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WO2021167310A1 - Composition inhibitrice d'atopie contenant une aldéhyde déshydrogénase - Google Patents

Composition inhibitrice d'atopie contenant une aldéhyde déshydrogénase Download PDF

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WO2021167310A1
WO2021167310A1 PCT/KR2021/001935 KR2021001935W WO2021167310A1 WO 2021167310 A1 WO2021167310 A1 WO 2021167310A1 KR 2021001935 W KR2021001935 W KR 2021001935W WO 2021167310 A1 WO2021167310 A1 WO 2021167310A1
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present
aldehyde dehydrogenase
aldehyde
aldh
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Korean (ko)
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권흥택
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Pico Entech Co Ltd
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Pico Entech Co Ltd
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    • 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
    • A23L33/14—Yeasts or derivatives thereof
    • A23L33/145—Extracts
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/06—Fungi, e.g. yeasts
    • A61K36/062—Ascomycota
    • A61K36/064—Saccharomycetales, e.g. baker's yeast
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/43—Enzymes; Proenzymes; Derivatives thereof
    • A61K38/44—Oxidoreductases (1)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P11/00—Drugs for disorders of the respiratory system
    • A61P11/06—Antiasthmatics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P17/00—Drugs for dermatological disorders
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P17/00—Drugs for dermatological disorders
    • A61P17/06—Antipsoriatics
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/14—Fungi; Culture media therefor
    • C12N1/16—Yeasts; Culture media therefor
    • C12N1/18—Baker's yeast; Brewer's yeast

Definitions

  • the present invention relates to an atopic inhibitor containing aldehyde dehydrogenase. More particularly, the present invention relates to a composition for treating atopy, allergy or psoriasis, containing an aldehyde dehydrogenase that reduces IgE.
  • the present invention relates to a treatment for atopic dermatitis, treatment for psoriasis, and treatment for allergies, containing aldehyde dehydrogenases and their variants as active ingredients, including fusion proteins or recombinant proteins including an action site involved in aldehyde dehydrogenation. .
  • Atopic dermatitis is a chronic, recurrent inflammatory skin disease that causes itching, an excessive allergic reaction that can occur without antigen contact, and an abnormal inflammatory reaction.
  • Atopic dermatitis is a skin disease that reacts sensitively to food or inhalable substances, and is caused by environmental factors, genetic predisposition, immunological abnormalities, and abnormal skin barrier.
  • Environmental factors of atopic dermatosis include environmental pollution such as soot, an increase in the use of food additives, the use of carpets, beds, and sofas, and an increase in substances that cause allergies such as house dust mites. It seems that an allergic reaction to an antigen that is easy to invade due to a decrease in skin defense function is involved, and environmental factors such as house dust mites and mold are involved as representative antigens.
  • atopic allergy psoriasis, etc. are both hereditary and environmental diseases. These skin diseases are considered to be diseases in which severe pruritus (itching), dry skin, and dermatitis (eczema) are the main symptoms, and type 1 and type 4 allergic reactions are related.
  • Topical steroids and local immunomodulators have been developed for the treatment of atopic dermatitis, and antihistamines are often used to suppress itching. Treatment is also needed to avoid allergens, irritants, and stress that aggravate or cause skin symptoms.
  • interleukins are soluble proteins and are a type of cytokine, which is a substance that transmits signals between cells. Because it is a cytokine produced by lymphocytes, it is also called lymphokine, and because it is a signaling substance between leukocytes, it is called interleukin.
  • interleukins are known to be involved in the regulation of various bioactivity, in particular, the regulation of various immune responses by binding to a receptor on the surface of a target cell.
  • interleukin 4 or interleukin 13 is known to be related to skin diseases such as atopy and allergic psoriasis.
  • Interleukin 4 (hereinafter, abbreviated as IL-4) secreted from T cells is a cytokine of about 20 kD consisting of 129 amino acids, and is an important cytokine that increases B-cell differentiation factors and B-cell proliferation factors, especially IgE. .
  • IL-4 is initially secreted by basophils and additionally secreted by activated T-cells (TH2). It has been reported to contribute to B cell activation, IgE type conversion, and induction of Th2 differentiation.
  • IL-4 production by Th2 cells is also accelerated by IL-4, and the receptor for IL-4 is a 130 kD protein that binds two cysteines, IL-4R ⁇ + ⁇ b and IL-4R ⁇ +IL-13R ⁇ . receptors are present.
  • IL-4 has many biological functions, such as stimulating activated B cells and T cells, and differentiating B cells into plasma cells, and is an important regulator of humoral and adaptive immunity. It is also known that IL-4 induces class switching, which is converted from B progenitor cells to B cells capable of producing IgE and IgG4, as well as promotes the production of MHC class II.
  • IL-4 acts synergistically with IL-13 and is the causative agent of IgE- or eosinophil-mediated inflammatory responses.
  • interleukin 13 is a protein composed of 122 amino acids, and the receptor for IL-13 is CD132 ( ⁇ c). IL-13 is functionally similar to IL-4 and is secreted from Th2 cells, dendritic cells, and mast cells. make it
  • the receptor for IL-13 consists of IL-13R ⁇ and IL-4R ⁇ , the IL-13 receptor also responds to IL-4.
  • the receptor for Il-13 is expressed in human B cells, but not in mouse B cells. IL-13 also differentiates monocytes into dendritic cells.
  • IL-13 inhibitors or IL-13 signaling blockers can be used as therapeutic agents for atopy, psoriasis, and allergies.
  • Immune disease is an immune disease in which an immune response essential for maintaining human health is expressed in an excessive or harmful form.
  • the IL-4 or IL-13 inhibitor composition of the present invention inhibits IgE production, and thus can be used as a treatment for allergies, psoriasis, and atopic dermatitis.
  • the aldehyde dehydrogenation reaction is a reaction for changing acetaldehyde or various aldehydes present in human metabolites into organic acids that can be discharged as waste products.
  • the enzyme that promotes this aldehyde dehydrogenation reaction decomposes the aldehyde, which is toxic to cells, and relieves carbonyl stress applied to cells and tissues.
  • ALDH aldehyde dehydrogenases
  • WO2016021847 describes an oral anti-atopic composition comprising rice prolamin as an active ingredient.
  • the composition described in this document not only has anti-atopic efficacy, but also has high safety and can be widely used by including the protein of commercial food as an active ingredient.
  • WO2008/044894 discloses an atopy-inhibiting composition containing a bamboo extract.
  • organosulfur compound sulforaphane (1-isothiocyanato-4R-(methylsulfinyl)butane) which is at least one compound that modulates the expression, amount, or enzymatic activity of one or more acetaldehyde dehydrogenases in a cell.
  • a method for reducing peak blood levels of acetaldehyde and increasing the rate of catabolism of acetaldehyde in a subject comprising administering an effective amount of a derivative of sulforaphane, an analog of sulforaphane, wherein the expression of acetaldehyde dehydrogenase genes ALDH2, provided is a method encoded by one or more of ALDH1A, ALDH3A1, ALDH1B1, ALDH 1a1, or a combination thereof.
  • ALDH which is the subject of the present invention, may inhibit or may inhibit interleukin-4 or interleukin-13.
  • This prior art describes in detail mitochondrial aldehyde dehydrogenase-2 (ALDH2), a tetrameric protein consisting of four identical subunits encoded in the nuclear genome and transported into the mitochondria, each of 500 amino acid residues. wherein the tetramer is a dimer of a dimer, wherein the interface between the monomers forming the dimer is different from the interface between the two dimers forming the tetramer, and each subunit has three major sites, i.e., an aldehyde Dehydrogenation catalytic sites, coenzyme or NAD+-binding sites, and oligomerization sites are disclosed.
  • ADH2 mitochondrial aldehyde dehydrogenase-2
  • ALDH2 related diseases and conditions include ischemic stress, chronic free-radical related diseases, acute free-radical related diseases, insensitivity to nitroglycerin (eg, in angina and heart failure), hypertension, diabetes. , osteoporosis, cancer, Fanconi anemia, Alzheimer's disease, Parkinson's disease, alcoholism, alcohol intolerance, alcohol addiction, alcohol abuse disease, alcohol intoxication, alcohol dependence, alcohol poisoning
  • modulators of ALDH2, agonists of ALDH2, or antagonists of ALDH2, including symptoms of alcohol consumption, and drug addiction include ischemic stress, chronic free-radical related diseases, acute free-radical related diseases, insensitivity to nitroglycerin (eg, in angina and heart failure), hypertension, diabetes. , osteoporosis, cancer, Fanconi anemia, Alzheimer's disease, Parkinson's disease, alcoholism, alcohol intolerance, alcohol addiction, alcohol abuse disease, alcohol intoxication, alcohol dependence, alcohol poisoning
  • Agonists of ALDH2 are also useful for reducing the levels of compounds such as ethanol, methanol, ethylene glycol monomethyl ether, polyvinyl chloride, heterogeneous aldehydes and aldehydes in vivo in a subject.
  • An agonist of ALDH2 is also useful for reducing the level of a compound that generates an aldehyde substrate of ALDH2 in a subject upon ingestion, absorption, or inhalation of a compound that generates an aldehyde substrate of ALDH2.
  • Antagonists of ALDH2 are useful for treating or inhibiting diseases such as cancer, where ALDH2 antagonists are used as an adjuvant to standard cancer treatment.
  • Aldehyde Dehydrogenase Inhibitors a Comprehensive Review of the Pharmacology, Mechanism of Action, Substrate Specificity, and Clinical Application, by The American Society for Pharmacology and Experimental Therapeutics Pharmacol Rev. 64:520-539, 2012.
  • 19 types of ALDH play a very important role in eliminating biotoxicity by decomposing various types of aldehydes produced in the body or aldehydes supplied from outside the body. and drugs that inhibit the action of aldehydes that cause a strong oxidative reaction in human tissue cells are described in detail.
  • the above prior art did not suggest or suggest that ALDHs have immunomodulatory use by inhibiting IL-4 or IL-13.
  • the present inventors confirmed the novel fact that all or part of various ALDH fusion proteins and recombinant proteins containing an aldehyde dehydrogenation reaction promoting site inhibit IL-4 or IL-13, so that the ALDH-containing composition of the present application can be used as a pharmaceutical or New uses as functional foods have been invented.
  • the present inventors inhibit the secretion of IL-4 and IL-13 from Th2 cells by using the ALDH-containing composition, thereby reducing the IgE production of activated B lymphocytes, thereby reducing inflammation such as histamine produced by degranulation of mast cells.
  • the production of mediators was also reduced, confirming the fact that diseases caused by an increase in IgE production could be prevented or treated.
  • the present inventors found that when reactive oxygen species (ROS) generated in the metabolic process of cells is excessively generated, it causes abnormalities in the immune system and IL-4 and IL- as one of the causes of autoimmune diseases.
  • ROS reactive oxygen species
  • the expression of 13 is increased, and the resulting oxidative stress causes contraction of smooth muscle in the lungs, increased vascular permeability, damage to drug-responsive receptors, increased secretion of mucus, secretion of inflammatory mediators, and damage to airway epithelial cells, leading to atopy, psoriasis, and psoriasis.
  • allergic dermatitis, and bronchial asthma were confirmed to be the cause of various diseases.
  • the IL-4 or IL-13 inhibition test results of the composition of the present invention the IL-4 or IL-13 inhibitory effect of ALDH and the IL-4 or IL-13 effect on the human immune system reported to date, It enables new pharmaceutical uses of ALDH as immunomodulators.
  • the primary object of the present invention is to contain all or a part of a fusion protein or recombinant protein (hereinafter abbreviated as ALDH) comprising an aldehyde dehydrogenase that inhibits IL-4 or IL-13 and a site that promotes a dehydrogenation reaction. It is to provide a therapeutic agent for immune diseases such as atopy, allergy, and psoriasis.
  • ALDH fusion protein or recombinant protein
  • the primary object of the present invention is to provide a composition containing, as an active ingredient, aldehyde dehydrogenation promoting enzymes and variants thereof, including fusion proteins or recombinant proteins comprising a site of action involved in aldehyde dehydrogenation reaction. .
  • Another object of the present invention is an immune response inhibitor or autoimmune disease containing aldehyde dehydrogenation promoting enzymes and variants thereof as active ingredients, including fusion proteins or recombinant proteins comprising a site involved in aldehyde dehydrogenation reaction.
  • aldehyde dehydrogenation promoting enzymes and variants thereof as active ingredients, including fusion proteins or recombinant proteins comprising a site involved in aldehyde dehydrogenation reaction.
  • Another object of the present invention is to provide a therapeutic agent for atopic dermatitis, psoriasis, and allergic dermatitis, which contains aldehyde dehydrogenases and variants thereof as active ingredients, including fusion proteins or recombinant proteins comprising a site of action involved in aldehyde dehydrogenation reaction will do
  • aldehyde dehydrogenase and its variants containing a fusion protein containing a part or all of the action site that promotes the aldehyde dehydrogenase reaction of the aldehyde dehydrogenase, recombinant protein containing It can be achieved by providing a therapeutic agent for atopic dermatitis, psoriasis, and allergic dermatitis.
  • the gist of the object and solution of the present invention as described above is a novel use of a known substance, ALDH, as an IL-4 or IL-13 inhibitor. Therefore, the ALDH that can be used as an IL-4 or IL-13 inhibitor of the present invention can include all of the various ALDHs, which are known substances exhibiting aldehyde dehydrogenation activity to date, and all of these are atopic dermatitis, psoriasis, and allergies of the present invention. It can be used as an active ingredient in a dermatitis treatment agent, and is included in the scope of the present invention.
  • KCTC International Depositary Organization
  • KCTC13925BP Saccharomyces cerevisiae KwonP-1
  • KCTC14122BP Saccharomyces cerevisiae KwonP-2
  • KCTC14123BP Saccharomyces cerevisiae KwonP-3
  • ALDH-producing strains were deposited, and the measurement test results for atopic, psoriasis, and allergic dermatitis treatments according to the IL-4 or IL-13 inhibitory effect of various known ALDHs were reviewed. It is disclosed through the specification of the application.
  • ALDH or a composition containing ALDH of the present invention exhibits a pharmaceutical effect for preventing and treating various immune diseases due to the inhibitory action of interleukin-4 or interleukin-13.
  • ALDH contained in the yeast extract according to the present invention showed immunoglobulin E (IgE) and total number of inflammatory cells and lymphocytes, monocytes, neutrophils, and eosinophils in the bronchoalveolar lavage fluid of asthma-induced mice in the in vivo experiment of FIG. It has the activity of inhibiting the increase and the secretion of airway mucus, and by reducing the Th2 cytokines IL-4 and IL-13 and airway inflammation, it can be used as a therapeutic agent for atopic dermatitis, psoriasis, allergic dermatitis, and as a health food.
  • IgE immunoglobulin E
  • the ALDH-containing composition according to the present invention is effective in treating and preventing inflammatory reactions caused by oxidative stress such as inflammation, allergy, asthma, and the like.
  • ALDH is extracted from edible yeast, it has the advantage that the possibility of side effects is low even if it is taken.
  • A CON
  • B atopy-induced group with DNCB
  • C dexamethasone administration group
  • D 50 mg/kg of the composition of the present invention
  • E 150 mg/kg of the composition of the present invention
  • F 300 mg/kg of the present composition
  • FIG. 2 is a graph showing changes in blood leukocytes (WBC) in atopy-induced control group and experimental group mice.
  • WBC blood leukocytes
  • DNCB-induced increase in the amount of leukocytes in the blood was reduced by 65% in the case of dexamethasone administration, and reduced by about 74% in the case of administration of the composition of the present invention (150mg/Kg).
  • FIG. 3 is a graph showing changes in eosinophils in atopy-induced control and experimental mice.
  • the increase in blood eosinophil due to atopy induction by DNCB was reduced by 60% when dexamethasone was administered, and decreased by about 55% when the composition of the present invention (150mg/Kg) was administered.
  • FIG. 4 is a graph showing changes in blood neutrophils in atopic-induced control and experimental mice. Blood neutrophils increased by DNCB induction were reduced by 35% by administration of dexamethasone, and reduced by about 46% by administration of the composition of the present invention (150mg/Kg).
  • 5 is a graph showing changes in IL-6 in atopy-induced control group and experimental group mice.
  • FIG. 6 is a graph showing changes in IL-6 mRNA in atopy-induced control and experimental mice.
  • Example 1-1 Saccharomyces cerevisiae yeast fermentation process for preparing the composition of the present invention
  • the ALDH-containing Saccharomyces cerevisiae yeast seed (KCTC13925BP) was fermented for 24 hours in an incubator at 160 rpm and 30°C using YPD medium (a medium containing yeast extract, peptone, and glucose) in a 200 mL flask and cultured. and this culture was carried out for 72 hours through a 5L fermenter (Marado-05D-PS, CNS, Korea). After completion of the culture, the yeast was centrifuged using a high-speed centrifuge (Supra R22, Hanil, Korea).
  • YPD medium a medium containing yeast extract, peptone, and glucose
  • Example 1-2 ALDH-containing yeast lysate preparation process
  • the centrifuged ALDH-containing yeast was frozen in a cryogenic freezer (CLN-52U, Nihon freezer, Japan) for 2 days, and then freeze-dried for 2 days in a freeze dryer (FDU-7006, Operon, Korea).
  • a freeze dryer FDU-7006, Operon, Korea.
  • PBS phosphate-buffered saline
  • 0.5 mm glass beads for cell disruption 11079105, Biospec 10 g was put into the bead homogenizer (Mixer Mill MM400, Retsch, Germany) to disrupt yeast over a total of 3 times for 2 minutes each.
  • After centrifugation using a high-speed centrifuge (Supra R22, Hanil, Korea), only the supernatant was separated and freeze-dried for 2 days with a freeze dryer (FDU-7006, Operon, Korea).
  • Example 2-1 Preparation of atopy-induced test animals.
  • mice with an average weight of about 20 g were purchased from Samtaco Bio Korea.
  • DNCB mouse dermatitis inducer
  • Example 2-2 autopsy of animals
  • composition of the present invention was orally administered in the second week of induction and for 5 days, and autopsies were performed the next day.
  • the back skin tissue and spleen were divided into two, one for checking the secretion of cytokine, and the other for checking gene expression. It was stored at -80°C for use.
  • Example 2-3 Analysis of blood IgE content in atopic mice
  • the blood collected from the test animals was placed in a 1.5 mL E-tube and centrifuged for 20 minutes at 12,000 rpm and 4°C. After that, only the supernatant was separated and used for the experiment. Quantification of IgE was performed by the Enzyme-Linked Immunosorbent Assay (ELISA) method, and quantitative analysis was performed using the IgE ELISA set (BD bioscience). After the reaction, the absorbance at the final 450 nm wavelength was measured with an ELISA reader (Multiskan sky, Thermo Fisher, USA), and the results are shown in FIG. 1 . In the group administered with the ALDH-containing composition of the present invention, IgE was reduced by about 40% or more compared to the atopy-induced experimental group.
  • ELISA Enzyme-Linked Immunosorbent Assay
  • FIG. 3 Compared to the atopy-inducing group (DNCB), IgE was reduced by 38% in the group administered with the composition of the present invention (150 mg/Kg), and IgE was reduced by 45% in the group administered with the composition of the present invention (300 mg/Kg). Atopic therapeutically effective amount of the composition of the present invention was evaluated as 150 mg/Kg in a mouse animal test.
  • Example 2-4 Confirmation of the pharmacological effect of inhibiting interleukin-13 by the ALDH-containing composition of the present invention in atopic mice
  • Example 2-5 Hematology analysis of atopic mice
  • eosinophilic and neutrophilic infiltration in tissues is a characteristic phenomenon (Horn BR, Robin ED, Theodore J, Van Kessel A. Total eosinophil counts in the management of bronchial asthma) (N Engl J Med 1975;292:1152-5).
  • the more the eosinophil-type cells differentiate the more the bone marrow is biased toward an atopic state, which can cause allergic inflammatory reactions.
  • blood cells such as leukocytes, eosinophils, and neutrophils were analyzed in atopic-induced mice to confirm the pharmacological effect of the ALDH-containing yeast lysate of the present invention.
  • the analysis of blood cells was performed immediately after the autopsy of the blood contained in the heparin tube using a hemocytometer (Hemavet 950FS, Drew Scientific Inc, Korea).
  • the neutrophil (NE) content in blood was reduced by 35% in the case of dexamethasone administration, and by about 46% in the case of administration of 150 mg/kg of the composition of the present invention (Fig. 4), the blood neutrophil (NE) content characteristically in allergic reactions such as atopy
  • the ALDH-containing composition of the present invention is similar to that of the control group or showed excellent pharmacological effects.
  • Example 2-6 Analysis of IL-6 changes in atopic-induced mice and the group administered with the composition of the present invention
  • composition of the present invention was orally administered to Balb/c mice induced by atopy for 2 weeks with DNCB solution for 5 days.
  • skin tissue was used as a sample for serum, ELISA and RT-PCR, and dexamethasone was used as a positive control.
  • the expression level of IL-6 cyokine in the skin tissue was measured by hemocytometry.
  • IL-6 expression levels in mouse skin tissues were analyzed by ELISA assay and real-time PCR to quantify IL-6 protein and mRNA levels.
  • the effect of equal or superiority was confirmed in the group administered with the composition of the present invention compared to the group administered with dexamethasone.
  • the IL-6 content in the skin tissue increased by DNCB induction was reduced by 50%, and in the case of administration of the composition of the present invention (150mg/Kg), it was reduced by about 48%.
  • Example 3-1 Preparation of atopic dermatitis human cell model using TNF- ⁇ and IFN- ⁇
  • HaCaT cells which are human keratinocytes, were used as the cells.
  • the medium was DMEM (Dolbicco's Modified Eagle's Medium, USA) in which 10% Fetal bovine serum (FBS) and + 1% P/S (Penicillin/Streptomycin: P/S) were added, seeding and After 23 hours, the ALDH-containing composition of the present invention was treated at concentrations of 0.5, 1, and 2 ⁇ g/mL, and after 1 hour, inflammation was induced by treatment with 10 ng/mL of TNF- ⁇ and IFN- ⁇ .
  • DMEM Dolbicco's Modified Eagle's Medium, USA
  • FBS Fetal bovine serum
  • P/S Penicillin/Streptomycin
  • Inflammation was induced in HaCaT cells with TNF- ⁇ and IFN- ⁇ , and after 1 hour, the medium in a 6-well plate was transferred to an E-tube, and centrifugation was performed at 13,000 rpm and 4°C for 10 minutes. After that, only the supernatant was transferred to a new tube and used for the experiment.
  • Enzyme-Linked Immunosorbent Assay For the analysis, an IgE ELISA set (BD bioscience, US) was used. Capture Ab diluted in coating buffer was added to micro wells and incubated at 4 °C overnight. After washing 3 times, put assay diluent into each well and incubate at room temperature. After washing 3 times, standard and sample diluted with assay diluent were added to each well and incubated at room temperature. After washing 5 times, a working detector was put into each well and incubated at room temperature. After washing 7 times, the substrate solution was added to each well and incubated for 30 minutes in dark conditions at room temperature. To terminate the reaction, 50 ⁇ L of Stop Solution was added to each well, and absorbance at 450 nm wavelength was measured with an ELISA reader (Multiskan sky, Thermo Fisher, USA).
  • the human keratinocyte HaCaTcell line was pretreated with the composition of the present invention at each concentration for 20 minutes, and TNF ⁇ /IFN ⁇ was added to the pretreated cells to induce an intracellular inflammatory state.
  • the expression level of IL-6 a major marker of atopic disease animal models, was measured. Compared to the IL6 expression level of the atopic-induced cell line, it was confirmed that the IL-6 expression level decreased by 0.03%, 14%, and 27% for each concentration in the cells treated with the composition of the present invention. (Fig. 8)
  • mice Female and male ICR mice (7 weeks old) were received and acclimatized for 7 days. During the acclimatization period, general symptoms were observed, and only healthy animals were used for the test. Feed and water were ingested ad libitum, and group separation was performed so that there were 5 males and 5 females in each group based on the average body weight of about 20 g the day before oral administration.
  • the test substance was prepared by dissolving in physiological saline so that the doses of the test animals were 0, 750, 3000, and 5000 mg/Kg, respectively, based on the content of the ALDH-containing yeast lysate of the present invention.
  • the standard of administration dose was in accordance with the Korea National Toxicology Program (KNTP) toxicity test manual of the Ministry of Food and Drug Safety.
  • KNTP National Toxicology Program
  • the maximum application dose of 5000 mg/Kg guided by the KNTP manual was applied as the maximum concentration in this experiment.
  • Samples prepared for each group were orally administered once to each test animal, and physiological saline was administered to the normal group (G1).
  • Symptoms were observed at least once a day from the date of acquisition to the day of autopsy for all animals in the test group, and symptoms were observed for 7 days after oral administration. After the type of symptom observation, an autopsy was performed, and changes in each organ were visually observed at the time of autopsy.
  • psoriasis treatment, atopic treatment, and allergy treatment containing aldehyde dehydrogenase (ALDH) of the present invention psoriasis treatment, atopic treatment, and allergy treatment containing aldehyde dehydrogenase (ALDH) of the present invention, pharmacological effect compared to control drug, administration method, therapeutically effective amount for disease model, acute toxicity
  • ADH aldehyde dehydrogenase

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Abstract

La présente invention concerne un agent thérapeutique pour la dermatite atopique, le psoriasis et des allergies contenant une aldéhyde déshydrogénase, l'agent thérapeutique pour la dermatite atopique, le psoriasis et l'allergie contenant, en tant que principes actifs, des enzymes accélérant la déshydrogénation d'aldéhyde et des variants de celles-ci, comprenant une protéine de fusion ou une protéine recombinante comprenant un site d'action impliqué dans la déshydrogénation d'aldéhyde.
PCT/KR2021/001935 2020-02-18 2021-02-16 Composition inhibitrice d'atopie contenant une aldéhyde déshydrogénase Ceased WO2021167310A1 (fr)

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KR10-2020-0019824 2020-02-18
KR20200019793 2020-02-18
KR20200019824 2020-02-18
KR10-2020-0019793 2020-02-18
KR1020210020183A KR102460570B1 (ko) 2020-02-18 2021-02-15 신규한 돌연변이 효모를 함유하는 아토피 억제 조성물
KR10-2021-0020183 2021-02-15

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Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2011158689A1 (fr) * 2010-06-19 2011-12-22 天野エンザイム株式会社 Agent permettant de réduire l'acétaldéhyde dans la cavité orale
JP2013247939A (ja) * 2012-06-04 2013-12-12 Kiso Machi 新規酵母及びこれを含む医薬又は飲食品
KR20180003344A (ko) * 2016-06-30 2018-01-09 (주)아모레퍼시픽 효모 유래 세포밖 소포체를 포함하는 항염 조성물

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2011158689A1 (fr) * 2010-06-19 2011-12-22 天野エンザイム株式会社 Agent permettant de réduire l'acétaldéhyde dans la cavité orale
JP2013247939A (ja) * 2012-06-04 2013-12-12 Kiso Machi 新規酵母及びこれを含む医薬又は飲食品
KR20180003344A (ko) * 2016-06-30 2018-01-09 (주)아모레퍼시픽 효모 유래 세포밖 소포체를 포함하는 항염 조성물

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"Study on the functional role of acetaldehyde dehydrogenase in atopic dermatitis for the development of biomarker", FINAL REPORT: HEALTH TECHNOLOGY R&D PROJECT, 11 November 2015 (2015-11-11), Korea, pages 1 - 78, XP009529965 *
BAZEWICZ CHRISTOPHER G.: "Aldehyde dehydrogenase in regulatory T‐cell development, immunity and cancer", IMMUNOLOGY, vol. 156, 1 January 2018 (2018-01-01), pages 47 - 55, XP055840160 *

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