JP2005082491A - Immunosuppresant - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To obtain an immunosuppresant which is safe and excellent and can widely be used for foods, medicines, quasi-drugs, medical care devices, feeds, animal medicines, pet foods, and the like. <P>SOLUTION: This immunosuppresant is characterized by containing an acidic mucopolysaccharide and/or its derivative as an active ingredient. The acidic mucopolysaccharide is preferably a chondroitin sulfate, especially chondroitin sulfate A, chondroitin sulfate C, chondroitin sulfate D, chondroitin sulfate E, and/or their derivatives. The immunosuppresant is added to foods, medicines, quasi-drugs, medical care devices, feeds, animal drugs, pet foods, and the like. <P>COPYRIGHT: (C)2005,JPO&NCIPI

Description

  The present invention relates to an immunoregulator having an immunoregulatory function by oral, transdermal, intravenous injection or vaginal, containing chondroitin sulfate as an active ingredient.

With the advance of medicine, there is a marked increase in diseases caused by reduced immune function or impaired immune function associated with advanced treatments. On the other hand, patients with allergies such as atopic dermatitis, hay fever, and food ingredients are steadily increasing, which is a serious social problem.

In the field of pharmaceuticals, corticosteroids and interferons are used as immunoregulators as therapeutic agents for diseases such as autoimmune diseases and multiple myeloma. It is the present condition that it is accompanied.

  In the field of foods, the search for components having an immune control function has been conducted extensively, and it has been reported that tea, mushrooms, vegetables, fruits, fish oil, etc. have an immune control function, but the mechanism is clear. There are many things that are not.

  On the other hand, chondroitin sulfate binds to a serine residue via a bridging structure of two galactose molecules and one xylose molecule in vivo to form a complex with a protein, proteoglycan, and collagen, It is involved in the composition of extracellular matrix together with elastin, hyaluronic acid, etc., and is known to exist in various organs and body fluids such as cartilage, bone, cornea, cervical ligament, tendon, umbilical cord, skin, urine, blood .

  Sakai et al. Have already found that chondroitin sulfate has an immunoregulatory function (see Non-patent Document 1), but the relationship between the structure of chondroitin sulfate and the immunoregulatory function is not mentioned. It has been pointed out that chondroitin sulfate has various physiological activities depending on the chain length, the number of sulfone group bonds, and the sulfone group binding site, but the physiological role and state of existence in vivo are still unclear. Kawashima et al. Also reported that chondroitin sulfate E binds to L-selectin, P-selectin, CD44, and chemokine as ligands (see Non-Patent Document 2).

  On the other hand, it is described that polysulfated mucopolysaccharides have functions as preventive or therapeutic agents for allergic diseases (see Patent Document 1) and kallikrein-kinin system inhibitors (see Patent Document 2).

JP 11-335288 A JP-A-11-147901 Sakai, S., Akiyama, H., Harikai, N., Toyoda, H., Toida, T., Maitani, T., Imanari, I., (2002) Immunol. Lett., 84, 211-216. Kawashima, H., Hirose, M., Hirose, J., Nagakubo, D., Plaas, A. H. K., and Miyasaka, M. (2000) J. Biol. Chem. 275, 35448-35456.

An object of the present invention is to provide an immunoregulator that can be widely used for foods, pharmaceuticals, quasi-drugs, medical devices, feedstuffs, veterinary drugs, pet foods, etc. is there.

  The present inventors have eagerly investigated the search for immunoregulators with weak side effects, and have studied in detail the position and number (structure) of the sulfone group of chondroitin sulfate, which is an acidic mucopolysaccharide. It has been found that it has a control function (activity) and has led to the present invention.

  According to the present invention, there is provided an immunoregulator that is selected from chondroitin sulfate A, chondroitin sulfate C, chondroitin sulfate D, chondroitin sulfate E, and / or derivatives thereof.

  Chondroitin sulfate which is a component of the present invention is a compound contained in vertebrates, annelids, arthropods, molluscs and the like. Or a compound produced by a microorganism.

Chondroitin sulfate is an acidic mucopolysaccharide composed of disaccharide units. Structure of chondroitin sulfate A is _{(→ 4GlcAβ1 → 3GalNAc4Sβ1 →) n} , the structure of chondroitin sulfate C is _{(→ 4GlcAβ1 → 3GalNAc6Sβ1 →) n} , the structure of chondroitin sulfate D of _{(→ 4GlcA2Sβ1 → 3GalNAc6Sβ1 →) n} , chondroitin sulfate E structure Is (→ 4GlcAβ1 → 3GalNAc4S6Sβ1 →) _n . Where GlcA is D-glucuronic acid, β is the binding mode, the numbers before and after the arrow are the bonding positions, GALNAc is N-acetyl-D-galactosamine, S is the sulfone group, the number before S is the number of sulfone groups, n Indicates the degree of polymerization.

  In recent years, it has become clear that it is important to return the balance of helper T cell Th1 cells and helper T cell Th2 cells to normal for the treatment of allergic reactions. That is, cellular immunity decreases when Th2 cells become dominant over Th1 cells, and cellular immunity increases when Th1 cells become dominant. We examined the production of chondroitin sulfate and these derivatives by producing Th1-type cytokines (interferon γ, interleukin 2) and Th2-type cytokines (interleukin 5, interleukin 10). As a result, it has been found that chondroitin sulfate A, chondroitin sulfate C, chondroitin sulfate D, chondroitin sulfate E and derivatives have an immunoregulatory function that promotes Th1-type cytokine production and suppresses Th2-type cytokine production.

  The chondroitin sulfate component used as an active ingredient of the immunoregulator in the present invention may be any one that contains chondroitin sulfate A, chondroitin sulfate C, chondroitin sulfate D, chondroitin sulfate E, and derivatives thereof, Those directly extracted / separated / purified from animals, those produced by microorganisms, separated / purified, or chemically synthesized derivatives thereof can be used.

  The diluent or carrier of the immunoregulator of the present invention is not particularly limited as long as it does not interfere with the effects of the present invention. For example, sugars such as sucrose, glucose, dextrin, varicella, liquid sugar; ethanol, propylene glycol, Examples thereof include alcohols such as glycerin; sugar alcohols such as sorbitol and mannitol; polysaccharides such as gum arabic; or water. Examples of the additive include antioxidants, auxiliaries such as chelating agents, fragrances, spice extracts, preservatives, and the like.

  Specific products to which the immunoregulator of the present invention is applied are not particularly limited. For example, food (food and drink), cosmetics, pharmaceuticals, quasi drugs, medical supplies, feedstuffs, veterinary drugs, pets For food.

  Examples of foods targeted by the present invention include milk drinks, lactic acid bacteria drinks, soft drinks with fruit juice, soft drinks, carbonated drinks, fruit drinks, vegetable drinks, vegetable / fruit drinks, alcoholic drinks, powdered drinks, coffee drinks, and tea drinks. Beverages such as custard pudding, milk pudding, souffle pudding, pudding such as fruit pudding, desserts such as jelly, bavaroa and yogurt; ice cream, ice milk, lacto ice, milk ice cream, ice cream with fruit juice and soft Frozen confectionery such as cream, ice candy, sherbet, ice confectionery; gums such as chewing gum and bubble gum (plate gum, sugar-coated granule gum); coating chocolate such as marble chocolate, strawberry chocolate, blueberry chocolate and melon chocolate Add flavor Chocolate such as chocolate; hard candy (including bonbon, butterball, marble, etc.), soft candy (including caramel, nougat, gummy candy, marshmallow, etc.), caramel such as drop, toffee; hard biscuits, cookies, Baked confectionery such as oysters, rice crackers (and confectionery); soups such as consomme soup and potage soup; Pickles such as pickles, pickles, Fukujinzuke, Shiba pickles, ginger pickles, pickled plums; sauces such as separate dressing, non-oil dressing, ketchup, sauce, sauces; strawberry jam, blueberry jam, marmalade, apple jam, apricot jam, presabu Jams such as; Fruit syrups such as syrups, processed fruits such as cherries, apricots, apples, strawberries and peaches; processed meat products such as ham, sausage and grilled pork; fish ham, fish sausage, fish meat surimi, salmon, bamboo rings, hampen, Fish paste products such as fried Satsuma, Date roll, whale bacon; dairy and fat products such as butter, margarine, cheese, whipped cream; In addition, various prepared foods and various processed foods such as rice cakes and rice cakes can be mentioned. Beverages and confectionery are preferred.

  The food and drink of the present invention can be produced according to conventional production methods for various foods and drinks, except that the immunoregulator of the present invention is blended in any step of production. There is no restriction | limiting in particular in the compounding method and order of an immunoregulator.

  For example, in the case of frozen confectionery, first add the immunomodulating agent, acids, emulsifiers and stabilizers of the present invention to milk, cream, condensed milk, powdered milk, saccharides, fruits or strawberries as main ingredients, and then add fragrance and then add frozen confectionery. Examples include a method of preparing a mixed solution, adding a dye to the mixed solution, mixing, sterilizing, freezing after cooling, filling into a container, and cooling or freezing to prepare a final product.

  In the case of chewing gums, sugar, glucose, the immunoregulatory agent of the present invention, citric acid, etc. are added to a heated and softened gum base, and then fragrance and pigment are added and kneaded. A method of preparing a final product by making an appropriate thickness, cooling to room temperature, and cutting.

  In the case of desserts, the main ingredients of sugar, starch syrup, the immunoregulator of the present invention, citric acid and a coagulant (pectin, agar, gelatin, carrageenan, etc.) are mixed in an appropriate ratio, and flavors and A method of preparing a jelly that is a final product by adding a dye, dissolving by heating, filling in a container, and cooling can be mentioned. In the case of candies, for example, water is added to the main ingredients such as sugar and starch syrup, heated and dissolved, allowed to cool, then added with immunoregulators, then added with fragrances and pigments, molded, cooled to room temperature and finally A method for preparing candies can be mentioned.

  In addition, in the case of beverages, the immunity control agent or stabilizer of the present invention is added to sugars, fruit juices or acids as the main raw material, and then a fragrance and a pigment as necessary are added to the beverage and mixed, then sterilized, The method of cooling and filling a container can be mentioned.

  In the case of pickles, add the pickles by adding various seasonings such as salt and sugar, preservatives, and auxiliary ingredients such as the immune regulator of the present invention to the main ingredients such as vegetables, seaweed, mushrooms or fruits to be pickled. A method of preparing a final product by preparing and mixing the pickles with a fragrance and, if necessary, a coloring matter, filling the container, sterilizing and cooling can be mentioned. In the case of sauces and dressings, vegetable oil, soy sauce, fruit juice, saccharides, fruit juice, brewed vinegar or salt etc. are used as the main ingredients, and the immune regulator and stabilizer or emulsifier of the present invention are added thereto, and this dressing solution A method of preparing a final product by adding a fragrance and, if necessary, a dye and mixing the mixture and then sterilizing and cooling and then filling the container.

  Cosmetics targeted by the present invention include skin cosmetics (lotions, emulsions, creams, etc.), lipsticks, sunscreen cosmetics, makeup cosmetics, etc .; as pharmaceuticals, conventional immune regulators and anticancer agents as pharmaceutical compositions Oral agents such as tablets, granules, powders, capsules, syrups, drinks, etc., or suppositories, ointments, sprays, etc. Parenterals such as pills, injections, and haptics; quasi-drugs such as toothpastes, drinks, mouth fresheners, bad breath prevention agents, haps, ointments, soaps, face wash, creams, milky lotions, shampoos, Examples of feeds for rinsing include food for appreciation fish, food for aquaculture, livestock for livestock, feed for birds, feed for livestock, etc., but are not limited to these. No.

  These various products such as cosmetics, pharmaceuticals, quasi-drugs, and feedstuffs can be manufactured according to conventional methods of various products, except that the immunomodulator of the present invention is blended in any step of the manufacture. it can. There are no particular limitations on the timing of the immunoregulatory agent added to cosmetics, pharmaceuticals, quasi drugs or feed.

  The amount of the immunoregulatory agent of the present invention added to various products such as foods and drinks, cosmetics, pharmaceuticals, quasi drugs, and feed is not particularly limited. It can be appropriately selected and determined in consideration of the type and application of the object and the components contained therein.

  The present invention will be described in more detail based on examples, but the present invention is not limited to these examples.

Female mice BALB / c mice (7 weeks old) were sensitized with 20 μg of ovalbumin (OVA) together with 2 mg of aluminum hydroxide gel, followed by spleen cell suspension after secondary sensitization (5.0 × 10 ⁶ cells / mL) Was prepared. After stimulation with OVA in vitro, add chondroitin sulfate A (derived from bovine tracheal cartilage), chondroitin sulfate B (derived from porcine skin), chondroitin sulfate C (derived from shark cartilage) to a final concentration of 1 ppm / well, The cells were cultured under conditions of a culture temperature of 37 ° C. and a CO ₂ concentration of 5%, and various cytokines produced in the supernatant were quantified by ELISA.

Fig. 1 shows Th1-type cytokines, interferon γ (INF-γ), interleukin 2 (IL-2), and Th2 type, when chondroitin sulfate A, chondroitin sulfate B, and chondroitin sulfate C are added and cultured under the above conditions. The production ratio of cytokines interleukin 5 (IL-5) and interleukin 10 (IL-10) to the control is shown. FIG. 1 shows that Th1 type cytokine production is promoted when the Th1 type cytokine production ratio is increased, and Th2 type cytokine production is suppressed when the Th2 type cytokine production ratio is decreased. As shown in FIG. 1, chondroitin sulfate A and chondroitin sulfate C were found to promote Th1 type cytokine production and Th2 type cytokine production, but chondroitin sulfate B promoted Th1 type cytokine production and Th2 type cytokine production. Inhibition was not observed. That is, chondroitin sulfate A and chondroitin sulfate C showed an immune control function, but chondroitin sulfate B did not show an immune control function.

As in Example 1, female mice BALB / c mice (7 weeks old) were sensitized with 20 μg of ovalbumin (OVA) together with 2 mg of aluminum hydroxide gel, and after the second sensitization, the spleen cell suspension (5.0 × 10 ⁶ cells / mL) was prepared. After in vitro stimulation with OVA, chondroitin sulfate A (derived from bovine tracheal cartilage) with a different degree of sulfonation was added to a final concentration of 1 ppm / well, and the culture temperature was 37 ° C and the CO ₂ concentration was 5%. The various cytokines produced in the supernatant were quantified by ELISA. Chondroitin sulfates with different degrees of sulfonation were prepared using chemical methods.

In FIG. 2, pOS Chondroitin is chondroitin sulfate having 2 sulfone groups per disaccharide unit, fOS Chondroitin is chondroitin sulfate having 4 sulfone groups per disaccharide unit, and de OS Chondroitin has no sulfone group. Chondroitin sulfate is shown. pOS Chondroitin and fOS Chondroitin, like chondroitin sulfate A, were found to promote Th1-type cytokine production, but de OS Chondroitin, which has no sulfone group, promoted Th1-type cytokine production and Th2-type cytokine production. No inhibition was observed. That is, chondroitin sulfate with 2 sulfone groups per disaccharide unit and chondroitin sulfate with 4 sulfone groups per disaccharide unit were found to have immunoregulatory functions, but chondroitin sulfate without any sulfone groups had immune control functions. The function was not recognized.

In the same manner as in Example 1, female mice BALB / c mice (7 weeks old) were sensitized with 20 μg of ovalbumin (OVA) together with 2 mg of aluminum hydroxide gel and spleen cell suspension (5.0% after secondary sensitization). × 10 ⁶ cells / mL) was prepared. After in vitro stimulation with OVA, chondroitin sulfate D (derived from shark cartilage) and chondroitin sulfate E (derived from shark cartilage) are natural product-derived chondroitin sulfate having two sulfone groups per disaccharide unit so that the final concentration is 1 ppm / well Squid cartilage derived) was added, cultured under conditions of a culture temperature of 37 ° C. and a CO ₂ concentration of 5%, and various cytokines produced in the supernatant were quantified by ELISA.

As a result, as shown in FIG. 3, similar to chondroitin sulfate A, chondroitin sulfate D and chondroitin sulfate E were found to promote Th1-type cytokine production and suppress Th2-type cytokine production. That is, chondroitin sulfate D and chondroitin sulfate E having two sulfone groups per disaccharide unit were found to have an immunoregulatory function.

According to the present invention, chondroitin sulfate can be widely used as a safe and excellent immunoregulator for pharmaceuticals, foods, quasi drugs, medical devices, feedstuffs, veterinary drugs, pet foods, and the like.

Th1 type cytokines IFN-γ (interferon gamma), IL-2 (interleukin 2), Th2 type cytokines IL-5 (interleukin 5), IL-10 (inter) in spleen cells derived from OVA immunized mice The effects of chondroitin sulfate A, chondroitin sulfate B, and chondroitin sulfate C on leukin 10) production were observed.

Th1 type cytokines IFN-γ (interferon gamma), IL-2 (interleukin 2), Th2 type cytokines IL-5 (interleukin 5), IL-10 (inter) in spleen cells derived from OVA immunized mice The effect of the number of sulfone groups on leukin 10 production was examined.

Th1 type cytokines IFN-γ (interferon gamma), IL-2 (interleukin 2), Th2 type cytokines IL-5 (interleukin 5), IL-10 (inter) in spleen cells derived from OVA immunized mice The effects of chondroitin sulfate A, chondroitin sulfate D, and chondroitin sulfate E on the production of leukin 10) were observed.

Claims (4)

An immunoregulator comprising an acidic mucopolysaccharide and / or a derivative thereof as an active ingredient. The immunoregulator according to claim 1, wherein the acidic mucopolysaccharide is chondroitin sulfate. The immunoregulator according to claim 1 or 2, wherein the derivative according to claim 1 has 2 or 4 sulfone groups per chondroitin sulfate disaccharide unit. The immunoregulator according to claim 2 or 3, wherein the chondroitin sulfate is at least one selected from chondroitin sulfate A, chondroitin sulfate C, chondroitin sulfate D, and chondroitin sulfate E.

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