and flavanone methyl hesperidin represented by the following general formula (2). , [1].
• R 1 to R 9 are each independently a methyl group or a hydrogen atom. However, at least one of R 1 to R 9 is a methyl group.
• R 11 to R 18 are each independently a methyl group or a hydrogen atom. However, at least one of R 11 to R 18 is a methyl group.
• the methyl hesperidin is one or more selected from the group consisting of chalcone methyl hesperidin represented by the following general formula (3) and flavanone methyl hesperidin represented by the following general formula (4). , [2] The autophagy activator.
• R 20 to R 23 are each independently a methyl group or a hydrogen atom.
• R 24 to R 25 are each independently a methyl group or a hydrogen atom.
• flavanone-form methylhesperidins represented by the general formula (4) selected from the group consisting of flavanone-forms-1 to 4 having a combination of R 24 to R 25 shown in Table 2 below.
• An autophagy activating composition comprising the autophagy activator according to any one of [1] to [10] and a pharmaceutically acceptable carrier.
• the composition for autophagy activation according to [11] wherein the total content of the methyl hesperidin is 0.01 to 2% by mass with respect to the total amount of the composition for autophagy activation.
• the vitamin derivative or a salt thereof is at least one selected from the group consisting of ascorbyl phosphate, fatty acid ester of ascorbyl phosphate, tocopherol phosphate ester, and salts thereof, according to [13].
• Composition for activating autophagy [15] The composition for autophagy activation according to any one of [11] to [14], which further contains an inositol derivative in which a sugar is bound to inositol. [16] The composition for autophagy activation according to [15], wherein the sugar is glucose or an oligosaccharide containing glucose as a constituent unit.
• an autophagy activator and an autophagy activation composition containing the autophagy activator which can effectively activate autophagy.
• the present invention provides an autophagy activator containing methyl hesperidin as an active ingredient.
• autophagy is a mechanism for regenerating energy and removing damaged substances by decomposing old or damaged intracellular substances and organelles.
• the autophagy activator of the present embodiment can promote the expression of the LC3 gene, which is an autophagy marker, and the ATG5 gene and ATG7 gene contained in the autophagosome, and can activate autophagy.
• autophagy can be activated by suppressing the expression of the mTOR gene, which acts as an inhibitor of autophagy.
• the autophagy activator of the present embodiment is not particularly limited as long as it contains methyl hesperidin as an active ingredient.
• hesperidin is preferably methylated and solubilized in water.
• the methyl hesperidin mainly includes a chalcone-type compound represented by the following general formula (1) (chalcone-form methyl hesperidin) and a flavanone-type compound represented by the following general formula (2) (flavanone-form methyl hesperidin). It is known.
• R 1 to R 9 are each independently a methyl group or a hydrogen atom. However, at least one of R 1 to R 9 is a methyl group.
• R 11 to R 18 are each independently a methyl group or a hydrogen atom. However, at least one of R 11 to R 18 is a methyl group.
• the methyl hesperidin used in the autophagy activator of the present embodiment is composed of the chalcone methyl hesperidin represented by the general formula (1) and the flavanone methyl hesperidin represented by the general formula (2). It is preferably one or more selected.
• R 1 to R 9 are independently methyl groups or hydrogen atoms, and at least one of R 1 to R 9 is a methyl group.
• any 1 to 6 is preferably a methyl group, and any 2 to 5 is more preferably a methyl group.
• R 11 to R 18 are independently methyl groups or hydrogen atoms, and at least one of R 11 to R 18 is a methyl group. Of R 11 to R 18 , it is preferable that any 1 to 4 are methyl groups, and it is more preferable that any 1 to 3 are methyl groups.
• the compound represented by the following general formula (3) is preferable as the chalcone methyl hesperidin.
• the compound represented by the following general formula (4) is preferable as the flavanone methyl hesperidin.
• R 20 to R 23 are each independently a methyl group or a hydrogen atom.
• R 24 to R 25 are each independently a methyl group or a hydrogen atom.
• R 20 to R 23 are independently methyl groups or hydrogen atoms.
• the chalcone methyl hesperidin represented by the general formula (3) is at least one selected from the group consisting of chalcones -1 to 3 having a combination of R 20 to R 23 shown in Table 3 below. Is preferable.
• R 24 to R 25 are independently methyl groups or hydrogen atoms.
• the flavanone methyl hesperidin represented by the general formula (4) is one or more selected from the group consisting of flavanones -1 to 4 having a combination of R 24 to R 25 shown in Table 4 below. Is preferable.
• the methyl hesperidin used in the autophagy activator of the present embodiment may be one kind alone or a mixture of two or more kinds.
• the methyl hesperidin is a chalcone methyl hesperidin represented by the general formula (1) or a chalcone methyl hesperidin represented by the general formula (3), and a flavanone methyl hesperidin represented by the general formula (2). It may contain both of the flavanone-form methyl hesperidin represented by the general formula (4), or may contain only one of them.
• the methyl hesperidin may contain a chalcone-form methyl hesperidin represented by the general formula (3) and a flavanone-form methyl hesperidin represented by the general formula (4).
• methyl hesperidin may contain any one or more of the chalcones -1 to 3 and may contain any one or more of the flavanone -1 to 4.
• the autophagy activator of the present embodiment may contain a mixture of chalcone bodies-1 to 3 and flavanone bodies-1 to 3 as methyl hesperidin.
• Methyl hesperidin can be produced by a known method.
• methyl hesperidin for example, hesperidin produced from the peel of citrus fruits is dissolved in an aqueous solution of sodium hydroxide, a corresponding amount of dimethyl sulfate is allowed to act on the alkaline solution, the reaction solution is neutralized with sulfuric acid, and n-butyl alcohol is used. It can be produced by extracting, distilling off the solvent, and then recrystallizing with isopropyl alcohol (Sakiburo, Journal of Japanese Chemistry, (1958) Vol. 79, pp. 733-736; Japanese Patent No. 6312333).
• the method for producing methyl hesperidin is not limited to the above method.
• Methyl hesperidin is a commercially available product (for example, a product distributed as a pharmaceutical additive, a food additive, and a raw material for cosmetics, or "Methyl hesperidin” (Showa Denko KK), “Methyl hesperidin” (Tokyo Chemical Industry Co., Ltd.). , “Hesperidin Methyl Calcon” (Sigma), etc.) can also be purchased and used.
• the autophagy activator of the present embodiment can be used by administering itself to a patient for the purpose of treating neurodegenerative diseases such as Alzheimer's disease, Huntington's disease, and Parkinson's disease.
• the autophagy activator of the present embodiment can also be used by blending with pharmaceuticals and cosmetics for the purpose of activating autophagy. Further, it may be blended with the composition for activating autophagy described later and used.
• the autophagy activator of the present embodiment can effectively activate autophagy by promoting the expression of the LC3 gene.
• the autophagy activator of the present embodiment can effectively activate autophagy by promoting the expression of the ATG5 gene.
• the autophagy activator of the present embodiment can effectively activate autophagy by promoting the expression of the ATG7 gene.
• the autophagy activator of the present embodiment can effectively activate autophagy by suppressing the expression of the mTOR gene. Since the autophagy activator of the present embodiment can effectively activate autophagy, it can be used for the prevention or treatment of Alzheimer's disease.
• Amyloid ⁇ is known to cause a decrease in autophagy in nerve cells.
• amyloid ⁇ is known to induce a decrease in autophagy and cell death called apoptosis of nerve cells.
• the autophagy activator of the present embodiment can promote LC3 gene expression in the presence of amyloid ⁇ .
• the autophagy activator of the present embodiment can promote ATG5 gene expression in the presence of amyloid ⁇ .
• the autophagy activator of the present embodiment can promote ATG7 gene expression in the presence of amyloid ⁇ .
• the autophagy activator of the present embodiment can suppress apoptosis in the presence of amyloid ⁇ .
• the autophagy activator of the present embodiment can promote the expression of at least one gene selected from the group consisting of the LC3 gene, the ATG5 gene, and the ATG7 gene in the presence of amyloid ⁇ , particularly in nerve cells. can.
• the autophagy activator of the present embodiment can suppress apoptosis in the presence of amyloid ⁇ , especially in nerve cells.
• Promoting LC3 gene expression in the presence of amyloid ⁇ means that the autophagy activator of the present embodiment is administered in the presence of amyloid ⁇ , as compared with the case where the autophagy activator is not administered. This means that the expression level of the LC3 gene is increased. The same applies to the ATG5 gene and the ATG7 gene.
• Suppressing apoptosis in the presence of amyloid ⁇ means that by administering the autophagy activator of the present embodiment in the presence of amyloid ⁇ , the apoptosis is compared with the case where the autophagy activator is not administered. Means that is suppressed.
• LC3 microtubule assisted protein 1 light chain 3 alpha: NCBI Gene ID: 84557
• LC3-II is converted to LC3-II, which is attracted to the autophagosome membrane by adding phosphatidylethanolamine upstream of autophagy signal transduction. It binds to the membrane.
• LC3 is used as a marker for autophagosomes. Examples of the base sequence of the human LC3 gene include NM_032514.4 and NM_181509.3 registered in the NCBI Reference Sequence database.
• ATG5 (autophagy processed 5: NCBI Gene ID: 9474) binds to ATG12 and functions as an E1-like activating enzyme in a ubiquitin-like conjugated system.
• Examples of the base sequence of the human ATG5 gene include NM_001286106.1, NM_001286107.1, NM_001286108.1, NM_001286111.1.
• ATG7 (autophagy processed 7: NCBI Gene ID: 10533) functions as an E1-like activating enzyme that activates LC3 and ATG12 in an ATP-dependent manner.
• Examples of the base sequence of the human ATG7 gene include NM_001136031.3, NM_001144912.2, NM_0013492322.2, NM_001349233.2.
• MTOR (mechanistic target of rapamycin kinase: NCBI Gene ID: 2475) is a kind of phosphatidylinositol kinase-related kinase and mediates a cellular response to stress such as DNA damage and nutritional deficiency. mTOR functions as an inhibitor of autophagy. Examples of the base sequence of the human mTOR gene include NM_004958.4 registered in the NCBI Reference Sequence database.
• the autophagy activator of the present embodiment is administered to a patient having a high risk of developing neurodegenerative diseases such as Alzheimer's disease, Huntington's disease, and Parkinson's disease, and prevents neurodegenerative diseases such as Alzheimer's disease, Huntington's disease, and Parkinson's disease. May be used for. Further, the autophagy activator of the present embodiment may be administered to a patient who has developed a neurodegenerative disease such as Alzheimer's disease, Huntington's disease, or Parkinson's disease, and may be used to suppress the progression or worsening of the neurodegenerative disease. good.
• the autophagy activator of the present embodiment can be administered to a patient in the same manner as the composition for autophagy activation described later, and may be administered orally or parenterally. It may be administered intravenously, intraarterially, intramuscularly, intradermally, subcutaneously, intraperitoneally or the like, or may be administered intrarectally as a suppository, or may be administered to the skin as an external preparation for skin.
• composition for activating autophagy of the present embodiment contains the above-mentioned autophagy activator containing methyl hesperidin and a pharmaceutically acceptable carrier.
• composition for activating autophagy of the present embodiment follows a conventional method (for example, the method described in the Japanese Pharmacopoeia), the above-mentioned autophagy activator, a pharmaceutically acceptable carrier, and possibly other components. Can be produced by mixing and formulating.
• the term "pharmaceutically acceptable carrier” means a carrier that does not inhibit the physiological activity of the active ingredient and is not substantially toxic to the subject to be administered.
• does not show substantial toxicity means that the component does not show toxicity to the administration subject at the dose normally used.
• the pharmaceutically acceptable carrier is not particularly limited, and is not particularly limited.
• Polymer / thickening / gelling agent solvent, propellant, antioxidant, reducing agent, oxidizing agent, chelating agent, acid, alkali, powder, inorganic salt, water, metal-containing compound, unsaturated monomer , Polyhydric alcohol, polymer additive, wetting agent, thickener, tackifier, oily raw material, liquid matrix, fat-soluble substance, polymer carboxylate and the like.
• compositions for activating autophagy of the present embodiment may be used alone or in combination of two or more.
• the other components are not particularly limited, and are not particularly limited, such as preservatives, antibacterial agents, ultraviolet absorbers, whitening agents, vitamins and derivatives other than methyl hesperidin, anti-inflammatory agents, anti-inflammatory agents, hair growth agents, and blood circulation promotion.
• Agents stimulants, hormones, anti-wrinkle agents, anti-aging agents, tightening agents, cooling sensitizers, warming agents, wound healing promoters, stimulants, analgesics, cell activators, plant / animal / microbial extracts, Seed oil, antipruritic agent, keratin exfoliating / dissolving agent, antiperspirant, cooling agent, astringent agent, enzyme, nucleic acid, fragrance, pigment, colorant, dye, pigment, anti-inflammatory analgesic, antifungal agent, antihistamine, hypnotic sedative , Psychiatric stabilizers, antihypertensive agents, antihypertensive diuretics, antibiotics, anesthetics, antibacterial agents, antiepileptic agents, coronary vasodilators, crude drugs, antipruritic agents, keratin softening release agents, UV blocking agents, bactericides, Examples thereof include antioxidants, pH adjusters, additives, metal anesthetics and the like.
• Specific examples of these components include those described in International Publication No. 2016/076310. Further, specific examples of plant / animal / microbial extracts include lapsana comnis flowers / leaves / stems, tea leaves and the like. Specific examples of seed oil include Moringa oleifera seed oil. Specific examples of fragrances include perillaldehyde. As for the other components, one type may be used alone, or two or more types may be used in combination.
• the composition for activating autophagy of the present embodiment can contain the above-mentioned autophagy activator in a therapeutically effective amount.
• “Therapeutically effective amount” means the amount of drug effective for the treatment or prevention of a patient's disease.
• the therapeutically effective amount may vary depending on the condition, age, sex, body weight, etc. of the disease to be administered.
• the therapeutically effective amount of the above autophagy activator may be an amount in which methyl hesperidin can activate autophagy.
• the therapeutically effective amount of the autophagy activator may be an amount in which methyl hesperidin can promote the expression of at least one gene selected from the group consisting of the LC3 gene, the ATG5 gene, and the ATG7 gene. ..
• the therapeutically effective amount of the autophagy activator may be an amount in which methyl hesperidin can suppress the expression of the mTOR gene.
• the therapeutically effective amount of the autophagy activator may be an amount in which methyl hesperidin can suppress apoptosis in the presence of amyloid ⁇ .
• the therapeutically effective amount (total content of methyl hesperidin) of the autophagy activator in the autophagy activating composition of the present embodiment is, for example, 0.01 to the total amount of the autophagy activating composition. It may be 2% by mass, for example, 0.05 to 1.5% by mass, or 0.1 to 1.0% by mass, for example.
• the total content of methyl hesperidin means the content of the compound when one kind of methyl hesperidin is used alone, and when two or more kinds of methyl hesperidin are used in combination, these compounds are used. Means total content.
• composition for autophagy activation of the present embodiment may contain other autophagy activating components in addition to the autophagy activating agent.
• other autophagy-activating components include at least one vitamin derivative selected from the group consisting of vitamin C derivatives and vitamin E derivatives or a salt thereof, and an inositol derivative in which a sugar is bound to inositol.
• composition for activating autophagy of the present embodiment preferably contains a vitamin C derivative or a salt thereof in addition to the autophagy activator.
• a vitamin C derivative or a salt thereof By containing a vitamin C derivative or a salt thereof, the activation action of autophagy is further improved.
• vitamin C derivatives examples include ascorbic acid derivatives in which at least one hydroxyl group of ascorbic acid is derivatized.
• ascorbic acid derivative more specifically, ascorbyl phosphate (also referred to as ascorbic acid phosphate ester) in which any of the hydroxyl groups of ascorbic acid is phosphorically esterified; any of the hydroxyl groups of ascorbic acid is a phosphoric acid ester.
• Ascorbyl phosphate fatty acid ester obtained by esterifying other hydroxyl groups with fatty acid; ethyl ascorbic acid ethoxylated from any of the hydroxyl groups of ascorbic acid; ascorbic acid glucoside obtained by glucosidizing any of the hydroxyl groups of ascorbic acid.
• Ascorbic acid acylated by acylating one of the hydroxyl groups of ascorbic acid Ascorbyl acylated phosphate obtained by acylating one of the hydroxyl groups of ascorbic acid and phosphorylating the other hydroxyl group; Glyceryl ascorbic acid in which either is replaced with glycerin; a phosphate diester of ascorbic acid and tocopherol (specifically, dl- ⁇ -tocopherol 2-L) in which ascorbic acid and tocopherol are bound by an ester bond via phosphoric acid, respectively. -Ascorbic acid phosphate diester, etc.) and the like.
• Examples of the salt of the ascorbic acid derivative include a salt of an ascorbic acid derivative and an inorganic base, a salt of an ascorbic acid derivative and an organic base, and the like.
• Examples of the salt with the inorganic base include alkali metal salts such as sodium salt and potassium salt; alkaline earth metal salts such as calcium salt and magnesium salt; aluminum salt; ammonium salt; zinc salt and the like.
• Examples of the salt with an organic base include an alkylammonium salt and a salt with a basic amino acid.
• ascorbic acid derivative or salt thereof examples include (i) ascorbyl phosphate or a salt thereof, (ii) a fatty acid ester of ascorbyl phosphate or a salt thereof, (iii) ethylascorbic acid or a salt thereof, and (iv) ascorbin.
• Acid glucoside or a salt thereof is preferable, and (i) ascorbyl phosphate or a salt thereof, and (ii) a fatty acid ester of ascorbyl phosphate or a salt thereof are more preferable.
• Ascorbic Phosphate is a compound in which a phosphate group is introduced into at least one hydroxyl group of ascorbic acid.
• Ascorbyl phosphate a compound represented by the following chemical formula (5) is preferably mentioned.
• the compound represented by the following chemical formula (5) is an ascorbic acid-2-phosphate ester in which the hydroxyl group at the 2-position of ascorbic acid is protected by a phosphoric acid ester.
• Ascorbyl phosphate contains D-form and L-form stereoisomers and racemic DL-form.
• the ascorbyl phosphate in the present embodiment may be any of these stereoisomers, but from the viewpoint of availability, it is preferably L-form, and specifically, L-ascorbic acid-2- Phosphoric acid esters are preferred.
• salt of ascorbyl phosphate examples include a salt of ascorbyl phosphate and an inorganic base, a salt of ascorbyl phosphate and an organic base, and the like.
• Examples of the salt with the inorganic base include alkali metal salts such as sodium salt and potassium salt; alkaline earth metal salts such as calcium salt and magnesium salt; aluminum salt; ammonium salt; zinc salt and the like.
• Examples of the salt with an organic base include an alkylammonium salt and a salt with a basic amino acid.
• the salt of ascorbyl phosphate is preferably an alkali metal salt or an alkaline earth metal salt, more preferably a sodium salt or a magnesium salt, and even more preferably a magnesium salt.
• the magnesium salt of ascorbyl phosphate is preferable from the viewpoint of high stability and resistance to coloring.
• the ascorbyl phosphate or a salt thereof is preferably a salt of ascorbyl phosphate from the viewpoint of improving stability, and is an alkali metal salt of the compound represented by the above chemical formula (5) or the above chemical formula.
• the alkaline earth metal salt of the compound represented by (5) is more preferable, and the sodium salt of the compound represented by the chemical formula (5) or the magnesium salt of the compound represented by the chemical formula (5) is further preferable. ..
• the magnesium salt of the compound represented by the chemical formula (5) specifically, the magnesium salt of L-ascorbic acid-2-phosphate ester is particularly preferable.
• the sodium salt of the compound represented by the chemical formula (5) specifically, the sodium salt of L-ascorbic acid-2-phosphate ester is particularly preferable.
• composition for activating autophagy of the present embodiment ascorbyl phosphate or a salt thereof may be used alone or in combination of two or more.
• content thereof shall be 0.1 to 15% by mass with respect to the total amount of the composition for autophagy activation. Is preferable, 0.5 to 10% by mass is more preferable, and 1 to 5% by mass is further preferable.
• Ascorbyl phosphate or a salt thereof can be produced by a known production method, for example, the method described in JP-A-2-279690 and JP-A-6-345786.
• a specific method for producing ascorbic phosphate it can be obtained by reacting ascorbic acid with phosphorus oxychloride or the like to phosphorylate it.
• phosphorus is obtained by neutralizing an ascorbyl phosphate solution with a metal oxide such as magnesium oxide or a metal hydroxide such as sodium hydroxide.
• a salt of ascorbyl acid can be obtained.
• a sodium salt of ascorbic acid PS (compound name; L-ascorbic acid-2-phosphate ester) manufactured by Showa Denko Co., Ltd. (also known as L-sodium ascorbic acid-2-phosphate). ), Display name; Na ascorbyl phosphate), Ascorbic acid PM manufactured by Showa Denko Co., Ltd. (Compound name; Magnesium salt of L-ascorbic acid-2-phosphate ester (also referred to as L-ascorbic acid-2-phosphate magnesium) ), Display name; ascorbyl phosphate Mg) and the like.
• the fatty acid ester of ascorbyl phosphate is a compound in which a fatty acid is ester-bonded to at least one hydroxyl group of ascorbyl phosphate.
• the fatty acid is a linear or branched fatty acid having 6 to 22 carbon atoms (that is, a fatty acid having a linear or branched alkyl group bonded to a carboxy group having 5 to 21 carbon atoms).
• fatty acid ester of phosphoric acid ascorbyl examples include compounds represented by the following general formula (6).
• the compound represented by the following general formula (6) has ascorbic acid-2-phosphate-6-fatty acid in which phosphoric acid is ester-bonded to the hydroxyl group at the 2-position of ascorbic acid and fatty acid is ester-bonded to the hydroxyl group at the 6-position. Is.
• Rc 1 is a linear or branched-chain alkyl group having 5 to 21 carbon atoms.
• Rc 1 is a linear or branched-chain alkyl group having 5 to 21 carbon atoms. Specifically, a linear or branched pentyl group, a linear or branched hexyl group, a linear or branched heptyl group, a linear or branched octyl group, a direct chain.
• Chained or branched nonyl group linear or branched decyl group, linear or branched undecyl group, linear or branched dodecyl group, linear or branched chain Tridecyl group, linear or branched tetradecyl group, linear or branched pentadecyl group, linear or branched hexadecyl group, linear or branched heptadecyl group, straight chain Examples thereof include a linear or branched octadecyl group, a linear or branched nonadesyl group, a linear or branched icosyl group, and a linear or branched henicosyl group.
• Rc 1 is preferably a linear or branched alkyl group having 9 to 19 carbon atoms, and is preferably a linear or branched alkyl group having 11 to 17 carbon atoms.
• a branched alkyl group is more preferable, and a linear or branched alkyl group having 13 to 15 carbon atoms is more preferable, and the carbon atom number is 15 from the viewpoint of raw material availability and the like.
• Fatty acid esters of ascorbyl phosphate include D-form and L-form stereoisomers and racemic DL-forms.
• the fatty acid ester of ascorbyl phosphate in the present embodiment may be any of these three steric isomers, but from the viewpoint of availability, it is preferably L-form, and specifically, L-ascorbic acid.
• Fatty acid esters of -2-phosphate esters are preferred.
• the salt of fatty acid ester of ascorbyl phosphate include a salt of a fatty acid ester of ascorbyl phosphate and an inorganic base, a salt of a fatty acid ester of ascorbyl phosphate and a salt of an organic base, and the like. Be done.
• Examples of the salt with the inorganic base include alkali metal salts such as sodium salt and potassium salt; alkaline earth metal salts such as calcium salt and magnesium salt; aluminum salt; ammonium salt; zinc salt and the like.
• Examples of the salt with an organic base include an alkylammonium salt and a salt with a basic amino acid.
• the salt of the fatty acid ester of ascorbyl phosphate is preferably an alkali metal salt or an alkaline earth metal salt, more preferably a sodium salt or a magnesium salt, and even more preferably a sodium salt.
• Sodium salts of fatty acid esters of ascorbyl phosphate are preferred from the standpoint of stability and ease of formulation.
• the fatty acid ester of ascorbyl phosphate or a salt thereof is preferably a salt of the fatty acid ester of ascorbyl phosphate from the viewpoint of stability and ease of blending into a preparation, and is represented by the above general formula (6).
• the alkali metal salt of the compound to be used or the alkaline earth metal salt of the compound represented by the general formula (6) is more preferable, and the sodium salt of the compound represented by the general formula (6) or the general formula is described above.
• the magnesium salt of the compound represented by the formula (6) is more preferable, and the sodium salt of the compound represented by the above general formula (6), specifically, L-ascorbic acid-2-phosphoric acid-6-palmitic acid.
• the sodium salt of is particularly preferred.
• the fatty acid ester of ascorbyl phosphate or a salt thereof may be used alone or in combination of two or more.
• the content thereof is preferably 0.05 to 12% by mass, preferably 0.05 to 5% by mass. %, More preferably 0.1 to 2% by mass.
• the fatty acid ester of ascorbyl phosphate or a salt thereof can be produced by a known production method, for example, the method described in Patent 6265550.
• a specific method for producing a fatty acid ester of ascorbyl phosphate after producing ascorbyl phosphate by the same method as the above-mentioned method for producing ascorbyl phosphate, the ascorbyl phosphate and the fatty acid or an ester thereof are used. It can be obtained by subjecting it to a condensation reaction.
• a fatty acid ester solution of ascorbyl phosphate is mixed with a metal oxide such as magnesium oxide or a metal hydroxide such as sodium hydroxide.
• a salt of a fatty acid ester of ascorbyl phosphate can be obtained.
• a commercially available salt of ascorbyl phosphate fatty acid ester in the composition for activating autophagy of the present embodiment is a commercially available salt of ascorbyl phosphate fatty acid ester in the composition for activating autophagy of the present embodiment.
• Aprecier registered trademark
• Examples thereof include a sodium salt of 2-phosphate-6-palmitic acid (also referred to as a sodium salt of L-6-O-palmitoyle ascorbic acid-2-phosphate ester), a display name; palmitate ascorbyl phosphate 3Na) and the like.
• Ethyl ascorbic acid is a compound in which an ethyl group is introduced into at least one hydroxyl group of ascorbic acid.
• a compound represented by the following chemical formula (7) is preferably mentioned.
• the compound represented by the following chemical formula (7) is 3-O-ethylascorbic acid in which the hydrogen atom of the hydroxyl group at the 3-position of ascorbic acid is substituted with an ethyl group.
• Ethyl ascorbic acid has D-form and L-form stereoisomers and racemic DL-form.
• Ethyl ascorbic acid may be any of these stereoisomers, but from the viewpoint of availability, it is preferably L-form, and specifically, L-3-O-ethylascorbic acid (3). -O-ethyl-L-ascorbic acid) is preferable.
• salt of ethyl ascorbic acid examples include a salt of ethyl ascorbic acid and an inorganic base, a salt of ethyl ascorbic acid and an organic base, and the like.
• Examples of the salt with the inorganic base include alkali metal salts such as sodium salt and potassium salt; alkaline earth metal salts such as calcium salt and magnesium salt; aluminum salt; ammonium salt; zinc salt and the like.
• Examples of the salt with an organic base include an alkylammonium salt and a salt with a basic amino acid.
• ethylascorbic acid or a salt thereof is preferably ethylascorbic acid, and more preferably L-3-O-ethylascorbic acid, from the viewpoint of availability.
• ethylascorbic acid or a salt thereof may be used alone or in combination of two or more.
• the content thereof shall be 0.1 to 15% by mass with respect to the total amount of the composition for autophagy activation. Is preferable, 0.5 to 10% by mass is more preferable, and 1 to 5% by mass is further preferable.
• Ethyl ascorbic acid or a salt thereof can be produced by a known production method.
• a method for producing ethyl ascorbic acid a method of alkylating ascorbic acid in dimethyl sulfoxide (DMSO) with an alkyl halide in the presence of sodium methoxide; It can be manufactured by a method or the like.
• DMSO dimethyl sulfoxide
• a salt of ethyl ascorbic acid ethyl ascorbic acid solution is neutralized with a metal oxide such as magnesium oxide or a metal hydroxide such as sodium hydroxide to ethyl.
• a salt of ascorbic acid can be obtained.
• ethyl ascorbic acid examples include 3-O-ethyl-L-ascorbic acid (labeled name; 3-O-ethylascorbic acid) manufactured by Fuji Film Wako Pure Chemical Industries, Ltd.
• Ascorbic acid glucoside is a compound in which at least one hydroxyl group of ascorbic acid is glucosided.
• the glucosidic bond is preferably an ⁇ -glucoside bond.
• Ascorbic acid glucoside is preferably a compound represented by the following chemical formula (8).
• the compound represented by the following chemical formula (8) is ascorbic acid 2-glucoside in which glucose is bound to the hydroxyl group at the 2-position of ascorbic acid.
• Ascorbic acid has D-form and L-form stereoisomers, and racemic DL-form.
• the ascorbic acid in the ascorbic acid glucoside may be any of these stereoisomers, but from the viewpoint of availability, it is preferably L-form, and the ascorbic acid glucoside is specifically L-.
• Ascorbic acid 2-glucoside is preferred.
• the glucose in the ascorbic acid glucoside may be D-form or L-form, but is preferably D-form from the viewpoint of availability.
• the salt of ascorbic acid glucoside examples include a salt of ascorbic acid glucoside and an inorganic base, a salt of ascorbic acid glucoside and an organic base, and the like.
• Examples of the salt with the inorganic base include alkali metal salts such as sodium salt and potassium salt; alkaline earth metal salts such as calcium salt and magnesium salt; aluminum salt; ammonium salt; zinc salt and the like.
• Examples of the salt with an organic base include an alkylammonium salt and a salt with a basic amino acid.
• ascorbic acid glucoside or a salt thereof is preferably ascorbic acid glucoside, and more preferably L-ascorbic acid 2-glucoside, from the viewpoint of availability.
• composition for activating autophagy of the present embodiment ascorbic acid glucoside or a salt thereof may be used alone or in combination of two or more.
• content thereof shall be 0.1 to 15% by mass with respect to the total amount of the composition for autophagy activation. Is preferable, 0.5 to 10% by mass is more preferable, and 1 to 5% by mass is further preferable.
• Ascorbic acid glucoside or a salt thereof can be produced, for example, by the method described in JP-A-03-139288.
• Ascorbic acid glucoside it can be produced by binding one molecule of glucose to the hydroxyl group at the 2-position of ascorbic acid by an enzymatic reaction.
• Ascorbin is obtained by neutralizing the ascorbic acid glucoside solution with a metal oxide such as magnesium oxide or a metal hydroxide such as sodium hydroxide.
• a salt of acid glucoside can be obtained.
• ascorbic acid glucoside examples include ascorbic acid 2-glucoside (compound name; L-ascorbic acid 2-glucoside, display name; ascorbic acid) manufactured by Hayashibara Co., Ltd.
• the ascorbic acid derivative or a salt thereof may be used alone or in combination of two or more.
• As the ascorbic acid derivative or a salt thereof contained in the composition for activating autophagy of the present embodiment (i) ascorbyl phosphate or a salt thereof or (ii) from the viewpoint of further activating autophagy among the above. It is preferably a fatty acid ester of ascorbyl phosphate or a salt thereof, and more preferably (ii) a fatty acid ester of ascorbyl phosphate or a salt thereof.
• composition for autophagy activation of the present embodiment can further promote the expression of the LC3 gene, the ATG5 gene, and the ATG7 gene by containing an ascorbic acid derivative or a salt thereof in addition to methyl hesperidin.
• the composition for autophagy activation of the present embodiment can further suppress the expression of the mTOR gene by containing an ascorbic acid derivative or a salt thereof in addition to methyl hesperidin.
• the composition for autophagy activation of the present embodiment contains an ascorbic acid derivative or a salt thereof in addition to methyl hesperidin, whereby the LC3 gene and the ATG5 gene in the presence of amyloid ⁇ , especially in nerve cells, And the expression of the ATG7 gene can be further promoted.
• composition for autophagy activation of the present embodiment further suppresses apoptosis in the presence of amyloid ⁇ , especially in nerve cells, by containing an ascorbic acid derivative or a salt thereof in addition to methyl hesperidin. Can be done.
• composition for activating autophagy of the present embodiment preferably contains a vitamin E derivative or a salt thereof in addition to the autophagy activator.
• a vitamin E derivative or a salt thereof By containing the vitamin E derivative or a salt thereof, the activation action of autophagy is further improved.
• vitamin E derivative examples include tocopherol phosphate ester or a salt thereof.
• tocopherol phosphate ester examples include compounds represented by the following general formula (9).
• Rd 1 , Rd 2 and Rd 3 independently represent a hydrogen atom or a methyl group.
• the tocopherol phosphate ester is not particularly limited, and may be any of these tocopherol phosphate esters. Among these, ⁇ -tocopherol phosphate ester and ⁇ -tocopherol phosphate ester are preferable, and ⁇ -tocopherol phosphate ester is more preferable.
• the compound represented by the above general formula (9) has an asymmetric carbon atom at the 2-position of the chromane ring, d-form and l-form stereoisomers and dl-form exist.
• the tocopherol phosphate ester may be any of these stereoisomers, but the dl form is preferable.
• tocopherol phosphate ester dl- ⁇ -tocopherol phosphate ester and dl- ⁇ -tocopherol phosphate ester are preferable, and dl- ⁇ -tocopherol phosphate ester is more preferable.
• the salt of tocopherol phosphate ester is not particularly limited, and examples thereof include a salt with an inorganic base and a salt with an organic base.
• Examples of the salt with the inorganic base include alkali metal salts such as sodium salt and potassium salt; alkaline earth metal salts such as calcium salt and magnesium salt; aluminum salt; ammonium salt; zinc salt and the like.
• Examples of the salt with an organic base include an alkylammonium salt and a salt with a basic amino acid.
• alkali metal salt As the salt of tocopherol phosphate ester, an alkali metal salt is preferable, and a sodium salt is more preferable.
• Alkali metal salts of tocopherol phosphate esters, particularly sodium salts have the advantages of being highly soluble in water and being easy to handle because they are powdery in nature.
• Preferred embodiments of the tocopherol phosphate ester include an alkali metal salt (eg, sodium salt) of the compound represented by the above general formula (9), an alkali metal salt of ⁇ -tocopherol phosphate ester (eg, sodium salt), and ⁇ .
• alkali metal salt of tocopherol phosphate eg, sodium salt
• alkali metal salt of dl- ⁇ -tocopherol phosphate eg, sodium salt
• alkali metal salt of dl- ⁇ -tocopherol phosphate eg, sodium salt
• the alkali metal salts of tocopherol phosphate the sodium salt of ⁇ -tocopherol phosphate and the sodium salt of ⁇ -tocopherol phosphate are preferable, and the sodium salt of ⁇ -tocopherol phosphate is more preferable.
• the sodium salt of dl- ⁇ -tocopherol phosphate is commercially available from Showa Denko under the product name of TPNa (registered trademark) (display name: Na tocopheryl phosphate).
• TPNa registered trademark
• the TPNa is exemplified as a preferable example of the tocopherol phosphate ester.
• composition for activating autophagy of the present embodiment one kind of tocopherol phosphate ester or a salt thereof may be used alone, or two or more kinds may be used in combination.
• the content thereof is 0.01 to 10% by mass with respect to the total amount of the composition for autophagy activation. It is preferably 0.05 to 5% by mass, more preferably 0.1 to 3% by mass.
• the tocopherol phosphate ester or a salt thereof can be produced by a known production method, for example, the method described in JP-A-59-44375, International Publication No. 97/14705 and the like.
• a tocopherol phosphate ester can be obtained by allowing a phosphorylating agent such as phosphorus oxychloride to act on tocopherol dissolved in a solvent and appropriately purifying the tocopherol after completion of the reaction.
• the obtained tocopherol phosphoric acid ester is neutralized with a metal oxide such as magnesium oxide, a metal hydroxide such as sodium hydroxide, or ammonium hydroxide or alkylammonium hydroxide to obtain tocopherol phosphate.
• Ester salts can be obtained.
• composition for autophagy activation of the present embodiment can further promote the expression of the LC3 gene, the ATG5 gene, and the ATG7 gene by containing tocopherol phosphate ester or a salt thereof in addition to methyl hesperidin. ..
• the composition for autophagy activation of the present embodiment can further suppress the expression of the mTOR gene by containing tocopherol phosphate ester or a salt thereof in addition to methyl hesperidin.
• the composition for autophagy activation of the present embodiment contains tocopherol phosphate ester or a salt thereof in addition to methyl hesperidin, whereby the LC3 gene and the ATG5 gene in the presence of amyloid ⁇ are particularly contained in nerve cells.
• composition for autophagy activation of the present embodiment further suppresses apoptosis in the presence of amyloid ⁇ , especially in nerve cells, by containing tocopherol phosphate ester or a salt thereof in addition to methyl hesperidin. be able to.
• the composition for activating autophagy of the present embodiment preferably contains an inositol derivative in which a sugar is bound to inositol, in addition to the autophagy activator.
• the inositol derivative is a compound composed of inositol and a sugar, and specifically, is a compound in which a sugar is bound to at least one hydroxyl group of inositol.
• Inositol is a cyclic hexahydric alcohol represented by C 6 H 6 (OH) 6 .
• inositols There are nine inositols: cis-inositol, epi-inositol, allo-inositol, myo-inositol, muco-inositol, neo-inositol, ciro-inositol (there are D and L bodies), and sillo-inositol.
• steric isomers There are steric isomers.
• myo-inositol having physiological activity is preferable as the inositol constituting the inositol derivative.
• the structural formula of myo-inositol is shown below.
• Examples of the method for producing inositol include a method for extracting from rice bran, a chemical synthesis method, and a fermentation method.
• the sugar constituting the inositol derivative may be a monosaccharide or an oligosaccharide.
• the monosaccharide means a sugar that is not further hydrolyzed, and means a compound that is a component when forming a polysaccharide. It can also be said that a monosaccharide is the smallest unit of a sugar.
• the oligosaccharide is an oligomer of a sugar in which a plurality of monosaccharides are bound by glycosidic bonds.
• Monosaccharides include glucose (dextrose), fructose (fructose), galactose, ribose, xylose, mannitol, sorbitol, xylitol, erythritol, pentaerythritol and the like.
• oligosaccharides specifically, disaccharides such as sucrose (sucrose), lactose (lactose), maltose (malt sugar), isomaltose, trehalose, cellobiose, and martitol; Such as trisaccharides; sucrose such as stakiose; hexasaccharides such as ⁇ -cyclodextrin; seven sugars such as ⁇ -cyclodextrin; octasaccharides such as ⁇ -cyclodextrin.
• the sugar constituting the inositol derivative is preferably glucose or an oligosaccharide containing glucose as a monosaccharide unit.
• the monosaccharide unit means a chemical structure corresponding to a monosaccharide, and can also be said to be a chemical structure derived from a monosaccharide.
• the oligosaccharide containing glucose as a single sugar unit may be an oligosaccharide in which only glucose is bound by a glycosidic bond, or an oligosaccharide in which at least one molecule of glucose and a sugar other than glucose are bound by a glycosidic bond. May be.
• the molecular weight of the oligosaccharide containing glucose as a monosaccharide unit may be, for example, about 300 to 3000.
• the sugar may be bound to any one of the six hydroxyl groups present in the inositol molecule, or may be bound to any two or more.
• one molecule of inositol may be bound to one or more monosaccharides
• one molecule of inositol may be bound to one or more oligosaccharides
• one molecule of inositol may be bound to one or more monosaccharides.
• the sugar and one or more oligosaccharides may be bound.
• the total number of sugars (monosaccharides and / or oligosaccharides) bound to one molecule of inositol is 1 or more in terms of monosaccharide unit, for example, 2 or more, for example, 3 or more. There may be, for example, 4 or more, and for example, 10 or more.
• the conversion to a monosaccharide unit indicates how many monosaccharide units the sugar bound to one molecule of inositol is composed of. When a plurality of sugars are bound to one molecule of inositol, it means the total value of the monosaccharide units of the plurality of sugars.
• monosaccharides such as glucose (dextrose), fructose (fructose), galactose, ribose, xylose, mannitol, sorbitol, xylitol, erythritol, and pentaerythritol are converted into monosaccharide units.
• disaccharides such as sucrose (sucrose), lactose (lactose), maltose (maltose), isomaltose, trehalose, cellobiose, and maltose are converted into single sugar units, it is 2.
• trisaccharides such as raffinose, melezitose, and maltotriose are converted into monosaccharide units, it is 3.
• stachyose and other tetrasaccharides are converted into monosaccharide units, it is 4, and when hexasaccharides such as ⁇ -cyclodextrin are converted into monosaccharide units, it is 6, and ⁇ -cyclodextrin and other seven sugars are converted into monosaccharide units. It is 7 when converted, and 8 when converted into a monosaccharide unit of octasaccharide such as ⁇ -cyclodextrin.
• the inositol derivative preferably uses ⁇ -cyclodextrin as a raw material sugar from the viewpoint of facilitating the acquisition of an inositol derivative having a high degree of purification.
• ⁇ -Cyclodextrin is industrially inexpensive and can be stably supplied.
• the sugar constituting the inositol derivative contains glucose as a constituent unit.
• cheaper starch or the like is used as the sugar as the raw material of the inositol derivative, various sugars are transferred to various places during the synthesis of the inositol derivative, so that the degree of purification of the obtained inositol derivative tends to be unstable. There is.
• the inositol derivative may be in the form of a pharmaceutically acceptable salt.
• pharmaceutically acceptable salt means the form of a salt that does not inhibit the bioactivity of the inositol derivative.
• the pharmaceutically acceptable salt of the inositol derivative is not particularly limited, and is, for example, a salt with an alkali metal (sodium, potassium, etc.); a salt with an alkaline earth metal (magnesium, calcium, etc.); an organic base (pyridine, etc.). , Triethylamine, etc.), salt with amine, etc.
• the inositol derivative may be in the form of a solvate. Further, the inositol derivative may be in the form of a solvate of a salt of the inositol derivative.
• the solvate is not particularly limited, and examples thereof include hydrates and ethanol solvates.
• one kind of inositol derivative may be used alone, or two or more kinds may be used in combination.
• the inositol derivative is preferably a mixture of two or more kinds of inositol derivatives, more preferably a mixture of 2 to 40 kinds of inositol derivatives, and more preferably a mixture of 2 to 30 kinds of inositol derivatives. Is more preferable, and a mixture of 10 to 30 kinds of inositol derivatives is particularly preferable.
• the inositol derivative preferably contains an inositol derivative in which the total amount of sugars bound to one molecule of inositol is 10 or more in terms of monosaccharide unit.
• the inositol derivative is preferably an inositol derivative in which glucose or an oligosaccharide containing glucose as a single sugar unit is bound to inositol, and is preferably a mixture of two or more kinds of the inositol derivatives, and is preferably 2 to 40 kinds. It is more preferably a mixture of the inositol derivatives, further preferably a mixture of 2 to 30 types of the inositol derivatives, and particularly preferably a mixture of 10 to 30 types of the inositol derivatives.
• the inositol derivative contains an inositol derivative in which glucose or an oligosaccharide containing glucose as a monosaccharide unit is bound to inositol, and the total of glucose bound to one molecule of inositol and an oligosaccharide containing glucose as a monosaccharide unit is converted into a monosaccharide unit. It is preferable to contain 10 or more inositol derivatives.
• the content thereof is preferably 0.1 to 2% by mass, preferably 0 to 2% by mass, based on the total amount of the composition for autophagy activation. .2 to 1.5% by mass is more preferable, and 0.5 to 1.5% by mass is further preferable.
• the method for synthesizing the inositol derivative is not particularly limited, and it can be appropriately synthesized by a conventionally known method.
• inositol and cyclodextrin which is one of oligosaccharides, may be reacted in the presence of cyclodextrin glucanotransferase to synthesize an inositol derivative (see, for example, Japanese Patent Application Laid-Open No. 63-196596). ..
• an inositol derivative may be synthesized by a method of obtaining a glucosyl compound by using a glucosyl phosphite ester as a sugar donor (see, for example, JP-A-6-298783).
• the composition for autophagy activation of the present embodiment can further promote the expression of the LC3 gene, the ATG5 gene, and the ATG7 gene by containing an inositol derivative in addition to methyl hesperidin.
• the composition for autophagy activation of the present embodiment can further suppress the expression of the mTOR gene by containing an inositol derivative in addition to methyl hesperidin.
• the composition for autophagy activation of the present embodiment contains an inositol derivative in addition to methyl hesperidin, so that the LC3 gene, ATG5 gene, and ATG7 gene in the presence of amyloid ⁇ are particularly contained in nerve cells. Expression can be further promoted.
• the composition for autophagy activation of the present embodiment can further suppress apoptosis in the presence of amyloid ⁇ , particularly in nerve cells, by containing an inositol derivative in addition to methyl hesperidin.
• composition for activating autophagy of the present embodiment may be a pharmaceutical composition or a cosmetic.
• the present invention provides a pharmaceutical composition for autophagy activation, which comprises the above-mentioned autophagy activator and a pharmaceutically acceptable carrier.
• the pharmaceutically acceptable carrier is not particularly limited, and a carrier generally used for pharmaceutical products can be used in addition to those listed above.
• a carrier generally used for pharmaceutical products can be used in addition to those listed above.
• the Japanese Pharmacopoeia the Pharmaceutical Standards outside the Japanese Pharmacopoeia, the Pharmaceutical Additives Standard 2013 (Yakuji Nippo, 2013), the Pharmaceutical Additives Dictionary 2016 (edited by the Japan Pharmaceutical Additives Association, Yakuji Nippo, 2016), Handbook of General raw materials described in Pharmaceutical Excipients, 7th edition (Pharmaceutical Press, 2012) and the like can be used.
• the pharmaceutically acceptable carrier one type may be used alone, or two or more types may be used in combination.
• the pharmaceutical composition of the present embodiment may contain other components in addition to the autophagy activator and a pharmaceutically acceptable carrier.
• the other ingredients are not particularly limited, and general pharmaceutical additives can be used.
• an active ingredient other than the above-mentioned autophagy activator can also be used.
• pharmaceutical additives and active ingredients as other ingredients include, for example, the Japanese Pharmacopoeia, the Pharmaceutical Standards outside the Japanese Pharmacopoeia, the Pharmaceutical Additive Standard 2013 (Yakuji Nippo Co., Ltd., 2013), and the addition of pharmaceuticals.
• the dosage form of the pharmaceutical composition of the present embodiment is not particularly limited, and can be a dosage form generally used as a pharmaceutical preparation.
• dosage forms for oral administration such as tablets, coated tablets, pills, powders, granules, capsules, liquids, suspensions, emulsions; and parenteral such as injections, suppositories, external preparations for skin, etc. Examples thereof include a dosage form to be administered.
• Pharmaceutical compositions of these dosage forms can be formulated according to a conventional method (for example, the method described in the Japanese Pharmacopoeia).
• the method for administering the pharmaceutical composition of the present embodiment is not particularly limited, and the pharmaceutical composition can be administered by a method generally used as a method for administering the drug.
• it may be orally administered as a tablet, a coated tablet, a pill, a powder, a granule, a capsule, a liquid, a suspending agent, an emulsion, etc.
• It may be mixed with a general infusion solution such as, and administered intravenously, intraarterial, intramuscularly, intradermally, subcutaneously, intraperitoneally, etc. It may be administered to the skin.
• the dose of the pharmaceutical composition of the present embodiment can be a therapeutically effective amount.
• the therapeutically effective amount may be appropriately determined depending on the patient's symptoms, body weight, age, sex, etc., the dosage form of the pharmaceutical composition, the administration method, and the like.
• the dose of the pharmaceutical composition of the present embodiment is 0.01 to 500 mg per administration unit form as the total content of methyl hesperidin in the case of oral administration, and the total content of methyl hesperidin in the case of injection.
• the amount is 0.02 to 250 mg per unit form of administration
• the total content of methyl hesperidin in the case of suppositories is 0.01 to 500 mg per unit form of administration
• the total content of methyl hesperidin in the case of external preparations for skin examples thereof include 0.01 to 500 mg per administration unit form.
• the administration interval of the pharmaceutical composition of the present embodiment may be appropriately determined depending on the patient's symptoms, body weight, age, sex, etc., the dosage form of the pharmaceutical composition, the administration method, and the like. For example, it may be once a day or about 2 to 3 times.
• the pharmaceutical composition of the present embodiment can be used for the treatment or prevention of diseases caused by a decrease in autophagy activity.
• diseases include neurodegenerative diseases such as Alzheimer's disease, Huntington's disease, Parkinson's disease and SENDA's disease; inflammatory bowel diseases such as Crohn's disease; and cancer.
• the pharmaceutical composition of the present embodiment is administered to a neurodegenerative disease such as Alzheimer's disease, Huntington's disease, Parkinson's disease, SENDA's disease; inflammatory bowel disease such as Crohn's disease; , Inflammatory bowel disease, or can be used to control the progression of cancer. Further, the pharmaceutical composition of the present embodiment is administered to a neurodegenerative disease such as Alzheimer's disease, Huntington's disease, Parkinson's disease, SENDA's disease; an inflammatory bowel disease such as Crohn's disease; or a cancer patient to cause a neurodegenerative disease. , Inflammatory bowel disease, or can be used to treat cancer. In addition, the pharmaceutical composition of the present embodiment can be used for treating a disease caused by amyloid ⁇ .
• the pharmaceutical composition of the present embodiment can be used for treating a disease caused by a decrease in the expression level of the LC3 gene, the ATG5 gene, or the ATG7 gene.
• the pharmaceutical composition of the present embodiment can be used for treating a disease caused by an increase in the expression level of mTOR.
• the pharmaceutical composition of the present embodiment can be suitably used for treating Alzheimer's disease in particular.
• the pharmaceutical composition of the present embodiment can also be administered to patients at high risk of developing neurodegenerative diseases such as Alzheimer's disease, Huntington's disease, Parkinson's disease, and SENDA's disease to prevent neurodegenerative diseases. ..
• the pharmaceutical composition of the present embodiment can be administered to a patient at high risk of developing inflammatory bowel disease such as Crohn's disease and used to prevent inflammatory bowel disease.
• the pharmaceutical composition of the present embodiment can be administered to a patient at high risk of developing cancer and used to prevent cancer.
• the present invention provides a cosmetic for autophagy activation, which comprises the autophagy activator described above and a pharmaceutically acceptable carrier.
• the pharmaceutically acceptable carrier is not particularly limited, and a carrier generally used for cosmetics can be used in addition to those listed above.
• a carrier generally used for cosmetics can be used in addition to those listed above.
• commentary on the second edition of the cosmetic raw material standard (edited by the Japan Official Regulations Association, Yakuji Nippo Co., Ltd., 1984), the cosmetic raw material non-standard ingredient standard (supervised by the Examination Division, Pharmaceutical Affairs Bureau, Ministry of Health and Welfare, Yakuji Nippo Co., Ltd., 1993), cosmetic raw material standard.
• the cosmetic of this embodiment may contain other components in addition to the autophagy activator and the pharmaceutically acceptable carrier.
• the other ingredients are not particularly limited, and general cosmetic additives can be used.
• an active ingredient other than the above-mentioned autophagy activator can also be used.
• cosmetic additives and active ingredients as other ingredients include, for example, commentary on the second edition of the cosmetic raw material standard (edited by the Japan Official Regulations Association, Yakuji Nippo, 1984), non-standard cosmetic raw material ingredients.
• the form of the cosmetic of the present embodiment is not particularly limited, and can be a form generally used as a cosmetic.
• hair cosmetics such as shampoo, rinse and hair conditioner
• basic cosmetics such as wash pigments, cleansing agents, lotions, milky lotions, lotions, creams, gels, sunscreen agents, packs, masks and beauty liquids
• foundations makeup base, make-up cosmetics such as lipsticks, lip gloss, cheeks; body cleansers, body powders, deodorant cosmetics and the like.
• These cosmetics can be manufactured according to a conventional method.
• the dosage form of the cosmetic of the present embodiment is not particularly limited, and is, for example, oil in water (O / W) type, water in oil (W / O) type, W / O / W type, O / W.
• Emulsified type such as / O type, emulsified polymer type, oily, solid, liquid, paste, stick, volatile oil type, powder, jelly, gel, paste, cream, sheet, film , Mist-like, spray-type, aerosol-like, multi-layered, foam-like, flake-like and the like.
• the amount of the cosmetic used in this embodiment is not particularly limited, but can be an amount effective for activating autophagy.
• the amount of the cosmetic used in this embodiment is 0.01 to 500 mg per use as the total content of methyl hesperidin, and may be, for example, 0.15 to 300 mg, for example, 0.15 to 0.15. It may be 200 mg, for example 0.2 to 100 mg.
• the interval of use of the cosmetics of this embodiment is not particularly limited, but may be, for example, once a day or about 2 to 3 times a day.
• the cosmetic of this embodiment can be used to alleviate the symptoms caused by the decrease in autophagy activity.
• it may be used in routine skin care and makeup by subjects at high risk of developing these symptoms in order to prevent the onset of symptoms resulting from decreased autophagy activity.
• the invention provides a method of activating autophagy, comprising the step of administering methyl hesperidin to a subject.
• the present invention provides a method for promoting expression of the LC3 gene, ATG5 gene, or ATG7 gene, which comprises a step of administering methyl hesperidin to a subject.
• the present invention provides a method for suppressing the expression of the mTOR gene, which comprises a step of administering methyl hesperidin to a subject.
• the present invention provides a method for suppressing apoptosis in the presence of amyloid ⁇ , which comprises a step of administering methyl hesperidin to a subject.
• the present invention provides methyl hesperidin for activating autophagy.
• the invention provides methyl hesperidin for promoting expression of the LC3 gene, ATG5 gene, or ATG7 gene.
• the present invention provides methyl hesperidin for suppressing the expression of the mTOR gene.
• the present invention provides methyl hesperidin for suppressing apoptosis in the presence of amyloid ⁇ .
• the invention provides methyl hesperidin for the prevention or treatment of Alzheimer's disease, Huntington's disease, Parkinson's disease, SENDA disease, Crohn's disease, or cancer.
• the present invention provides the use of methyl hesperidin for producing an autophagy activator.
• the invention provides the use of methyl hesperidin to produce an agent for promoting LC3 gene, ATG5 gene, or ATG7 gene expression.
• the invention provides the use of methyl hesperidin to produce an agent that suppresses mTOR gene expression.
• the present invention provides the use of methyl hesperidin for producing an agent for promoting LC3 gene, ATG5 gene, or ATG7 gene expression in the presence of amyloid ⁇ .
• the invention provides the use of methyl hesperidin to produce an inhibitor of apoptosis in the presence of amyloid ⁇ .
• the present invention provides the use of methyl hesperidin for producing a composition for autophagy activation.
• the invention provides the use of methyl hesperidin to produce a composition for promoting expression of the LC3 gene, ATG5 gene, or ATG7 gene.
• the present invention provides the use of methyl hesperidin for producing a composition for suppressing mTOR gene expression.
• the present invention provides the use of methyl hesperidin for producing a composition for promoting expression of the LC3 gene, ATG5 gene, or ATG7 gene in the presence of amyloid ⁇ .
• the present invention provides the use of methyl hesperidin for producing a composition for suppressing apoptosis in the presence of amyloid ⁇ .
• methyl hesperidin may be used in combination with at least one selected from the group consisting of an ascorbic acid derivative or a salt thereof, a tocopherol phosphate ester or a salt thereof, and an inositol derivative in which a sugar is bound to inositol. preferable.
• Methyl hesperidin (product name: methyl hesperidin) sold by Showa Denko KK was used.
• the total content of the chalcone bodies -1 to 3 and the flavanone bodies -1 to 3 is 97.5% by mass or more in the total amount of the composition.
• vitamin C derivatives The following vitamin C derivatives were used in the following examples and formulations.
• APM Magnesium salt of L-ascorbic acid-2-phosphate ester (display name; ascorbyl phosphate Mg, product name; ascorbic acid PM, manufactured by Showa Denko KK)
• APPS Sodium salt of L-ascorbic acid-2-phosphate-6-palmitic acid (display name; ascorbyl palmitate 3Na, product name; APPS, manufactured by Showa Denko)
• vitamin E derivative In the following examples and formulation examples, the following vitamin E derivatives were used.
• ⁇ -TPNa dl- ⁇ -sodium tocopheryl phosphate (display name: Na tocopheryl phosphate, product name; TPNa (registered trademark), manufactured by Showa Denko KK)
• ⁇ -TPNa dl- ⁇ -sodium tocopheryl phosphate (manufactured by Showa Denko KK)
• the inositol derivative A produced by the method described in International Publication No. 2019/045113 was used. Specifically, myo-inositol (manufactured by Tsukino Rice Fine Chemicals) and ⁇ -cyclodextrin (manufactured by Shiomizu Port Refinery) are reacted in the presence of cyclodextrin glucanotransferase (manufactured by Novozyme) to react with myo-inositol.
• Inositol derivative A which is a mixture of inositol derivatives to which glucose or an oligosaccharide having glucose as a monosaccharide unit is bound to glucose, was prepared.
• LC-MS liquid chromatography-mass spectrometry
• Methyl hesperidin DMSO solution Methyl hesperidin was dissolved in DMSO.
• Hesperidin DMSO solution Hesperidin (manufactured by Tokyo Chemical Industry Co., Ltd.) was dissolved in DMSO.
• APM aqueous solution APM was dissolved in purified water.
• APPS aqueous solution APPS was dissolved in purified water.
• ⁇ -TPNa solution ⁇ -TPNa was dissolved in a 0.05% (V / V) ethanol aqueous solution.
• ⁇ -TPNa solution ⁇ -TPNa was dissolved in a 0.05% (V / V) ethanol aqueous solution.
• Aqueous solution of inositol derivative A Inositol derivative A was dissolved in purified water.
• ⁇ Preparation of aging fibroblasts Human normal fibroblasts (NB1RGB; RIKEN BRC Cellbank) were cultured in D-MEM medium (Sigma-Aldrich) supplemented with 10% fetal bovine serum (MP Biomedicals) until confluent. Then, it was treated with 250 ⁇ M hydrogen peroxide solution for 2 hours, and cultured in D-MEM medium supplemented with fresh 10% fetal bovine serum for 24 hours. This treatment with hydrogen peroxide solution and the operation of cell culture were repeated three times, and the obtained fibroblasts were designated as senescent fibroblasts.
• Example 1 ⁇ Evaluation test of gene expression promoting effect ⁇ The prepared aging fibroblasts were prepared at a seeding density of 10,000 cells / cm 2 and cultured for 24 hours in D-MEM medium (manufactured by Sigma-Aldrich) supplemented with 10% fetal bovine serum (manufactured by MP Biomedicals). did. Then, in Example 1, a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10 -3 % (V / V) and the final concentration of DMSO was 0.1% (V / V). ..
• Example 2 a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10-2 % (V / V) and the final concentration of DMSO was 0.1% (V / V).
• Example 3 a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10-2 % (V / V) and the final concentration of DMSO was 0.1% (V / V), and APM was added.
• APM aqueous solution was added to the medium so that the final concentration of APM was 100 ⁇ M.
• Example 4 a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10-2 % (V / V) and the final concentration of DMSO was 0.1% (V / V), and APPS was added. An APPS aqueous solution was added to the medium so that the final concentration of the was 10 ⁇ M.
• Example 5 a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10-2 % (V / V) and the final concentration of DMSO was 0.1% (V / V).
• the ⁇ -TPNa solution was added to the medium so that the final concentration of TPNa was 10 ⁇ M.
• Example 6 a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10-2 % (V / V) and the final concentration of DMSO was 0.1% (V / V), and ⁇ A ⁇ -TPNa solution was added to the medium so that the final concentration of -TPNa was 10 ⁇ M.
• Example 7 a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10-2 % (V / V) and the final concentration of DMSO was 0.1% (V / V), and inositol was added.
• Reference Example 1 the above-mentioned normal human fibroblasts were prepared at a seeding density of 10,000 cells / cm 2 , and 10% fetal bovine serum (manufactured by MP Biomedicals) was added to the D-MEM medium (Sigma-). The cells were cultured in Aldrich for 24 hours, and DMSO alone was added to the medium so that the final concentration of DMSO was 0.1% (V / V). The medium was then cultured for 24 hours at 37 ° C. under 5% CO 2 .
• RNA was extracted from the aged fibroblasts or human normal fibroblasts of each example, and cDNA was synthesized from the obtained RNA.
• cDNA was synthesized from the obtained RNA.
• the expression level of each gene was quantified by quantitative real-time PCR using primers specific for the LC3 gene, ATG5 gene, and ATG7 gene (manufactured by Takara Bio Inc.).
• GAPDH primary; manufactured by Takara Bio Inc.
• the expression level of GAPDH which is a housekeeping gene whose gene expression does not change due to the addition of a compound, was quantified, and the expression level of each gene was standardized based on the value.
• the relative gene expression level was determined when the expression level of each gene in Comparative Example 1 was 1.00. The results are shown in Table 6.
• the senile fibroblasts had the LC3 gene, the ATG5 gene, and the ATG5 gene, respectively. It was confirmed that the expression levels of the ATG7 genes were all reduced. Comparing the aged fibroblasts cultured with the addition of the hesperidin DMSO solution of Comparative Example 2 and the aged fibroblasts cultured with the addition of only the DMSO of Comparative Example 1, the aged fibroblasts of Comparative Example 2 were compared. The expression levels of the LC3 gene, ATG5 gene, and ATG7 gene were lower than those of the aging fibroblasts of Example 1.
• the LC3 gene and the ATG5 gene were compared with the aged fibroblasts cultured with the addition of only DMSO of Comparative Example 1. , And the expression level of the ATG7 gene were both increased, and in particular, the expression level of the LC3 gene was increased.
• additional components APM, APPS, ⁇ -TPNa, ⁇ -TPNa or inositol derivative A
• the expression levels of the LC3 gene, ATG5 gene and ATG7 gene were further increased.
• the expression of each of the above genes was further promoted.
• ⁇ Evaluation of mTOR gene expression inhibitory effect in human aging fibroblasts The aging fibroblasts prepared above were prepared at a seeding density of 10000 cells / cm 2 and in D-MEM medium (manufactured by Sigma-Aldrich) supplemented with 10% fetal bovine serum (manufactured by MP Biomedicals) 24. Cultured for hours. Then, in Example 8, a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10 -3 % (V / V) and the final concentration of DMSO was 0.1% (V / V). ..
• Example 9 a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10-2 % (V / V) and the final concentration of DMSO was 0.1% (V / V).
• Example 10 a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10-2 % (V / V) and the final concentration of DMSO was 0.1% (V / V), and APM was added.
• APM aqueous solution was added to the medium so that the final concentration of APM was 100 ⁇ M.
• Example 11 a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10-2 % (V / V) and the final concentration of DMSO was 0.1% (V / V), and APPS was added. An APPS aqueous solution was added to the medium so that the final concentration of the was 10 ⁇ M.
• Example 12 a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10-2 % (V / V) and the final concentration of DMSO was 0.1% (V / V).
• the ⁇ -TPNa solution was added to the medium so that the final concentration of TPNa was 10 ⁇ M.
• Example 13 a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10-2 % (V / V) and the final concentration of DMSO was 0.1% (V / V), and ⁇ A ⁇ -TPNa solution was added to the medium so that the final concentration of -TPNa was 10 ⁇ M.
• Example 14 a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10-2 % (V / V) and the final concentration of DMSO was 0.1% (V / V), and inositol was added.
• the above-mentioned normal human fibroblasts were prepared at a seeding density of 10,000 cells / cm 2 , and 10% fetal bovine serum (manufactured by MP Biomedicals) was added to the D-MEM medium (Sigma-).
• the cells were cultured in Aldrich for 24 hours, and DMSO alone was added to the medium so that the final concentration of DMSO was 0.1% (V / V).
• the medium was then cultured for 24 hours at 37 ° C. under 5% CO 2 .
• RNA was extracted from aging fibroblasts or human normal fibroblasts of each example using a Nucleospin (registered trademark) RNA kit (manufactured by Takara Bio Inc.), and cDNA was synthesized from the obtained RNA.
• a Nucleospin (registered trademark) RNA kit manufactured by Takara Bio Inc.
• cDNA was synthesized from the obtained RNA.
• the expression level of the mTOR gene was quantified using a primer specific to the mTOR gene (manufactured by Takara Bio Inc.) by quantitative real-time PCR.
• GAPDH primary; manufactured by Takara Bio Inc.
• the expression level of GAPDH which is a housekeeping gene whose expression does not change due to the addition of a compound, was quantified, and the expression level of each gene was standardized based on the value.
• the relative gene expression level was determined when the expression level of the mTOR gene in Comparative Example 3 was 1.00. The results are shown in Table 7.
• the aged fibroblasts cultured with the addition of the autophagy activator of Examples 8 to 14 had an expression level of the mTOR gene as compared with the aged fibroblasts cultured with the addition of only DMSO of Comparative Example 3. Was decreased, and the expression of the mTOR gene was suppressed.
• SH-SY5Y cells were prepared at a seeding density of 10000 cells / cm 2 and placed in D-MEM / Ham's F-12 medium (manufactured by Sigma-Aldrich) supplemented with 10% fetal bovine serum (manufactured by MP Biomedicals). Was cultured for 24 hours. Then, in Example 15, a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10 -3 % (V / V) and the final concentration of DMSO was 0.1% (V / V). ..
• Example 16 a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10-2 % (V / V) and the final concentration of DMSO was 0.1% (V / V).
• Example 17 a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10-2 % (V / V) and the final concentration of DMSO was 0.1% (V / V), and APM was added.
• APM aqueous solution was added to the medium so that the final concentration of APM was 100 ⁇ M.
• Example 18 a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10-2 % (V / V) and the final concentration of DMSO was 0.1% (V / V), and APPS was added. An APPS aqueous solution was added to the medium so that the final concentration of the was 10 ⁇ M.
• Example 19 a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10-2 % (V / V) and the final concentration of DMSO was 0.1% (V / V).
• the ⁇ -TPNa solution was added to the medium so that the final concentration of TPNa was 10 ⁇ M.
• Example 20 a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10-2 % (V / V) and the final concentration of DMSO was 0.1% (V / V), and ⁇ A ⁇ -TPNa solution was added to the medium so that the final concentration of -TPNa was 10 ⁇ M.
• Example 21 a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10-2 % (V / V) and the final concentration of DMSO was 0.1% (V / V), and inositol was added.
• an amyloid ⁇ solution prepared by dissolving amyloid ⁇ (manufactured by Sigma-Aldrich) in a 0.01% (V / V) DMSO aqueous solution was prepared, and the amyloid ⁇ was prepared so that the final concentration of amyloid ⁇ in each medium was 20 ⁇ M.
• ⁇ solution was added to each medium.
• Reference Example 3 only DMSO was added so that the final concentration of DMSO was 0.1% (V / V), and the amyloid ⁇ solution was not added.
• Each medium was then cultured for 48 hours at 37 ° C. under 5% CO 2 .
• RNA was extracted from the SH-SY5Y cells of each example using a Nucleospin (registered trademark) RNA kit (manufactured by Takara Bio Inc.), and cDNA was synthesized from the obtained RNA.
• a Nucleospin (registered trademark) RNA kit manufactured by Takara Bio Inc.
• cDNA was synthesized from the obtained RNA.
• the expression level of each gene was quantified by quantitative real-time PCR using primers specific for the LC3 gene, ATG5 gene, and ATG7 gene (manufactured by Takara Bio Inc.).
• GAPDH GAPDH
• Takara Bio Inc. a housekeeping gene whose expression does not change due to the addition of a compound.
• the expression level of each gene was standardized based on the value.
• the relative gene expression level was determined when the expression level of each gene in Reference Example 3 was 1.00. The results are shown in Table 8.
• the SH-SY5Y cells cultured with the addition of the autophagy activator of Examples 15 to 21 and the amyloid ⁇ solution were compared with the SH-SY5Y cells cultured with the addition of DMSO and the amyloid ⁇ solution of Comparative Example 5. Therefore, the expression levels of the LC3 gene, the ATG5 gene, and the ATG7 gene were all increased. Further, even in the SH-SY5Y cells cultured with the addition of hesperidin having a final concentration of 10-2 % (V / V) in Comparative Example 6, the LC3 gene, the ATG5 gene, and the LC3 gene and the ATG5 gene were compared with those of the SH-SY5Y cells in Comparative Example 5.
• the expression level of the ATG7 gene was increased.
• the SH-SY5Y cells cultured with the addition of 10-2 % (V / V) of methyl hesperidin at the final concentration of Example 16 showed a remarkable increase in the expression level of all the genes.
• additional components APM, APPS, ⁇ -TPNa, ⁇ -TPNa or inositol derivative A.
• the expression levels of the LC3 gene, ATG5 gene and ATG7 gene were further increased. The expression of each of the above genes was further promoted.
• ⁇ Evaluation of apoptosis inhibitory effect caused by amyloid ⁇ in human neuroblastoma The proportion of apoptotic cells in human neuroblastoma (SH-SY5Y; obtained from ATCC) in the presence of an autophagy activator was measured by the following test method, and the apoptosis inhibitory effect of the autophagy activator was evaluated.
• amyloid ⁇ which is known to induce a decrease in autophagy and cell death called apoptosis of nerve cells, was added to each medium for the test.
• SH-SY5Y cells were prepared at a seeding density of 50,000 cells / cm 2 and placed in D-MEM / Ham's F-12 medium (manufactured by Sigma-Aldrich) supplemented with 10% fetal bovine serum (manufactured by MP Biomedicals). Was cultured for 24 hours. Then, in Example 22, a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10 -3 % (V / V) and the final concentration of DMSO was 0.1% (V / V). ..
• Example 23 a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10-2 % (V / V) and the final concentration of DMSO was 0.1% (V / V).
• Example 24 a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10-2 % (V / V) and the final concentration of DMSO was 0.1% (V / V), and APM was added.
• APM aqueous solution was added to the medium so that the final concentration of APM was 100 ⁇ M.
• Example 25 a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10-2 % (V / V) and the final concentration of DMSO was 0.1% (V / V), and APPS was added. An APPS aqueous solution was added to the medium so that the final concentration of the was 10 ⁇ M.
• Example 26 a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10-2 % (V / V) and the final concentration of DMSO was 0.1% (V / V).
• the ⁇ -TPNa solution was added to the medium so that the final concentration of TPNa was 10 ⁇ M.
• Example 27 a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10-2 % (V / V) and the final concentration of DMSO was 0.1% (V / V), and ⁇ A ⁇ -TPNa solution was added to the medium so that the final concentration of -TPNa was 10 ⁇ M.
• Example 28 a methyl hesperidin DMSO solution was added to the medium so that the final concentration of methyl hesperidin was 10-2 % (V / V) and the final concentration of DMSO was 0.1% (V / V), and inositol was added.
• an amyloid ⁇ solution prepared by dissolving amyloid ⁇ (manufactured by Sigma-Aldrich) in a 0.01% (V / V) DMSO aqueous solution was prepared, and the amyloid ⁇ was prepared so that the final concentration of amyloid ⁇ in each medium was 30 ⁇ M.
• ⁇ solution was added to each medium.
• Reference Example 4 only DMSO was added so that the final concentration of DMSO was 0.1% (V / V), and the amyloid ⁇ solution was not added.
• Each medium was then cultured for 48 hours at 37 ° C. under 5% CO 2 .
• each SH-SY5Y cell was washed with phosphate buffer (PBS, manufactured by Wako Pure Chemical Industries, Ltd.), and the number of cells showing strong Hoechst fluorescence like apoptosis due to chromatin aggregation under a fluorescence microscope (Hoechst (+)). ) Cells) were measured.
• PBS phosphate buffer
• the SH-SY5Y cells cultured with the addition of the autophagy activator of Examples 22 to 28 and the amyloid ⁇ solution were compared with the SH-SY5Y cells cultured with the addition of DMSO and the amyloid ⁇ solution of Comparative Example 7. It was confirmed that the proportion of apoptotic cells decreased. Further, even in the SH-SY5Y cells cultured with the addition of hesperidin having a final concentration of 10-2 % (V / V) in Comparative Example 8, the proportion of apoptotic cells was lower than that in the SH-SY5Y cells of Comparative Example 7. Was. However, in the SH-SY5Y cells cultured with the addition of methyl hesperidin having a final concentration of 10-2 % (V / V) in Example 23, the proportion of apoptotic cells was significantly reduced.
• additional components APM, APPS, ⁇ -TPNa, ⁇ -TPNa or inositol derivative A.
• the proportion of apoptotic cells was lower.
• Table 10 shows Formulation Examples 1 to 5 of an external preparation as a composition for activating autophagy.
• an autophagy activator capable of effectively activating autophagy and an autophagy activation composition containing the autophagy activator are provided.

Landscapes

• Health & Medical Sciences (AREA)
• Life Sciences & Earth Sciences (AREA)
• Chemical & Material Sciences (AREA)
• Medicinal Chemistry (AREA)
• Pharmacology & Pharmacy (AREA)
• Animal Behavior & Ethology (AREA)
• General Health & Medical Sciences (AREA)
• Public Health (AREA)
• Veterinary Medicine (AREA)
• Epidemiology (AREA)
• Molecular Biology (AREA)
• Organic Chemistry (AREA)
• Engineering & Computer Science (AREA)
• Bioinformatics & Cheminformatics (AREA)
• Chemical Kinetics & Catalysis (AREA)
• General Chemical & Material Sciences (AREA)
• Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
• Neurosurgery (AREA)
• Neurology (AREA)
• Biomedical Technology (AREA)
• Hospice & Palliative Care (AREA)
• Psychiatry (AREA)
• Proteomics, Peptides & Aminoacids (AREA)
• Immunology (AREA)
• Obesity (AREA)
• Hematology (AREA)
• Diabetes (AREA)
• Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)

Priority Applications (5)

Applications Claiming Priority (2)

Publications (1)

Family

ID=80776133

Family Applications (1)

Country Status (6)

Cited By (1)

Citations (18)

Family Cites Families (7)

• 2021
  • 2021-09-17 WO PCT/JP2021/034285 patent/WO2022059775A1/ja not_active Ceased
  • 2021-09-17 KR KR1020237009525A patent/KR20230054431A/ko active Pending
  • 2021-09-17 JP JP2022550627A patent/JPWO2022059775A1/ja active Pending
  • 2021-09-17 CN CN202180071407.1A patent/CN116322705B/zh active Active
  • 2021-09-17 EP EP21869457.8A patent/EP4215199A4/en not_active Withdrawn
  • 2021-09-17 US US18/026,057 patent/US20230355646A1/en active Pending

Patent Citations (19)

Non-Patent Citations (22)

Also Published As

Similar Documents

Legal Events