Classifications

• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23L—FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES, NOT OTHERWISE PROVIDED FOR; PREPARATION OR TREATMENT THEREOF
  • A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
  • A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
  • A23L33/125—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives containing carbohydrate syrups; containing sugars; containing sugar alcohols; containing starch hydrolysates
• • 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
  • A61K31/7004—Monosaccharides having only carbon, hydrogen and oxygen atoms
• • 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
  • A61K8/00—Cosmetics or similar toiletry preparations
  • A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
  • A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
  • A61K8/60—Sugars; Derivatives thereof
  • A61K8/604—Alkylpolyglycosides; Derivatives thereof, e.g. esters
• • 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
  • A61P1/18—Drugs for disorders of the alimentary tract or the digestive system for pancreatic disorders, e.g. pancreatic enzymes
• • 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
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P35/00—Antineoplastic agents
• • 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
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
  • A61Q19/00—Preparations for care of the skin
  • A61Q19/08—Anti-ageing preparations

Definitions

• the present invention relates to cell growth retarding agents containing D-allose and/or derivatives thereof and/or mixtures thereof as active ingredients, cosmetics containing the same, and methods for delaying cell growth.
• Non-Patent Documents 1 and 2 Such a cellular phenomenon is called “cell slow cycling”, which is a cell function acquired in the process of evolution to survive in a harsh environment, and it has been confirmed that a similar cell population exists in cancer.
• Cell slow cycling Such a phenomenon of cells in which the cell cycle slowly rotates without stopping, called “cell slow cycling”, is thought to be a latent ability in the cells acquired in the process of evolution of living organisms, but there are limits to this phenomenon. This phenomenon is observed only in the situation of , and it is considered that it cannot normally occur (Non-Patent Document 4).
• Non-Patent Document 5 Non-Patent Document 5
• the purpose of the present invention is to provide a cell growth retardant based on a new mechanism, cosmetics containing the same, and a method for delaying cell growth.
• D-allose or a derivative thereof which is one of the rare sugars, has the effect of extending the period of staying in the G0/1 phase of the cell cycle.
• the present invention has been completed.
• Patent Document 1 It has been reported that the rare sugar D-allose arrests the cell cycle via TXNIP and delays the G2 phase of the cell cycle (Patent Document 1 and Non-Patent Document 5). Certain allitols have been reported to have an anti-obesity effect (Patent Document 2), but all of them can extend the period of staying in the G0/1 phase of the cell cycle, especially temporarily arrest the cell cycle. It is not described that the period of staying in the G0/1 phase of the cell cycle can be extended, and the inventors of the present application made it clear for the first time.
• the present invention includes the following inventions.
• the cell growth retardant according to item 1 or 2 which does not arrest the cell cycle.
• the cell growth retarding agent according to any one of Items 1 to 4 which delays the growth of cancer cells.
• the cell growth retardant according to any one of Items 1 to 5 which delays the growth of pancreatic cancer cells or brain tumor cells.
• a cell growth retardant containing D-allose and/or a derivative thereof and/or a mixture thereof as an active ingredient is administered to a subject in need thereof to prolong the period of staying in the G0/1 phase of the cell cycle.
• a method of slowing cell proliferation in a subject comprising: [10] The method according to item 9, wherein the derivative of D-allose is allitol.
• the present invention it is possible to specifically extend the period of staying in the G0/1 phase of the cell cycle without arresting the cell cycle. It has become possible to reproduce the phenomenon called "slow cycling".
• the present invention which has this action, has a wide range of applications, such as preventing age-related changes in various tissues and organs, suppressing the growth of cancer, and making it possible to use it as an anti-aging agent for the skin (e.g., cosmetics). becomes possible.
• FIG. 8 shows cell cycle ratios (G0/1 phase, S phase, and G2/M phase) in cultured human pancreatic cancer cells PK-1 cultured according to the schedule in FIG. Evaluation of glycolysis by flux analyzer system. Evaluation of cell proliferation ability in cultured human pancreatic cancer cell PK-1. Cell proliferation ability was compared by WST-1 assay 48 hours after administration of medium alone (control), D-mannitol or allitol. Evaluation of apoptosis in cultured human pancreatic cancer cells PK-1. Percentages of apoptotic cells 48 hours after administration of medium alone (control), D-mannitol or allitol were compared by flow cytometric analysis of Annexin V-stained cells. Cell cycle delay from G0/1 phase to S phase.
• Cultivation was performed according to the schedule shown in FIG. 8, except that D-mannitol or allitol was administered instead of D-glucose or D-allose.
• Each graph shows the percentage of cell cycle (G0/1 phase, S phase, and G2/M phase) of cultured human pancreatic cancer cell PK-1.
• Evaluation of glycolysis by flux analyzer system. Results of principal component analysis based on metabolome analysis of substances related to central energy metabolism in human pancreatic cancer cells PK-1 and PANC-1 cultured without administration of sugar (control) and with the addition of D-glucose or D-allose.
• Heat map analysis results based on metabolome analysis of substances related to central energy metabolism in human pancreatic cancer cells PK-1 cultured without administration of sugar (control), with the addition of D-glucose or D-allose.
• Gluconeogenesis in human pancreatic cancer cells PK-1 and PANC-1 cultured with no sugar administration (control), D-glucose or D-allose added, metabolites related to polyol pathway and pentose phosphate pathway (PPP) Results of heat map analysis based on metabolome analysis.
• Substance names in black Substances that were not measured or that did not change with the addition of D-allose.
• the present invention provides a cell growth retardant that contains D-allose and/or a derivative thereof and/or a mixture thereof as an active ingredient and prolongs the period of staying in the G0/1 phase of the cell cycle.
• the present invention may be provided as a pharmaceutical product or as a food product.
• the food of the present invention may be, for example, a food with health claims (for example, a food for specified health use or a food with nutrient claims) or a dietary supplement.
• the present invention administers a cell growth retardant containing D-allose and/or a derivative thereof and/or a mixture thereof as an active ingredient to a subject in need thereof, and Methods of slowing cell proliferation in a subject are provided, comprising increasing the duration of time spent in .
• the present invention newly found that the rare sugar D-allose and/or its derivatives and/or mixtures thereof has a hitherto unknown effect of extending the period of staying in the G0/1 phase of the cell cycle. It was completed by discovering
• cell slow cycling refers to a state in which the cell cycle progresses more slowly than conventional cells without arresting the cell cycle ("cell cycle arrest"), e.g. .
• cell cycle arrest a state in which the cell cycle progresses more slowly than conventional cells without arresting the cell cycle
• cultured in a medium containing D-allose and/or derivatives thereof and/or mixtures thereof The cells undergoing cytotoxicity are specifically prolonged in the cell cycle, particularly in the G0/1 phase of the cell cycle, resulting in a prolonged cell cycle.
• the term "cell cycle” refers to the cycle of events that constitute cell division, including mitosis, cytokinesis and interphase, in eukaryotes. Historically, proliferating somatic cells divided the cell cycle into a mitotic phase (M phase) and an interphase. Along with this phase (S phase), the entire cell cycle is divided into 4 phases by adding M phase to G1 phase, S phase and G2 phase, and these cycles are usually called cell cycle. Therefore, in one aspect, the cell cycle means a cycle in which genetic information doubled in the S phase is equally distributed in the M phase to replicate cells.
• the time required for each phase of one cell cycle can be calculated by a known method. It can be calculated by measuring internal DNA content, or the like. Many cell types that have stopped proliferating leave the cell cycle in the middle of the G1 phase and shift to the G0 phase.
• G0 phase refers to the period during which cells stop progressing through the cell cycle at the G1 phase.
• the arrest of cell cycle progression occurs in the G1 phase, G2 phase, M phase, etc.
• the G0 phase refers to the case of exiting the cell cycle in the G1 phase, and also according to the general definition in this specification.
• a typical example of the G0 phase is when cells differentiate or age and stop growing. Entry and exit between the G0 and G1 phases is thought to occur prior to the checkpoint during the G1 phase.
• CDKs, cyclins, and CDK inhibitors are known to be the main groups of factors that control cell cycle progression.
• CDKs are the catalytic subunits of protein kinases. It can be said that it is a driving device for cell cycle progression.
• cyclins can be said to be activators and CKIs to be inhibitors.
• CDKs and cyclins each constitute a protein family, and the timing of promoting cell cycle progression differs depending on the combination of their subtypes.
• mammalian somatic cells include the cyclin B-Cdc2 (CDK1) complex (Cdc2 kinase) in the M phase, the cyclin D-CDK4 complex and the cyclin E-CDK2 complex in the G1 phase, and the cyclin A-CDK2 complex in It is believed to cause S phase progression.
• CKI is believed to inhibit the progression of G1 and S phases or induce withdrawal to G0 phase.
• the term "period of staying in the G0/1 phase of the cell cycle” refers to the time from when the G1 phase starts (i.e., when the M phase ends) to when the S phase starts (i.e., DNA synthesis starts and/or the period from the time a cell in the G0 phase enters the cell cycle (that is, the cell cycle returns from the G0 phase to the G1 phase) to the time the S phase begins.
• D-allose can specifically prolong the duration of cell cycle G0/1 phase without arresting the cell cycle. It was found that the specific prolongation of the period of stay in the G0/1 phase of the cell cycle delays the subsequent transition to the S phase and further delays the subsequent transition to the G2/M phase.
• the present invention exerts an effect of suppressing glycolytic activity in cells.
• glycolysis refers to a biochemical reaction pathway that exists in a living body (cell). It refers to the metabolic process for converting into a form that is easy for living organisms to use, and refers to the same term as generally used. While it is a representative metabolic system that can occur even in an anaerobic state, it also functions as a part of aerobic respiration by transferring the obtained reducing power and pyruvate to the electron transport system and the citric acid cycle.
• Glycolytic activity can be evaluated by measuring intracellular activity. Glycolytic activity can be assessed, for example, without limitation, by measuring the cellular oxygen consumption rate (OCR value), extracellular acidification rate (ECAR value), or OCR and ECAR values. (See Jing Zhang, and Qing Zhang, Using Seahorse Machine to Measure OCR and ECAR in Cancer Cells Methods Mol Biol. 2019; 1928: 353-363.).
• OCR value cellular oxygen consumption rate
• ECAR value extracellular acidification rate
• OCR and ECAR values See Jing Zhang, and Qing Zhang, Using Seahorse Machine to Measure OCR and ECAR in Cancer Cells Methods Mol Biol. 2019; 1928: 353-363.
• to suppress the activity of the glycolytic system means, for example, that the extracellular acidification rate (ECAR value) is controlled (for example, cells cultured in the absence of D-allose and/or allitol ), and the oxygen consumption rate (OCR value) is significantly reduced compared to a control (e.g., cells cultured in the absence of D-allose and/or allitol) may be maintained at a high level.
• ECAR value extracellular acidification rate
• OCR value oxygen consumption rate
• Intracellular activity can be measured by using, for example, an extracellular flux analyzer XFe24 (for 24well) manufactured by Agilent Technologies (formerly Seahorse Bioscience), which is the main energy metabolism pathway of cells, glycolysis, and aerobic respiration by mitochondria.
• the state of cells can be measured over time in a non-invasive and highly sensitive manner.
• the above device is an example of a device that can measure intracellular activity, and any device or method can be used to measure intracellular activity, so the use of the device is not limited.
• the present invention can exert the effect of prolonging the period of staying in the G0/1 phase of the cell cycle, and thus can delay the growth of cancer cells.
• Cancers to which the present invention can be applied include, for example, leukemia (e.g., acute myelogenous leukemia, chronic myeloid leukemia, acute lymphocytic leukemia, chronic lymphocytic leukemia), malignant lymphoma (Hodgkin's lymphoma, non-Hodgkin's lymphoma (e.g., adult T-cell leukemia, follicular lymphoma, diffuse large B-cell lymphoma)), multiple myeloma, myelodysplastic syndrome, head and neck cancer, gastrointestinal cancer (e.g., esophageal cancer, esophageal adenocarcinoma, gastric cancer, colon cancer, colon cancer, rectal cancer), liver cancer (e.g., hepatocellular carcinoma), gallbladder/cholangio
• the present invention can exert the effect of prolonging the period of staying in the G0/1 phase of the cell cycle, so it may be used to delay cell senescence.
• cosmetics used for aging and protective agents for tissues or organs for transplantation in the future, application to foods and pharmaceuticals that suppress biological aging is also possible.
• D-allose that can be used in the present invention is overwhelmingly small compared to D-glucose (glucose) that exists in large amounts in nature.
• D-glucose D-glucose
• monosaccharides that are the basic units of sugars (there are 34 types of monosaccharides (hexoses) with 6 carbon atoms, 16 types of aldoses, 8 types of ketoses, and 10 types of sugar alcohols), there are a large amount in nature.
• Monosaccharides (aldoses, ketoses) and their derivatives (sugar alcohols) which exist only in trace amounts in nature, are defined as "rare sugars", in contrast to "natural monosaccharides" represented by existing D-glucose (glucose). attached.
• D-psicose and D-allose are currently mass-producible rare sugars.
• D-allose is a D-form of allose classified as an aldose and is a hexose.
• Methods for obtaining "D-allose” include a method of synthesizing from D-psicose using L-rhamnose isomerase and a method of obtaining D-psicose by allowing D-xylose isomerase to act on a D-psicose-containing solution.
• D-allose in the present invention is not limited to those methods, and may be obtained by any method such as isomerization by a chemical treatment method.
• D-psicose which is a raw material for D-allose, is generally produced by treating fructose with an enzyme (epimerase), but is not limited thereto, and is obtained by a production method using a microorganism that produces the enzyme.
• It may be a substance, a substance extracted from a natural substance, a substance contained in a natural substance may be used as it is, or a substance isomerized by a chemical treatment method may be used.
• a method for purifying D-psicose using an enzyme is also known.
• D-allose can also be used in the form of D-allose-containing syrup.
• the D-allose-containing syrup can be obtained by appropriately mixing it with a general syrup (liquid sugar). )), it is a “food” sold at general stores and can be easily obtained.
• a method for obtaining a D-allose-containing syrup for example, is to react a monosaccharide (D-glucose or D-fructose) with an alkali to cause a Robry-Debruyn-Van Eckenstein rearrangement reaction, a retro-aldol reaction, and a subsequent aldol reaction.
• a monosaccharide D-glucose or D-fructose
• the above reaction is called an alkali isomerization reaction
• the resulting syrup containing various monosaccharides can be broadly referred to as a "rare sugar-containing syrup”
• D-glucose and / or D-fructose is used as a raw material and alkali isomerized syrup is exemplified until the D-glucose and/or D-fructose content is 55 to 99% by mass.
• the above-mentioned "rare sugar sweet” is a syrup containing rare sugar obtained by the method disclosed in WO 2010/113785 using isomerized sugar as a raw material, and mainly D-psicose and D - manufactured to contain allose.
• Rare sugars contained in the rare sugar-containing syrup obtained by this method are 0.5 to 17% by mass of D-psicose and 0.2 to 10% by mass of D-allose relative to the total sugar. According to Takahashi et al. (Applied Glycoscience, Vol. 5, No. 1, 44-49 (2015)), D-psicose 5.4 g/100 g, D-sorbose 5.3 g/100 g, D-tagatose 2 0 g/100 g, D-allose 1.4 g/100 g, and D-mannose 4.3 g/100 g.
• Raw materials used in the production of the rare sugar-containing syrup include starch, sugar, isomerized sugar, fructose, and glucose.
• Isomerized sugar is widely regarded as a mixed sugar whose main composition is D-glucose and D-fructose in a specific composition ratio, and is generally obtained by hydrolyzing starch with an enzyme such as amylase or an acid. It also refers to a liquid sugar composed mainly of glucose and fructose obtained by isomerizing a sugar solution mainly composed of glucose with glucose isomerase or alkali.
• fructose fructose liquid sugar those with a fructose content (percentage of fructose in sugar) of less than 50% are called “fructose fructose liquid sugar", those with a content of 50% or more and less than 90% are called “fructose liquid sugar”, and 90% or more.
• high-fructose liquid sugar a product obtained by adding sugar to high-fructose liquid sugar in an amount not exceeding the glucose-fructose liquid sugar is called “sugar mixed fructose-glucose liquid sugar”
• the rare sugar-containing syrup of the present invention is called Any isomerized sugar may be used as a raw material of .
• a rare sugar-containing syrup made from D-fructose contains 5.2% D-psicose, 1.8% D-allose, 15.0% glucose, and 69.3% D-fructose.
• the rare sugar-containing syrup made from isomerized sugar as a raw material contains 3.7% D-psicose, 1.5% D-allose, 45.9% glucose, and 37.7% D-fructose. contains 5.7% D-psicose, 2.7% D-allose, 47.4% glucose, and 32.1% D-fructose. changes.
• D-allose may be separated and purified from these syrups and used, use of the syrup as it is is also conceivable.
• D-allose that can be used in the present invention may be D-allose and/or its derivatives and/or mixtures thereof.
• a compound obtained by changing the molecular structure of a certain starting compound by a chemical reaction is called a derivative of the starting compound.
• derivative of D-allose means a compound obtained by converting the molecular structure of D-allose as a starting compound by a chemical reaction; and a compound similar to D-allose (e.g., D-glucose).
• the "D-allose derivative” that can be used in the present invention may include allitol, which is a reduced product of D-allose.
• allitol (D-allo-hexitol; or (2S,3S,4R,5R)-hexane-1,2,3,4,5,6-hexaol.)
• Allitol is a sugar alcohol having 6 carbon atoms obtained by reducing D-psicose or D-allose.
• Zina is also called “Ryobu”.
• Patent Document 2 is known to be contained in the plant body.
• the allitol that can be applied to the present invention may be one extracted from a plant of the genus Zina, and the existence of a catalyst containing a metal selected from elements of group 8 of the periodic table from D-psicose. Allitol produced by a hydrogenation reaction may also be used, but is not limited to these. Also, allitol applicable to the present invention may be allitol and/or its derivatives and/or mixtures thereof.
• the derivatives of D-allose are sugar alcohols in which the carbonyl group of D-allose is an alcohol group (that is, allitol), uronic acid in which the hydroxymethyl group of D-allose is converted to a carboxyl group, D - aldonic acid in which the formyl group at the 1-position of allose is a carboxyl group, aldanoic acid in which the formyl group at the 1-position of D-allose and the hydroxymethyl group at the end of the main chain are both changed to carboxyl groups, or D-allose may be an amino sugar in which the alcohol group of is substituted with an amino group.
• D-allose may be an amino sugar in which the alcohol group of is substituted with an amino group.
• the derivative of D-allose is such that any hydroxy group of D-allose (e.g., 2-, 3-, 4-, 5- and/or 6-position hydroxy groups) is a hydrogen atom, a halogen atom , amino group, carboxyl group, nitro group, cyano group, lower alkyl group, lower alkoxy group, lower alkanoyl group, lower alkanoyloxy group, lower alkoxycarbonyl group, mono- or di-lower alkyl-substituted amino group, aralkyl group, aryl group or It may be a D-allose derivative substituted with a heteroaryl group.
• any hydroxy group of D-allose e.g., 2-, 3-, 4-, 5- and/or 6-position hydroxy groups
• any hydroxy group of D-allose is a hydrogen atom, a halogen atom , amino group, carboxyl group, nitro group, cyano group, lower alkyl group, lower alkoxy group, lower alkan
• the "derivative of D-allose” is a compound obtained by chemically converting the molecular structure of allitol as a starting compound; , a compound obtained by converting the molecular structure of the starting compound by a chemical reaction, and having the same structure as the compound obtained by converting the molecular structure of allitol by a chemical reaction (“structural analogue of allitol" ) may be included.
• a derivative of D-allose for example, any hydroxy group of allitol (e.g., 1-, 2-, 3-, 4-, 5- and/or 6-position hydroxy groups) is a hydrogen atom, halogen atom, amino group, carboxyl group, nitro group, cyano group, lower alkyl group, lower alkoxy group, lower alkanoyl group, lower alkanoyloxy group, lower alkoxycarbonyl group, mono- or di-lower alkyl-substituted amino group, aralkyl group, aryl or heteroaryl groups.
• any hydroxy group of allitol is a hydrogen atom, halogen atom, amino group, carboxyl group, nitro group, cyano group, lower alkyl group, lower alkoxy group, lower alkanoyl group, lower alkanoyloxy group, lower alkoxycarbonyl group, mono- or di-lower alkyl-substituted amino group, aralkyl group, ary
• Halogen atoms refer to fluorine, chlorine, bromine, and iodine atoms.
• Alkyl moieties in lower alkyl groups and lower alkoxy groups, lower alkoxycarbonyl groups, and mono- or di-lower alkyl-substituted amino groups are linear, branched or cyclic C1-C6 alkyl groups, specific examples being methyl, ethyl , propyl, isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, pentyl, isopentyl, neopentyl, hexyl, isohexyl, cyclopropyl, cyclobutyl, 2-methylcyclopropyl, cyclopropylmethyl, cyclopentyl, cyclohexyl and the like.
• I can.
• Lower alkanoyl groups and lower alkanoyl moieties of lower alkanoyloxy groups are linear, branched or cyclic C1-C7 alkanoyl groups, specific examples being formyl, acetyl, propionyl, butyryl, isobutyryl, valeryl, isovaleryl, pivaloyl , hexanoyl, cyclopropylcarbonyl, cyclobutylcarbonyl, 2-methylcyclopropylcarbonyl, cyclohexylcarbonyl and the like.
• the aralkyl group is a C7-C20 aralkyl group, and specific examples include benzyl, phenethyl, ⁇ -methylbenzyl, benzhydryl, trityl, naphthylmethyl and the like.
• the aryl group is a C6-C14 aryl group, and specific examples include phenyl and naphthyl.
• the heteroaryl group is a C3-C8 heteroaryl group which is a monocyclic, polycyclic or condensed ring containing 1 to 4 of each of the same or different N, O and S atoms, and specific examples as 2-pyridyl, 3-pyridyl, 4-pyridyl, 2-quinonyl, 3-quinonyl, 4-quinonyl, 5-quinonyl, 6-quinonyl, 7-quinonyl, 8-quinonyl, 2-indolyl, 3-indolyl, 4-indolyl, 5-indolyl, 6-indolyl, 7-indolyl, 2-furyl, 3-furyl, 2-thienyl, 3-thienyl, 2-pyrrolidyl, 3-pyrrolidyl, 2-imidazolyl, 4-imidazolyl, 5- Imidazolyl, 2-oxazolyl, 4-oxazolyl, 5-oxazolyl, 2-thiazolyl, 4-
• derivatives of D-allose that can be used in the present invention are, for example, 2-deoxy-D-allose, 5-deoxy-D-allose, 6-deoxy-D-allose, 3-deoxy-D-allose D-allose derivatives such as (3-deoxy-D-glucose) and the like may also be used.
• D-allose and/or its derivatives and/or mixtures thereof that can be used in the present invention are interpreted to include pharmacologically acceptable salts and/or hydrates thereof.
• food means food in general, but in addition to general food including so-called health food, it also includes food with health claims such as food for specified health use and food with nutrient function claims. Supplements (supplements, dietary supplements), feeds, food additives, etc. are also included in the food of the present invention.
• the composition for suppressing the production of cytokines of the present invention contains a food called D-allose as an active ingredient, and includes sweeteners, seasonings, food additives, food materials, food and drink, health food and drink, and pharmaceuticals. - It may be used in the form of a quasi-drug or feed, and in either form, it is possible to suppress the production of cytokines.
• the composition of the present invention can be administered by any administration route.
• the route of administration includes topical administration (dermal, inhalation, enema, eye drops, ear drops, nasal, intravaginal, etc.), enteral administration (oral, tube, transinfusion, etc.), parenteral administration (intravenous, transarterial, percutaneous, intramuscular injection, etc.).
• the dose of D-allose and/or a derivative thereof and/or a mixture thereof that can be applied as the cell growth retardant of the present invention can be appropriately adjusted as long as the dose exhibits the effects of the present invention.
• it may be administered at 1 mg/kg body weight/day to 1000 mg/kg body weight/day, and it is also possible to adjust the dosage appropriately according to age and symptoms.
• 1 mg/kg body weight/day to 1000 mg/kg body weight/day such as 10 mg/kg body weight/day to 800 mg/kg body weight/day, 50 mg/kg body weight/day to 500 mg/kg body weight, which can be ingested by enteral administration /day.
• rare sugar D-allose and/or derivatives thereof and/or mixture thereof in daily diet when rare sugar D-allose and/or derivatives thereof and/or mixture thereof is used as a component of food amount can be safely taken.
• the basis for this is that the rare sugar D-allose is an aldose, and from that aspect it is a highly safe compound that can be administered to humans.
• the cell growth retardant of the present invention may be provided as a composition, and D-allose and/or its derivatives and/or mixtures thereof may be added as general excipients, stabilizers, preservatives, binders, disintegrants. It is formulated by blending appropriate excipients such as tablets, powders, granules, capsules, solutions, syrups, elixirs, or appropriate dosage forms such as oily or aqueous suspensions. may be provided.
• D-allose and/or derivatives thereof and/or mixtures thereof further contain pharmaceutically acceptable additives such as fillers, extenders, binders, disintegrants. , dissolution accelerators, wetting agents, lubricants, etc. can be selected and mixed as necessary to form a formulation.
• solid preparations can be produced by adding excipients, disintegrants, binders, lubricants, etc. to D-allose and/or its derivatives and/or mixtures thereof, mixing them, and compressing and shaping them. can be done. Lactose, starch, mannitol and the like are generally used as excipients.
• As the disintegrant calcium carbonate, carboxymethylcellulose calcium, etc. are generally used. Gum arabic, carboxymethylcellulose, or polyvinylpyrrolidone is used as the binder. Talc, magnesium stearate, and the like are known as lubricants.
• Tablets can be masked or coated with known coatings to make them enteric-coated preparations.
• Ethyl cellulose, polyoxyethylene glycol, or the like can be used as the coating agent.
• the composition of the present invention is a food (for example, medical food, food for specified health use, health supplement, health food, food with nutrient function claims, supplement, It can also be provided as a dietary supplement, herbal tea, etc.).
• D-allose may be administered at doses of 1 mg/kg body weight/day to 1000 mg/kg body weight/day (eg, 50 mg to 50 g/day for a 50 kg adult).
• agent of the present invention When the agent of the present invention is used as cosmetics, lotion, emulsion, foundation, lipstick, lip balm, cleansing cream, massage cream, mask, hand cream, hand powder, body shampoo, body lotion, body cream, bath cosmetics, etc. You may use it as a form.
• ingredients that are usually used in external skin preparations such as cosmetics, pharmaceuticals, and quasi-drugs, such as antioxidants, oils, and UV protection agents, to the extent that they do not impair the effects of the present invention.
• surfactants, thickeners, alcohols, powder components, coloring materials, aqueous components, water, various skin nutrients and the like can be appropriately blended as needed.
• the agent of the present invention can be applied as cosmetics, quasi-drugs, etc. applied to the outer skin, and its dosage form is not limited as long as it can be applied to the skin, and is solution-based, solubilized-based, and emulsified-based. , powder dispersion system, water-oil two-layer system, water-oil-powder three-layer system, ointment, lotion, gel, aerosol, etc., can be applied.
• Subjects to which the present invention can be applied may be animals including humans (humans, mammals such as cows, pigs, dogs and cats, birds such as chickens, etc.), and are not particularly limited.
• D-allose a rare sugar, showed a dose-dependent cell growth inhibitory effect in cultured human brain tumor cell lines, U251 cells and U87 cells, as shown in Fig. 1.
• Cells were cultured with DMEM, 50 mM-D glucose or 50 mM-D allose for 24 hours. The cultured cells were then harvested with trypsin, washed twice with PBS, and fixed with 70% ethanol at -20°C overnight. After washing with PBS, cells were stained with 500 ⁇ l of PI reagent for 30 min at room temperature in the dark, and after washing with PBS flow cytometry was used to monitor emission at 617 nm. The data were analyzed with Kaluza analysis software (Ver.1.2, Beckman Coulter), and the effect on the cell cycle was evaluated by changes in the ratio of cell distribution in each phase of the cell cycle.
• CytoTell (trademark), a fluorescent probe for flow cytometry that has recently been used as a cell proliferation monitoring reagent, was taken up by brain tumor cells for 24 hours. , and 50 mM D-allose were administered and allowed to stand for 96 hours, and the attenuation of fluorescent probe intensity in each cell was evaluated (Fig. 4-1).
• PK-1 cells 25 or 50 mM D-allose or D-glucose was added to PK-1 cells, a cultured human pancreatic cancer cell line, and after culturing for 48 hours, a WST-1 assay was performed to compare the effects on cell proliferation.
• D-allose exhibited a cell growth inhibitory effect as shown in Figure 5 in PK-1 cells, a cultured pancreatic cancer cell line.
• Annexin V staining was performed and assessed for apoptosis using flow cytometry, confirming that 50 mM D-allose, the dose that produced an inhibitory effect on pancreatic cancer PK-1 cells, did not produce apoptosis. (Fig. 6).
• PK-1 cells were serum starved overnight and then fixed with 70% ethanol at ⁇ 20° C. overnight.
• PI propidium iodide
• RNase solution Cell Signaling Technology, Danvers, MA, USA
• D-allose does not stop the cell cycle, but through the action of significantly delaying the transition from the G0/1 phase to the S phase, the overall speed of the cell cycle is delayed, leading to the effect of suppressing proliferation. It was thought that
• Example 3 Analysis using cultured human pancreatic cancer cells>
• D-mannitol one of sugar alcohols, was used as a control.
• PK-1 cells 50 mM allitol or D-mannitol was added to PK-1 cells, a cultured human pancreatic cancer cell line, and after 48 hours of culture, WST-1 assay was performed to compare the effects on cell proliferation. again,
• Allitol like D-allose, showed a cell growth inhibitory effect in PK-1 cells, a cultured pancreatic cancer cell line (Fig. 11).
• Annexin V staining was performed and apoptosis was evaluated using flow cytometry. As a result, it was confirmed that apoptosis did not occur at 50 mM allitol, the dose that produced an inhibitory effect on the proliferation of pancreatic cancer PK-1 cells (Fig. 12).
• Allitol like D-allose, does not arrest the cell cycle, but by significantly delaying the transition from the G0/1 phase to the S phase, slows the overall cell cycle speed and suppresses cell proliferation. It was suggested that it might be effective.
• Example 2 an extracellular flux analyzer XFe24 (for 24well) manufactured by Agilent Technologies (formerly Seahorse Bioscience) was used to measure the oxygen consumption rate (OCR value) and extracellular acidity of cultured human pancreatic cancer PK-1 cells. The conversion rate (ECAR value) was measured, and the effect on glycolysis was also examined.
• OCR value oxygen consumption rate
• ECAR value extracellular acidity of cultured human pancreatic cancer PK-1 cells
• D-allose and allitol significantly delay the transition from the G0/1 phase to the S phase by suppressing glycolysis, which leads to "cell slow cycling", resulting in cancer cell growth. It was thought that it might have an inhibitory effect.
• Example 4 Metabolome analysis using human pancreatic cancer cell lines> No sugar (control) or 50 mM D-glucose or D-allose was added to cultured human pancreatic cancer cell lines PK-1 cells and PANC-1 (5.0 ⁇ 10 6 cells). h, 37° C., 5% CO 2 atmosphere. After that, for each cell, samples were prepared according to the protocol provided by Human Metabolome Technologies Co., Ltd.
• PCA principal component analysis
• heat map analysis FIGS. 16 to 19
• pathway map analysis FIG. 20
• Example 5 Visualization of cell cycle of human pancreatic cancer cell line PK-1 cells to which D-allose was added>
• Human pancreatic cancer cell line PK-1 was spiked with tFucci(CA)5 (Plasmid #153521) purchased from Addgene (Massachusetts, USA) (reference: Ando R., et al., Two new coral fluorescent proteins of distinct colors for sharp visualization of cell-cycle progression. bioRxiv, 2020, https://www.biorxiv.org/content/10.1101/2020.03.30.015156v2) to visualize the cell cycle.
• Cells transfected with tFucci(CA)5 appear red in the G1 phase, green in the S phase, and yellow-orange in the G2/M phase when observed using a fluorescence microscope (FIG. 21). Using these cells, the influence of the presence or absence of the addition of D-allose (50 mM) on the cell cycle of human pancreatic cancer cell line PK-1 cells was observed.

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