Immunosuppressive composition comprising L-AHG

Abstract

The present invention relates to an immunosuppressive composition comprising 3, 6-anhydro-L-galactose (L-AHG) as an active ingredient, and to a pharmaceutical composition, a functional health food, and a functional cosmetic composition for preventing or treating immune diseases comprising the active ingredient. The immunosuppressive composition containing L-AHG, the pharmaceutical composition, the functional health food, and the functional cosmetic composition according to the present invention have excellent effects of selectively and effectively suppressing activation, proliferation, and differentiation of CD4T cells, which are the main target of an immunosuppressive agent, and thus, can be used as an immunosuppressive agent. The invention is very useful in the pharmaceutical, food and cosmetic industries.

Description

Background Art Technical Field

[0001] The present disclosure relates to an immunosuppressive composition comprising 3,6-anhydro-L-galactose (L-AHG) as an active ingredient, a pharmaceutical composition for preventing or treating immune diseases, a healthy functional food for preventing or improving immune diseases, and a functional cosmetic composition comprising the active ingredient for preventing or improving hypersensitivity skin reactions.

[0002] Related technologies

[0003] In some cases, the immune system may behave inappropriately. As a typical example, in some cases, the immune system overreacts due to the inability to distinguish between self and non-self molecules, resulting in hypersensitivity reactions, or induces immune responses against self molecules, resulting in autoimmune diseases. In addition, from the perspective of allograft maintenance and survival, immune rejection of an allograft transplanted for therapeutic purposes by the recipient's immune system can be considered an inappropriate immune response.

[0004] Allergic reactions or type 1 hypersensitivity reactions in humans are mainly reactions involving immunoglobulin E (IgE) antibodies. When specific antigens (allergens) bind to IgE on the surface of mast cells and basophils in the body, various mediators that cause hypersensitivity reactions and inflammation are released from these cells. The main symptoms of hypersensitivity reactions include sneezing, wheezing, asthma, atopic dermatitis, rash (urticaria), etc.

[0005] Autoimmunity, in which the immune system loses its ability to distinguish between self and non-self molecules and immunologically attacks the host, leads to a variety of chronic and debilitating diseases. Major diseases caused by autoimmunity include multiple sclerosis, rheumatoid arthritis, systemic lupus erythematosus, type 1 diabetes, psoriasis, Hashimoto's thyroiditis, and acute inflammatory bowel disease [IBD; Crohn's disease / ulcerative colitis], which is a more recent problem in younger populations.

[0006] When the body's immune system encounters foreign cells or tissues, it responds strongly to eliminate these foreign factors. However, in some cases, even if the patient's immune system recognizes foreign factors and initiates an immune response to reject them, transplantation of cells, tissues or organs from other donors may be the only way to treat the patient's disease. For example, it is estimated that in the United States alone, more than 100,000 people need treatment through kidney transplantation each year. However, the immune system attacks and eliminates any transplanted cells, tissues or organs that are not recognized as self through immune rejection. Therefore, immune rejection has become a serious obstacle to life-saving transplantation therapy. An additional risk of transplantation is that the immune cells contained in the donated tissue (e.g., a bone marrow transplant to restore immunity) recognize the new host as non-self and respond against the new host. This response is called graft-versus-host disease (GVHD) and can be fatal.

[0007] Therefore, immunosuppressive drugs have been used to control inappropriate immune responses that occur in the human immune system, such as hypersensitivity reactions, autoimmune diseases, immune rejection of allogeneic transplants, and graft-versus-host disease. Currently used immunosuppressants include corticosteroids (prednisone), antiproliferative agents [mycophenolate mofetil ], calcineurin inhibitors [(tacrolimus (Envarsus or Protopic), cyclosporine ( or )], mTOR inhibitors [sirolimus ], immune cell targeting polyclonal antibodies and monoclonal antibodies, etc. The currently used immunosuppressants still cannot completely eliminate side effects, leading to susceptibility to infectious diseases, cardiovascular risks, and damage to other organs such as the kidney and liver, and the efficacy is also unsatisfactory. Therefore, it is necessary to develop new safe and targeted specific immunosuppressants with minimal side effects.

[0008] In recent years, due to the enhanced understanding of the human immune system and the development of biotechnology, various types of immunosuppressants can be used alone or in combination according to the patient's condition and symptoms for the treatment of patients with hypersensitivity reactions, patients with autoimmune diseases, and organ transplant patients (including stem cell or bone marrow transplant patients) who need to be administered immunosuppressants clinically.

[0009] The target of action of typical immunosuppressants used for patients with the purpose of artificially suppressing human immune function is concentrated on T cells. In fact, suppressing the function of CD4 T (Th) cells, which is mainly in T cells, is its representative mechanism of action. This is because cytokines derived from effector Th cells are not only important in the cellular immune response in which T cells play a leading role, but also in the humoral immune response in which antibody molecules derived from B cells play a leading role. In particular, in several diseases such as Sjögren's syndrome, Behcet's disease, systemic sclerosis, polymyositis and dermatomyositis, it is reported that cytotoxic CD4+T cells are an important cause of disease, and inhibiting T cell activation may be an important treatment option.

[0010] Therefore, by appropriately blocking the activation, proliferation and differentiation of resting (G0) Th cells into effector Th cells, the immunosuppressive effect can be well exerted. The importance of Th cells in the induction and regulation of immune responses is well demonstrated by the occurrence of immunodeficiency syndrome in patients with Th cell dysfunction caused by infection with human immunodeficiency virus (HIV).

[0011] In order to alleviate the side effects, potential risks or complications caused by the use of immunosuppressants developed to date, there is a continuous need to develop new immunosuppressants that can specifically suppress only inappropriate immune responses. Summary of the invention

[0012] The present disclosure is proposed to solve the above-mentioned problems arising in the prior art. The purpose of the present disclosure is to provide a novel immunosuppressive composition that can effectively inhibit the activation, proliferation and differentiation of CD4 T cells, which are the main targets of immunosuppressants, a composition for preventing or treating immune diseases, a healthy functional food for preventing or improving immune diseases, and a functional cosmetic composition for preventing or improving hypersensitivity skin reactions.

[0013] In order to achieve the above-mentioned object, the present disclosure provides an immunosuppressive composition comprising 3,6-anhydro-L-galactose (L-AHG) as an active ingredient.

[0014] The composition preferably inhibits the activation, proliferation or differentiation of CD4 T cells.

[0015] The composition preferably blocks cell cycle progression at the G1 or S phase.

[0016] The composition preferably inhibits differentiation into Th1 cells or Th2 cells.

[0017] The composition preferably inhibits the production of interferon-γ (INF-γ), interleukin-2 (IL-2), interleukin-4 (IL-4) or interleukin-13 (IL-13).

[0018] The present disclosure also provides a pharmaceutical composition for preventing or treating an immune disease, comprising 3,6-anhydro-L-galactose (L-AHG) as an active ingredient.

[0019] The immune disease is preferably selected from the group consisting of hypersensitivity diseases, autoimmune diseases, immune rejection, graft-versus-host disease, Sjögren's syndrome and Behcet's disease.

[0020] The hypersensitivity disease is preferably selected from the group consisting of allergy, sneezing, wheezing, asthma, atopic dermatitis and urticaria.

[0021] The autoimmune disease is preferably selected from the group consisting of multiple sclerosis, rheumatoid arthritis, systemic lupus erythematosus, type 1 diabetes, psoriasis, Hashimoto's thyroiditis, systemic lupus erythematosus and acute inflammatory bowel disease.

[0022] The present disclosure also provides a health functional food for preventing or treating immune diseases, comprising 3,6-anhydro-L-galactose (L-AHG) as an active ingredient.

[0023] The immune disease is preferably selected from the group consisting of hypersensitivity diseases, autoimmune diseases, immune rejection, graft-versus-host disease, Sjögren's syndrome and Behcet's disease.

[0024] The hypersensitivity disease is preferably selected from the group consisting of allergy, sneezing, wheezing, asthma, atopic dermatitis and urticaria.

[0025] The autoimmune disease is preferably selected from the group consisting of multiple sclerosis, rheumatoid arthritis, systemic lupus erythematosus, type 1 diabetes, psoriasis, Hashimoto's thyroiditis, systemic lupus erythematosus and acute inflammatory bowel disease.

[0026] The present disclosure also provides a functional cosmetic composition for preventing or improving hypersensitivity skin reactions, comprising 3,6-anhydro-L-galactose (L-AHG) as an active ingredient.

[0027] The hypersensitivity skin reaction is preferably selected from the group consisting of solar dermatitis, contact dermatitis and atopic dermatitis.

[0028] The immunosuppressive composition, pharmaceutical composition, health functional food, and functional cosmetic composition containing L-AHG according to the present disclosure have excellent effects of selectively and effectively inhibiting the activation, proliferation, and differentiation of CD4 T cells, which are the main targets of immunosuppressants, and are therefore very useful in the pharmaceutical industry, the food industry, and the cosmetics industry. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] Figure 1 CD4 (T H ) and CD8(T C ) cell distribution.

[0030] Figure 2 Shown to be carried out 3 Results of [H]-thymidine incorporation assays to evaluate the effects of AGH, neoagarobiose (NA2), neoagarotetraose (NA4), and neoagarohexose (NA6), obtained from enzymatic hydrolysis of agarose, on the proliferation of T cells activated with immobilized anti-CD3 / anti-CD28.

[0031] Figure 3 Results indicating that L-AGH specifically blocks cell cycle progression in CD4 T cells are shown. G0 T cells were treated with nocodazole 20 hours after their activation.

[0032] Figure 4 Shown are results indicating that L-AGH blocks cell cycle progression of CD4 T cells not treated with nocodazole.

[0033] Figure 5 Results showing that L-AGH treatment reduced the secretion of Th1 signature cytokines (INF-γ and IL-2) and Th2 signature cytokines (IL-4 and IL-13) secreted by T cells activated with immobilized anti-CD3 / anti-CD28 are shown. DETAILED DESCRIPTION

[0034] Hereinafter, the present disclosure will be described in detail.

[0035] In the present disclosure, a combination of recombinant GH16B β-agarase, GH50A β-agarase [Kwon et al., 2020] and GH117 α-neoagarobiohydrolase (GH117A α-NABH) [Jang et al., 2021] derived from a freshwater agar-degrading bacterium Cellvibrio sp. KY-GH-1 strain (KCTC 13629BP) isolated from a non-marine environment is used for agarose saccharification by an enzymatic process, and then a mixture of monomeric 3,6-anhydro-L-galactose (L-AHG) and D-galactose is produced as the final degradation product, which is fractionated by size exclusion chromatography using a Sephadex G-10 column to finally purify L-AHG. In order to study the immunosuppressive activity of purified L-AHG, a model of in vitro induction of activation and cell cycle of resting (G0) T cells in mice was used to examine the effects of L-AHG on the activation and proliferation of resting Th cells. The results confirmed that resting Th cells were activated in the G0 phase by simultaneous stimulation of immobilized anti-CD3 bound to the T cell receptor complex and immobilized anti-CD28 bound to the T cell co-stimulatory receptor (CD28), and when the cell cycle proceeded in the order of G1→S→G2 / M phases, the presence of L-AHG could exert an immunosuppressive effect of reducing T cell proliferation by inhibiting the progression of the G1 and S phases of the cell cycle in activated Th cells.

[0036] Accordingly, the present disclosure provides an immunosuppressive composition comprising 3,6-anhydro-L-galactose (L-AHG) as an active ingredient.

[0037] The active ingredient can be obtained from acid hydrolysis or enzymatic degradation of agarose.

[0038] In a preferred embodiment, the active ingredient of the present disclosure can be obtained by acid hydrolysis of agarose.

[0039] In another embodiment, the active ingredient of the present disclosure can be obtained by performing agarosylation through an enzymatic process using a combination of recombinant GH16B β-agarase, GH50A β-agarase, and GH117 α-neoagarobiohydrolase derived from an agar-degrading bacterium Vibrio cellulosus KY-GH-1 strain (KCTC 13629BP), and then fractionating a mixture of monomeric 3,6-anhydro-L-galactose (L-AHG) and D-galactose produced as the final degradation product by size exclusion chromatography using a Sephadex G-10 column.

[0040] In another preferred embodiment, the active ingredient of the present disclosure can be obtained from the agarase-3,6-anhydro-L-galactosidase-arabinose isomerase complex enzyme product of agarose.

[0041] In addition to the above-mentioned several methods for obtaining L-AHG, the active ingredient of the present disclosure can also be obtained through commercial channels. For example, the active ingredient can be purchased from Toronto Research Chemicals (North York, Ontario, Canada).

[0042] The composition preferably inhibits the activation, proliferation or differentiation of CD4 T cells.

[0043] The composition preferably blocks cell cycle progression at the G1 or S phase.

[0044] The composition preferably inhibits differentiation into Th1 cells or T2 cells.

[0045] The composition preferably inhibits the production of interferon-γ (INF-γ), interleukin-2 (IL-2), interleukin-4 (IL-4) or interleukin-13 (IL-13).

[0046] The present disclosure also provides a pharmaceutical composition for preventing or treating an immune disease, comprising 3,6-anhydro-L-galactose (L-AHG) as an active ingredient.

[0047] The immune disease is preferably selected from the group consisting of hypersensitivity diseases, autoimmune diseases, immune rejection, graft-versus-host disease, Sjögren's syndrome and Behcet's disease.

[0048] The hypersensitivity disease is preferably selected from the group consisting of allergies, sneezing, wheezing, asthma, atopic dermatitis and urticaria.

[0049] The autoimmune disease is preferably selected from the group consisting of multiple sclerosis, rheumatoid arthritis, systemic lupus erythematosus, type 1 diabetes, psoriasis, Hashimoto's thyroiditis, systemic lupus erythematosus and acute inflammatory bowel disease.

[0050] The immunosuppressive compositions and pharmaceutical compositions comprising the active ingredients of the present disclosure can be formulated and used in a variety of dosage forms, including oral dosage forms such as powders, granules, tablets, capsules, suspensions, emulsions, syrups and aerosols, and injectable preparations such as sterile injection solutions, according to conventional methods depending on the respective purposes of use, and can be administered orally or by various routes, including intravenous, intraperitoneal, subcutaneous, intrarectal and topical routes.

[0051] The pharmaceutical composition may further comprise carrier, excipient or diluent. The example of the suitable carrier, excipient or diluent that may be comprised comprises lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, gum arabic, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methylcellulose, amorphous cellulose, polyvinylpyrrolidone, water, methyl hydroxybenzoate, propyl hydroxybenzoate, talcum, magnesium stearate and mineral oil. In addition, the pharmaceutical composition of the present disclosure may further comprise filler, anti-agglomeration agent, lubricant, wetting agent, flavoring agent, emulsifier, preservative etc.

[0052] In a preferred embodiment, solid preparations for oral administration include tablets, pills, powders, granules, capsules, etc., and these solid preparations are prepared by mixing the immunosuppressive composition or the pharmaceutical composition with one or more excipients (e.g., starch, calcium carbonate, sucrose, lactose, gelatin, etc.). In addition, in addition to simple excipients, lubricants such as magnesium stearate and talc may also be used.

[0053] In a preferred embodiment, oral liquid preparations include suspensions, internal solutions, emulsions, syrups, etc., and may contain various excipients such as wetting agents, sweeteners, flavoring agents and preservatives in addition to water and liquid paraffin (commonly used simple diluents).

[0054] In a preferred embodiment, preparations for parenteral administration include sterile aqueous solutions, non-aqueous solutions, suspensions, emulsions, lyophilized preparations, suppositories, etc. Non-aqueous solutions and suspensions may include propylene glycol, polyethylene glycol, vegetable oils (such as olive oil) and injectable esters (such as ethyl oleate). Injectable preparations may include conventional additives, such as solubilizers, isotonic agents, suspending agents, emulsifiers, stabilizers, preservatives, etc.

[0055] The active ingredients of the present disclosure are administered in a pharmaceutically effective amount. As used herein, the term "pharmaceutically effective amount" refers to an amount sufficient to treat a disease at a reasonable benefit / risk ratio applicable to any medical treatment. The effective dosage level of the active ingredient can be determined based on factors including the following: the type and severity of the patient's disease, the activity of the drug, sensitivity to the drug, the time of administration, the route of administration, the excretion rate, and the drugs used in combination with the active ingredient, as well as other factors well-known in the medical field. The immunosuppressive composition or pharmaceutical composition of the present disclosure can be administered alone or in combination with other therapeutic agents, and can be administered sequentially or simultaneously with conventional therapeutic agents. The immunosuppressive composition or pharmaceutical composition can be administered in a single dosage form or in multiple dosage forms. Taking all of the above factors into account, it is important to administer the immunosuppressive composition or pharmaceutical composition in the minimum amount that can show the maximum effect without causing side effects, and this amount can be easily determined by those skilled in the art.

[0056] In a preferred embodiment, the effective amount of the active ingredient in the immunosuppressive composition or pharmaceutical composition of the present disclosure can vary according to the age, sex and weight of the patient. In general, the active ingredient can be administered daily or every other day at a dosage of 1 mg to 1,000 mg / kg body weight, preferably 5 mg to 100 mg / kg body weight, more preferably 10 mg to 20 mg / kg body weight, or can be administered 1 to 3 times a day at this dosage. However, the dosage is not intended to limit the scope of the present disclosure in any way, because the dosage can be increased or decreased according to the route of administration, the severity of the disease, the sex, weight and age of the patient, etc.

[0057] The immunosuppressive compositions or pharmaceutical compositions of the present disclosure can be administered to a subject by various routes. All modes of administration are contemplated, and for example, the compositions can be administered orally, rectally, or by rectal, intravenous, intramuscular, subcutaneous, intrauterine, intrathecal, or intraventricular injection.

[0058] As used herein, the term "administering" means providing a given substance to a patient by any suitable method. The immunosuppressive composition or pharmaceutical composition of the present disclosure can be administered orally or parenterally by any general route, as long as it can reach the target tissue. In addition, the immunosuppressive composition or pharmaceutical composition of the present disclosure can also be administered using any device capable of delivering the active ingredient to the target cell.

[0059] In the present disclosure, the term "subject" is not particularly limited but includes, for example, humans, monkeys, cows, horses, sheep, pigs, chickens, turkeys, quails, cats, dogs, mice, rats, rabbits or guinea pigs, and preferably refers to mammals, more preferably humans.

[0060] The present disclosure also provides a health functional food for preventing or treating immune diseases, comprising 3,6-anhydro-L-galactose (L-AHG) as an active ingredient.

[0061] The immune disease is preferably selected from the group consisting of hypersensitivity diseases, autoimmune diseases, immune rejection, graft-versus-host disease, Sjögren's syndrome and Behcet's disease.

[0062] The hypersensitivity disease is preferably selected from the group consisting of allergies, sneezing, wheezing, asthma, atopic dermatitis and urticaria.

[0063] The autoimmune disease is preferably selected from the group consisting of multiple sclerosis, rheumatoid arthritis, systemic lupus erythematosus, type 1 diabetes, psoriasis, Hashimoto's thyroiditis, systemic lupus erythematosus and acute inflammatory bowel disease.

[0064] The health functional food of the present disclosure can be used in various ways in foods and beverages that effectively prevent and improve immune diseases. Foods containing the active ingredients of the present disclosure include various foods, such as beverages, chewing gum, tea, vitamin complexes, health supplement foods, etc., and can be used in the form of powders, granules, tablets, capsules or beverages.

[0065] The active ingredient of the present disclosure can be generally added in an amount of 0.001 to 10 wt % based on the total weight of the food. For a health drink composition, the active ingredient can be added in an amount of 0.001 to 10 g, preferably 0.01 to 1 g, based on 100 ml of the health drink composition.

[0066] In addition to containing the specified percentage of compounds as basic components, the health functional food of the present disclosure may additionally contain food-acceptable additives such as natural carbohydrates and various flavoring agents.

[0067] Examples of natural carbohydrates include traditional sugars such as monosaccharides (e.g., glucose, fructose, etc.), disaccharides (e.g., maltose, sucrose, etc.), polysaccharides (e.g., dextrin, cyclodextrin, etc.), and sugar alcohols such as xylitol, sorbitol, erythritol, etc.

[0068] Examples of flavoring agents include natural flavoring agents such as thaumatin, rebaudioside A or glycyrrhizin, saccharin, and synthetic flavoring agents such as aspartame. The amount of natural carbohydrates used is generally about 1 to 20 g, preferably about 5 to 12 g, based on 100 mL of the health functional food of the present disclosure.

[0069] In addition, the health functional food of the present invention may contain various nutrients, vitamins, minerals, flavoring agents (such as synthetic flavoring agents and natural flavoring agents), colorants, extenders, pectin acid and its salts, alginic acid and its salts, organic acids, protective colloid thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohol, carbonizing agents used in carbonated beverages, etc.

[0070] In addition, the health functional food of the present disclosure may include pulp for preparing natural fruit juice, fruit juice beverage or vegetable beverage. These components can be used alone or in combination. Based on 100 parts by weight of the active ingredient of the present disclosure, the content of these additives is usually selected in the range of 10 to about 50 parts by weight.

[0071] The present disclosure also provides a functional cosmetic composition for preventing or improving hypersensitivity skin reactions, comprising 3,6-anhydro-L-galactose (L-AHG) as an active ingredient.

[0072] The hypersensitivity skin reaction is preferably selected from the group consisting of solar dermatitis, contact dermatitis and atopic dermatitis.

[0073] Based on the total weight of the composition, the active ingredient in the cosmetic composition of the present disclosure may be included in an amount of 0.001 to 1 wt %, preferably 0.01 to 0.1 wt %, and more preferably 0.05 to 0.5 wt %. Within this content range, appropriate formulation stability can be ensured, and a desired antioxidant effect can be expected.

[0074] In addition to the active ingredients of the present disclosure, the composition of the present disclosure may further include known natural extracts or other ingredients for anti-oxidation, skin aging prevention and skin regeneration promotion effects within the range that does not inhibit the effective activity of the active ingredients of the present disclosure.

[0075] In addition, the cosmetic composition may include components conventionally added to cosmetic compositions, such as fatty substances, organic solvents, solubilizers, thickeners, gelling agents, emollients, antioxidants, suspending agents, stabilizers, foaming agents, fragrances, surfactants, water, ionic or nonionic emulsifiers, fillers, masking agents, chelating agents, preservatives, vitamins, blocking agents, moisturizers, essential oils, dyes, pigments, and hydrophilic or hydrophobic activators, and may be formulated in any form using additives or carriers conventionally used in the cosmetic field or dermatology field.

[0076] The cosmetic composition of the present disclosure can be prepared as any preparation commonly prepared in the art, and for example, it can be formulated into a solution, suspension, emulsion, paste, gel, cream, lotion, powder, pack, soap, surfactant-containing skin cleanser, oil, spray, etc., but is not limited thereto. More specifically, the cosmetic composition can be prepared into a preparation such as a skin softening lotion, a nourishing lotion, a nourishing cream, a massage cream, an essence, an eye cream, a foundation, an emulsion foundation, a wax foundation, a cleansing cream, a cleansing foam, a cleansing water, a mask, a spray, a powder, a pact, a lip gloss, a lipstick, an eye shadow, a shampoo, or a conditioner.

[0077] When the preparation of the present disclosure is a paste, cream or gel, the carrier components that can be used are animal oil, vegetable oil, wax, paraffin, starch, tragacanth, cellulose derivatives, polyethylene glycol, silicone, bentonite, silicon dioxide, talc, zinc oxide and the like.

[0078] When the preparation of the present disclosure is a powder or a spray, the carrier component that can be used is lactose, talc, silicon dioxide, aluminum hydroxide, calcium silicate or polyamide powder, and in particular, when the preparation is a spray, it may further contain a propellant such as chlorofluorocarbon, propane / butane or dimethyl ether.

[0079] When the preparation of the present disclosure is a solution or emulsion, the carrier component that can be used is a solvent, a solubilizer or an emulsifier, examples of which include water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol oil, glycerol fatty acid ester, polyethylene glycol or sorbitan fatty acid ester.

[0080] When the preparation of the present disclosure is a suspension, the carrier component that can be used is a liquid diluent such as water, ethanol or propylene glycol, a suspending agent such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitan ester or polyoxyethylene sorbitan ester, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar, tragacanth gum, etc.

[0081] When the preparation of the present disclosure is a detergent containing a surfactant, the carrier components that can be used are fatty alcohol sulfates, fatty alcohol ether sulfates, sulfosuccinic acid monoesters, isethionates, imidazolinium derivatives, methyl taurate, sarcosinates, fatty acid amide ether sulfates, alkylamide betaines, fatty alcohols, fatty acid glycerides, fatty acid diethanolamides, vegetable oils, lanolin derivatives, ethoxylated glyceryl fatty acid esters, and the like.

[0082] The present disclosure also provides a reagent composition containing L-AHG for inhibiting T cell activation, proliferation or differentiation.

[0083] The T cells are preferably CD4 T cells.

[0084] Activation refers to the activation of resting T cells.

[0085] Proliferation refers to the increase in T cell number through cell cycle progression.

[0086] Differentiation refers to differentiation into Th1 cells and Th2 cells.

[0087] The reagent composition can be applied to T cells under ex vivo conditions, preferably in vitro conditions, to inhibit the activation, proliferation and differentiation of T cells.

[0088] The present disclosure also provides a method for inhibiting the activation, proliferation or differentiation of T cells, preferably CD4 T cells, comprising treating the T cells with L-AHG of the present disclosure under ex vivo conditions.

[0089] The term "ex vivo conditions" means that T cells exist in a state completely physically isolated from a living body. Ex vivo conditions are preferably in vitro conditions.

[0090] The T cells may be cells isolated from the blood of a subject, a subculture of cells, or a cell line.

[0091] L-AHG treatment can be performed, for example, by adding L-AHG to a culture medium containing T cells in a cell culture dish, and the treatment concentration is 1.0 to 1,000 μg / mL, preferably 12.5 to 400 μg / mL.

[0092] The inhibition of differentiation may be inhibition of differentiation into Th1 and Th2 cells. The inhibition of differentiation into Th1 and Th2 cells may result in reduced secretion of interferon-γ (INF-γ), interleukin-2 (IL-2), interleukin-4 (IL-4) or interleukin-13 (IL-13).

[0093] Hereinafter, the present disclosure will be described in more detail with reference to specific examples. The following examples describe preferred embodiments of the present disclosure, and the scope of the present disclosure should not be construed as being limited by the contents described in the following examples.

[0094] [Example]

[0095] 1. Materials and Methods

[0096] 1.1. Animals

[0097] C57BL / 6J male mice aged 5–6 weeks were purchased from OrientBio (322 Galmachi-ro, Jungwon-gu, Seongnam-si, Gyeonggi-do, South Korea) and used in the experiments and were maintained under specific pathogen-free conditions at the Kyungpook National University-Laboratory Animal Resources Center (680 Gukchaebosang-ro, Jung-gu, Daegu-si, South Korea).

[0098] 1.2. Chemicals, antibodies, reagents, and culture media

[0099] Mouse recombinant IL-2 (rIL-2) was purchased from Cambridge Bioscience (Cambridge, UK). Rabbit anti-mouse IgG was purchased from Jackson ImmunoResearch (West Grove, PA, USA), and anti-CD3 was purchased from BD Pharmingen (San Diego, CA, USA). Goat anti-mouse IL-2 antibody for neutralizing IL-2 activity was purchased from R&D System (Minneapolis, MN, USA), and rat anti-mouse IL-2R neutralizing antibody was purchased from Abcam (Cambridge, UK). ECL Western Blotting Kit was purchased from Thermo Fisher Scientific (Rockford, IL, USA). Immobilon-P membrane for Western blot analysis was purchased from EMD Millipore Corporation (Temecula, CA, USA). Microtubule depolymerizing agent nocodazole and all other electrophoresis reagents were purchased from Sigma Chemical (St. Louis, MO, USA).

[0100] In order to obtain the hydrolyzed products of agarose, neo-agarotetrose (NA4) and neo-agarothexaose (NA6), 9.0% agarose dissolved in water by heat treatment was hydrolyzed with GH16Bβ-agarase (1.6 μg / mL) under the optimal reaction conditions (1 mM MnCl2 and 10 mM TCEP, 50 mM Tris-HCl, pH 7.0, 50°C) for 14 hours while continuously stirring with a magnetic stirrer, thereby producing NA4 and NA6. The resulting NA4 / NA6 mixture was purified by size exclusion chromatography fractionation using a Sephadex G-15 column. In order to obtain new agarobiose (NA2) from agarose, 9% agarose was first hydrolyzed into new agarooligosaccharides (NAOS) including NA4 / NA6 by treating with GH16B β-agarase at 50°C, and then the temperature was lowered to 35°C, and then GH50A β-agarase (20 μg / mL) was added thereto and reacted for 14 hours to produce NA2. NA2 in the reaction product was purified by size exclusion chromatography fractionation using a Sephadex G-15 column. In addition, in order to obtain the constituent monomer L-AHG of agarose, 9% agarose was first hydrolyzed into NAOS including NA4 and NA6 by treating with GH16B β-agarase at 50°C, and then the temperature was lowered to 35°C, and then GH50A β-agarase (20 μg / mL) and GH117A α-NABH (60 μg / mL) were added thereto simultaneously and reacted for 14 hours, thereby obtaining a mixture of monomer L-AHG and D-galactose as the final product. In order to finally purify L-AHG from the mixture, the mixture was fractionated by size exclusion chromatography using a Sephadex G-10 column.

[0101] The medium used for T lymphocyte activation and culture was RPMI 1640 containing 10% FBS (HyClone, Logan Utah, USA), 20 mM HEPES, 50 μM β-mercaptoethanol, and 100 μg / mL gentamicin, which was designated as complete medium.

[0102] 1.3. Resting (G0) T cells 3 Isolation, activation and measurement of [H]thymidine incorporation

[0103] Isolation of resting (G0) T lymphocytes from spleens of 4-5 month old C57BL / 6J male mice was performed from spleen cells by negative selection using a T cell purification column kit (R&D Systems, Minneapolis, Minn., USA). At this time, activation of G0 T cells was performed using a modification of the method of Kim et al., wherein the surface of the culture plastic container was smoothly coated with anti-CD3 (2.0 μg / mL) and anti-CD28 (2.0 μg / mL) after pretreatment with rabbit anti-hamster IgG antibody (20 μg / mL).

[0104] To evaluate the proliferation of T cells activated by immobilized anti-CD3 / anti-CD28, 1×10 5 G0T cells and various concentrations of L-AHG were added to each well of a 96-well plate coated with anti-CD3 (2.0 μg / mL) and anti-CD28 (2.0 μg / mL) together with 200 μL of RPMI 1640 complete medium, and cell activation was initiated by culture. 29 hours after activation, 0.25 μCi of [ 3 H]-thymidine, and the incorporation into the DNA of proliferating T cells was quantified by liquid scintillation counting while the cells were cultured for an additional 12 h. 3 H]-thymidine amount.

[0105] For analysis of the cell cycle of immobilized anti-CD3 / anti-CD28 activated T cells or for analysis of cytokine concentrations in culture supernatants, 3 × 10 6 G0 T cells and L-AHG at various concentrations were added to each well of a 35-mm well plate coated with anti-CD3 (2.0 μg / mL) and anti-CD28 (2.0 μg / mL) together with 2 mL of RPMI 1640 complete medium, and cell activation was initiated by culture. 30 hours after activation, culture was stopped, and cells and culture supernatant were recovered by centrifugation.

[0106] Under the same culture conditions, in order to block the first cell cycle process initiated by G0 T cell activation at prometaphase by nocodazole treatment, T lymphocytes were activated for 20 hours, and then nocodazole was added thereto at a concentration of 0.2 μg / mL, followed by further culturing for an additional 10 hours. After completion of the culture, the culture was centrifuged to recover the cells and the culture supernatant.

[0107] 1.4. Stain the T cell surface with fluorescently labeled anti-CD4 / anti-CD8 antibodies and stain the intracellular DNA with DAPI

[0108] Resting (G0) T cells isolated from mouse splenocytes using a T cell purification column kit (R&D) and T cells obtained by activating G0 T cells with immobilized anti-CD3 / anti-CD28 for a given period of time were subjected to anti-CD3, anti-CD4, anti-CD8 and DAPI staining.

[0109] About 5×10 5 Each cell was suspended in 100 μL of 1X PBS / 1% FBS solution, and then 0.5 μg of anti-CD3, 0.5 μg of anti-CD4 and 0.5 μg of anti-CD8 were added thereto, and reacted at 4°C for 15 minutes, and then washed twice with 1X PBS / 1% FBS solution. Subsequently, the cells were fixed with 1% paraformaldehyde solution at 4°C for 15 minutes, and the fixed cells were washed twice with 1X PBS solution, and then stained with 10 μM DAPI for 15 minutes. The stained cells were analyzed with an Attune NxT flow cytometer (Thermo Fisher Scientific, Waltham, MA, USA) to determine the proportion of CD3 T cells, CD4 T cells and CD8 T cells and the cell cycle distribution based on the increase of blue fluorescence (depending on the amount of DAPI bound to intracellular DNA).

[0110] 1.5. Enzyme-linked immunosorbent assay (ELISA)

[0111] After recovering the culture supernatant of T cells, the concentrations of IFN-γ, IL-2, IL-4 and IL-13 were analyzed using an ELISA kit (Mouse Uncoated Elisa Kit, Invitrogen) according to the manufacturer's instructions. In short, 100 μl of capture antibody dilution was first added to each well of a 96-well plate and incubated at 4 ° C for 16 hours, followed by washing three times with a washing solution. Each well was blocked at 25 ° C for 1 hour with 200 μl of blocking solution and washed once with a washing solution. Subsequently, 100 μl of a solution (wherein the standard sample and culture supernatant were appropriately diluted) was added to each well and reacted at 25 ° C for 2 hours. After washing three times with a washing solution, 100 μl of a second antibody (detection antibody) solution was added to each well and reacted at 25 ° C for 1 hour. After washing three times with a washing solution, 100 μl of avidin-HRP solution was added to each well and reacted at 25 ° C for 30 minutes. After washing 5 times with washing solution, add 100 μ l of substrate solution to each well, and react at 25 ℃ for 15 minutes under shading conditions. The reaction is terminated by adding 100 μ l of reaction stop solution to each well, and the absorbance at 450nm and 570nm wavelengths is measured with Varioskan Lux Multimode microplate reader (Thermo Fisher Scientific). At this time, the value obtained by deducting the absorbance measured at 570nm wavelength from the absorbance measured at 450nm wavelength is compared with the standard curve to determine the concentration of the cytokine present in the culture supernatant. In order to prepare the standard curve, serially diluted recombinant IFN-γ, IL-2, IL-4 and IL-13 are used as standard samples.

[0112] 2. Results

[0113] 2.1. Effect of enzymatic hydrolysis products of agarose on G0 T cells stimulated by immobilized anti-CD3 / anti-CD28 3 Effect of [H]-thymidine incorporation

[0114] Resting (G0) lymphocytes were isolated from mouse spleens by Percoll gradient centrifugation. G0 T lymphocytes were isolated from G0 lymphocytes by negative selection using a T cell purification column kit (R&D Systems, Minneapolis, MN, USA). The surface of G0 T cells isolated by T cell purification columns was stained with fluorescently labeled anti-CD3, anti-CD4, and anti-CD8 antibodies, and 96% of the cells were identified as CD3+ T cells ( Figure 1 In addition, 50.6% of these CD3+ T cells were identified as CD4+ T (T H) cells, and 43.8% were identified as CD8+T (T C )cell.

[0115] Isolated resting T cells were stimulated in 96-well plates and stimulated with immobilized anti-CD3 / CD28. At this time, NA4 and NA6, each of which was obtained by hydrolyzing agarose with GH16Bβ-agarase, NA2, which was obtained by first hydrolyzing agarose with GH16Bβ-agarase and then further hydrolyzing with GH50Bβ-agarase, and L-AHG, which was obtained by first hydrolyzing agarose with GH16Bβ-agarase and then further hydrolyzing with both GH50Bβ-agarase and GH117Aα-NABH, were added to cells at concentrations of 12.5 to 400 μg / mL to study the effects of these agarose degradation products (L-AHG, NA2, NA4, and NA6) on cell cycle progression and proliferation after activation of resting T cells.

[0116] First, resting T cells (1×10 4 T cells) were stimulated and activated with immobilized anti-CD3 / anti-CD28 in the presence of different concentrations (0, 12.5, 25, 50, 100, 200, and 400 μg / mL) of L-AHG, NA2, NA4, or NA6 using a 96-well plate. 5 G0 T cells / well). 29 hours after activation, 0.25 μCi of [ 3 H] thymidine was then further cultured for an additional 12 hours. The incorporation of [H] thymidine into DNA following T cell proliferation during this additional culture period was measured using a liquid scintillation counter. 3 The amount of [H] thymidine was quantified to determine the extent of T cell proliferation that occurred in each well.

[0117] Results, such as Figure 2 As shown, it was confirmed that 12.5 μg / ml of L-AHG had no significant effect on the proliferation of activated T cells, and thus at this L-AHG concentration [ 3 The level of [H] thymidine incorporation was almost the same as that of the control group without L-AHG treatment. In contrast, the presence of 25 to 400 μg / mL L-AHG inhibited [ 3 H] thymidine incorporation. In particular, the results showed that in the presence of 100 μg / mL of L-AHG, [ 3 H] thymidine incorporation was reduced to approximately 22% of that in the control group without L-AHG treatment, and at L-AHG concentrations of 200 to 400 μg / mL, [ 3H] thymidine incorporation was significantly reduced to unmeasurable levels. However, the results showed that under the same conditions, agarose degradation products such as NA2, NA4 and NA6 had an inhibitory effect on activated T cells at concentrations of 25 to 400 μg / mL. 3 H] Thymidine incorporation had no significant effect.

[0118] These results demonstrate that, among the enzymatic hydrolysates of agarose studied, only L-AHG has a significant physiological activity of inhibiting the proliferation of activated T cells.

[0119] Until recently, various physiological activities of agar degradation products (NA2, L-AHG, NAOS and AOS) have been reported. That is, these physiological activities include antioxidant, prebiotic, antitumor, anti-inflammatory, antidiabetic, anti-obesity and skin moisturizing and whitening activities. In addition, it has been reported that oral administration of NA4 in a mouse model showed a protective effect on fatigue and liver damage caused by strenuous exercise by regulating the intestinal microflora. However, the inhibitory effect of L-AHG on the proliferation of activated T cells is a new physiological activity that has not been reported so far.

[0120] Effect of L-AHG on cell cycle progression of G0 T cells stimulated by immobilized anti-CD3 / anti-CD28

[0121] The T lymphocytes of the immune system can be roughly divided into two types: CD4 T (T H ) lymphocytes and CD8 T (T C ) lymphocytes. The inventors of the present disclosure examined the 3 H] thymidine incorporation assay confirmed the inhibition of DNA synthesis in activated T cells. Will the proliferation inhibitory activity of L-AHG be exerted in CD4 T (T H ) cells and CD8 T (T C ) cells, or whether it occurs only in either of the two types of T cells.

[0122] For this reason, resting T cells are stimulated and activated with immobilized anti-CD3 / anti-CD28, and when these cells reach the time point of S phase starting after G0 exit and G1 entry and progress, that is, after 20 hours after activation, microtubule depolymerizing agent nocodazole (a cell cycle blocker, which blocks the cell cycle process of eukaryotic cells in the prometaphase of M phase) is added to the cells, and then further cultured for 10 hours. Through this unique experimental method, the first cell cycle started after resting T cells are activated passes through G1, S and G2 phases, and then blocked in the prometaphase of M phase due to microtubule damage caused by the action of nocodazole. Then, these cells are recovered, processed and stained with fluorescently labeled anti-CD4 antibodies, fluorescently labeled anti-CD8 antibodies and DNA fluorescent staining reagent DAPI, and analyzed with flow cytometer. Therefore, the inventors have studied how L-AHG affects the first cell cycle process (G1, S and G2 / M phase) started after resting CD4 T cells and resting CD8 T cells are activated.

[0123] like Figure 3 A and Figure 3 As shown in Figure B, as a result of analyzing the stained cells by flow cytometry, it is shown that when resting T cells are stimulated and activated with immobilized anti-CD / anti-CD28, 4.4%, 8.8% and 77.9% of CD8 T cells are distributed in G1, S and G2 / M phases, respectively, while 9.9%, 35.4% and 49.9% of CD4 T cells are distributed in G1, S and G2 / M phases, respectively. That is, 30 hours after activation, the percentage of CD8 T cells remaining in the G1+S phase without entering the G2 / M phase is only about 13.2%, but the percentage of CD4 T cells remaining in the G1+S phase is as high as about 45.3%. These results show that among resting T cells, CD8 T cells pass through the cell cycle process relatively faster and more smoothly than CD4 T cells after activation with immobilized anti-CD / anti-CD28.

[0124] It was found that the first cell cycle progression after activation of resting CD4 T and CD8 T cells identified as described above was significantly differently affected by L-AHG added at concentrations of 25 to 100 μg / ml. That is, after activation with immobilized anti-CD3 / anti-CD28, the percentage of resting CD4 T cells that remained in the G1 and S phases without entering the G2 / M phase increased in a manner dependent on the L-AHG concentration. In particular, the results showed that when 100 μg / mL of L-AHG was added, the percentages of cells in the G1 and S phases were 35.4% and 54.6%, respectively, while the percentage of cells in the G2 / M phase was only 8.8%, indicating that L-AHG significantly inhibited cell cycle progression. In contrast, cell cycle progression of resting CD8 T cells after activation was barely affected by the addition of 25 to 50 μg / mL of L-AHG, and when 100 μg / mL of L-AHG was added, the percentage of cells in the S phase increased to 46.6%, and the percentage of cells in G2 / M was 41.8%, while the change in the percentage of cells in the G1 phase was negligible.

[0125] These findings indicate that by adding L-AHG (25 to 100 μg / mL), the cell cycle progression of resting CD4 T cells after activation was inhibited mainly in the G1 and S phases in a concentration-dependent manner, and in particular, when 100 μg / ml of L-AHG was added, the percentage of cells remaining in the G1+S phase without entering the G2 / M phase was about 90.0% of the total cells. Under the same conditions, the addition of 25 to 50 μg / mL of L-AHG had no significant effect on the cell cycle progression of resting CD8 T cells after activation, and even when 100 μg / mL of L-AHG was added, the progression of the G1 phase was not affected, but the progression of the S phase was inhibited to a certain extent. Therefore, it has been confirmed that the presence of L-AHG (25-100 μg / mL) inhibits the cell cycle progression of resting CD4 T and CD8 T cells after activation in different ways, but the death of apoptotic cells in these cells (based on the proportion of apoptotic sub-G1 cells) is not induced by L-AHG.

[0126] At the same time, the results showed that when resting T cells were stimulated and activated with immobilized anti-CD3 / anti-CD28 under the same conditions and cultured for 30 hours without the addition of nocodazole, the inhibitory effect of L-AHG on the cell cycle progression of resting CD4 T and CD8 T cells after activation was not easily confirmed by flow cytometry ( Figure 4 A and Figure 4B). This is considered to be because the cell cycle progression initiated by the activation of resting CD4 T cells or resting CD8 T cells is inhibited by the presence of L-AHG in some cells but not in other cells, and these other cells progress to the G1, S, and G2 phases after completing the first cell cycle progression, and eventually, cells whose cell cycle progression is inhibited by L-AHG and cells that progress through the cell cycle even in the presence of L-AHG are mixed together and coexist.

[0127] In summary, the results of this study indicate that the inhibitory effect of L-AHG on the proliferation of activated T cells is mainly directed at CD4 T cells, rather than CD8 T cells, and inhibits the cell cycle progression of CD4 T cells in the G1 and S phases.

[0128] Effect of L-AHG on cytokine production by G0 T cells stimulated by immobilized anti-CD3 / anti-CD28

[0129] Unlike CD8 T cells, CD4 T cells differentiate into several functionally differentiated effector CD4 T cell subtypes and regulate various immune responses. Th1, Th2, Th17, Tfh, and Treg cells are known as effector CD4 T cell subtypes. These subtypes can be distinguished from each other based on the characteristics of the cytokine combinations they secrete.

[0130] Th1 cells produce IFN-γ and promote the activation of macrophages, so that microorganisms that can survive or reproduce in macrophages can be destroyed more effectively. Th2 cells participate in the recruitment and activation of eosinophils (IL-5) and mast cells / basophils (IL-4) by producing cytokines such as IL-4, IL-5 and IL-13, and also enhance the barrier immunity of mucosal surfaces (IL-13). They mainly promote defense reactions through IgE to help control parasitic infections. In particular, B cell class conversion also requires IL-4 and IL-13 to produce IgE. B cells may cause hypersensitivity reactions such as allergies and asthma by means of the IgE produced by Th2 cells. Th17 cells secrete IL-17A, IL17F and IL-22, induce local epithelial cells and stromal cells to produce chemokines, which recruit neutrophils to the site of infection. Therefore, they enhance the reaction of neutrophils to remove pathogens such as extracellular bacteria and fungi. In addition, they activate mucosal epithelial tissue and skin epithelial cells to produce antimicrobial peptides that attack bacteria. FH T cells are an effector CD4 T cell subtype that favor B cells in lymphoid tissues. FHT cells interact with naive B cells through mechanisms such as trafficking to B cell follicles and antigen recognition, and promote germinal center responses, contributing to antibody production and class switching of B cells. FH The cells produce large amounts of the cytokine IL-21, which supports B cell proliferation and its differentiation into antibody-producing plasma cells. Treg cells normally suppress T cell activity and innate immune cell activity by producing TGF-β and IL-10, thereby limiting the immune response and helping to prevent autoimmunity that develops during the immune response.

[0131] The stability of effector CD4 T cell subtypes is different from each other. Initial CD4 T cells have various potentials, while Th1 and Th2 cells show a relatively stable basal state and are highly resistant to conversion to other subtypes. At the same time, iTreg cells and Th17 cells are less stable and can be converted into other subtypes by the cytokines that mainly affect them. iTreg cells can be converted into Th17 cells in response to IL-6 and IL-1, and converted into Th1 cells in response to IL-12. Th17 cells can also be converted into Th1 cells in response to IL-12. Here, it should be understood that the conversion of iTreg cells to Th17 cells and the conversion of Th17 cells to Th1 cells are unidirectional or irreversible.

[0132] Based on the immune-related functions of effector CD4 T cell subtype differentiation, it would be more effective to use immunosuppressive agents with the ability to target Th1 and Th2 to exert specific inhibitory effects in effector CD4 T cell subtypes to control inappropriate immune responses in patients with hypersensitivity reactions, autoimmune diseases, and organ transplant patients (including stem cell or bone marrow transplant patients) to necessary levels.

[0133] The results showed that L-AHG (25 to 100 μg / mL) significantly inhibited the proliferation of activated CD4 T cells by inhibiting the cell cycle progression of activated CD4 T cells mainly in the G1 and S phases in a concentration-dependent manner.

[0134] Under the same conditions, resting T cells were activated by stimulating with immobilized anti-CD3 / anti-CD28 for 30 hours, and the culture supernatant was recovered and the concentrations of Th1 signature cytokines (IFN-γ and IL-2) known to be secreted by Th1 cells and Th2 signature cytokines (L-4 and IL-13) known to be secreted by Th2 cells were measured by ELISA.

[0135] The results showed that the culture supernatant of activated T cells contained not only IL-2 and IFN-γ secreted by the Th1 subtype, but also IL-4 and IL-13 secreted by the Th2 subtype, which were significantly reduced in a concentration-dependent manner by adding L-AHG (25, 50 and 100 μg / mL) ( Figure 5 A and B). Interestingly, a more pronounced decrease in the concentrations of Th2 signature cytokines was observed compared to the concentrations of Th1 signature cytokines. In particular, it was demonstrated that in the presence of 25 μg / mL of L-AHG, IFN-γ concentrations were barely affected, but IL-4 concentrations were significantly reduced to 38.5% of the concentration in the L-AHG untreated control group.

[0136] In conclusion, the above results indicate that when clonal expansion after cell proliferation and differentiation into effector Th1 and Th2 subtypes occur as a result of activating resting Th cells with immobilized anti-CD3 / anti-CD28 stimulation to initiate the cell cycle, the presence of L-AHG can inhibit differentiation into Th1 and Th2 subtypes by inhibiting the cell cycle progression of CD4 T cells, and can inhibit differentiation into Th2 subtype more strongly than Th1 subtype.

[0137] [Preservation of Microorganisms]

[0138] Depository: Korean Collection for Type Cultures (KCTC), Korea Research Institute of Bioscience and Biotechnology

[0139] Accession number: KCTC13629BP

[0140] Date of deposit: August 27, 2018

[0141] Budapest Treaty International Forms Relating to the International Recognition of the Deposit of Microorganisms in Patent Procedure

[0142] Deposit Certificate for Original Deposit under Rule 7.1

[0143] To: Kim Young-ho

[0144] Kyungpook National University, 80 Daehak-ro, Buk-gu, Daegu, South Korea

[0145]

[0146]

[0147] BP / 4 form (KCTC 17 form) Single page.

Claims

1. An immunosuppressive composition comprising 3,6-anhydro-L-galactose (L-AHG) as an active ingredient.

2. The immunosuppressive composition according to claim 1, which inhibits the activation, proliferation or differentiation of CD4 T cells.

3. The immunosuppressive composition of claim 2, which blocks cell cycle progression in the G1 or S phase. The immunosuppressive composition according to claim 1 , which inhibits differentiation into Th1 cells or Th2 cells.

5. The immunosuppressive composition according to claim 4, which inhibits the production of interferon-γ (INF-γ), interleukin-2 (IL-2), interleukin-4 (IL-4) or interleukin-13 (IL-13).

6. A pharmaceutical composition for preventing or treating an immune disease, comprising 3,6-anhydro-L-galactose (L-AHG) as an active ingredient.

7. The pharmaceutical composition according to claim 6, wherein The immune disease is selected from the group consisting of: hypersensitivity disease, autoimmune disease, immune rejection, graft-versus-host disease, Sjögren's syndrome and Behcet's disease.

8. The pharmaceutical composition according to claim 7, wherein The hypersensitivity disease is selected from the group consisting of allergy, sneezing, wheezing, asthma, atopic dermatitis and urticaria.

9. The pharmaceutical composition according to claim 7, wherein The autoimmune disease is selected from the group consisting of multiple sclerosis, rheumatoid arthritis, systemic lupus erythematosus, type 1 diabetes, psoriasis, Hashimoto's thyroiditis, systemic lupus erythematosus and acute inflammatory bowel disease.

10. A health functional food for preventing or improving immune diseases, comprising 3,6-anhydro-L-galactose (L-AHG) as an active ingredient.

11. The health functional food according to claim 10, wherein The immune disease is selected from the group consisting of: hypersensitivity disease, autoimmune disease, immune rejection, graft-versus-host disease, Sjögren's syndrome and Behcet's disease.

12. The health functional food according to claim 11, wherein The hypersensitivity disease is selected from the group consisting of allergy, sneezing, wheezing, asthma, atopic dermatitis and urticaria.

13. The health functional food according to claim 11, wherein The autoimmune disease is selected from the group consisting of multiple sclerosis, rheumatoid arthritis, systemic lupus erythematosus, type 1 diabetes, psoriasis, Hashimoto's thyroiditis, systemic lupus erythematosus and acute inflammatory bowel disease.

14. A functional cosmetic composition for preventing or improving hypersensitivity skin reactions, comprising 3,6-anhydro-L-galactose (L-AHG) as an active ingredient.

15. The functional cosmetic composition according to claim 14, wherein The hypersensitivity skin reaction is selected from the group consisting of solar dermatitis, contact dermatitis and atopic dermatitis.

Images