Agent for preventing or improving skin damage

Abstract

To find a new target molecule of skin damage caused by heat and to provide a new means for eliminating the skin damage caused by heat.SOLUTION: TRPV1 (transientreceptorpotentialvanilloidtype1) Agent for preventing or ameliorating skin damage caused by heat, comprising an antagonist as an active ingredient SELECTED DRAWING: None

Description

[Technical Field]

[0001] The present invention relates to an agent for preventing or improving skin damage. [Background technology]

[0002] Skin is subject to various types of damage due to exposure to sunlight and other factors. Sunlight is composed of ultraviolet rays (290-400 nm, 6.8%), visible light (400-760 nm, 38.9%), and infrared rays (760-3000 nm, 54.3%). Of these, light in the ultraviolet range in particular is known to have harmful effects on the skin, and is deeply involved in the formation of erythema and pigmentation caused by short-term exposure, as well as photoaging and carcinogenesis caused by long-term exposure. One technique for protecting the skin from UV damage is the application of sunscreen. However, applying sunscreen that only blocks UV rays may not provide sufficient sun protection, and some reports have suggested that skin damage can occur even when only UV rays are blocked (Non-Patent Document 1). On the other hand, infrared rays have also been reported to be involved in photoaging (spots and wrinkles) (Non-Patent Document 2). Furthermore, it is known that irradiating the skin with infrared rays causes a rise in temperature. Therefore, not only ultraviolet rays but also infrared rays are considered to be one of the environmental factors that significantly affect sunburn.

[0003] TRPV1 (transient receptor potential vanilloid type 1) is a member of a family of calcium ion-permeable nonselective cation channels. TRPV1 is directly activated by noxious stimuli such as ultraviolet light, heat, and capsaicin, and translocates Ca 2+ This TRPV1 activity is inhibited by specific antagonists. These conditions for TRPV1 activation imply that TRPV1 is the primary biological sensor for chemical stimuli and tissue damage. Since TRPV1 can be a target for preventing skin damage caused by exposure to sunlight, etc., a new TRPV1 activity inhibitor is needed to prevent or improve skin damage. For example, Patent Document 1 describes a composition for preventing skin aging and improving skin wrinkles, which contains a TRPV1 inhibitory peptide consisting of a specific amino acid sequence as an active ingredient. The TRPV1 inhibitory peptide described in Patent Document 1 suppresses the expression of pro-inflammatory cytokines induced by exposure to ultraviolet light, and increases intracellular Ca 2+ Since it inhibits the inflow and increase in skin thickness, it can be usefully used as an active ingredient in compositions for preventing skin aging, inhibiting the increase in skin wrinkles, increasing elasticity, whitening the skin, and alleviating inflammation, itching, or pain. [Prior art documents] [Patent documents]

[0004] [Patent Document 1] Special Publication No. 2015-515455 [Non-patent literature]

[0005] [Non-Patent Document 1] J. Dermatol. Sci. 50, 123-33 (2008) [Non-patent document 2] J. Invest. Dermatol. 128, 2491-97 (2008) Summary of the Invention [Problem to be solved by the invention]

[0006] The present invention relates to the discovery of a novel target molecule for heat-induced skin damage and to the provision of a new means for resolving heat-induced skin damage. [Means for solving the problem]

[0007] The present inventors have found that a TRPV1 antagonist that suppresses TRPV1 activity is effective in preventing or ameliorating skin damage such as sunburn caused by heat generated by exposure of the skin to infrared rays. The present invention has been completed based on these findings.

[0008] The present invention relates to an agent for preventing or improving heat-induced skin damage, which contains a TRPV1 antagonist as an active ingredient. [Effects of the Invention]

[0009] According to the present invention, a new agent is provided that can be used to prevent or improve heat-induced skin damage. DETAILED DESCRIPTION OF THE INVENTION

[0010] The active ingredient of the agent for preventing or improving heat-induced skin damage of the present invention is a TRPV1 antagonist. TRPV1 antagonists act as inactivators of TRPV1. It has been reported that suppressing TRPV1 activity can prevent photoaging caused by UV exposure (Japanese Patent Publication No. 2015-515455). However, nothing is known about their effect on heat-induced skin damage.

[0011] In this specification, "skin damage caused by heat" refers to a change (deterioration) in the normal condition of skin due to an external factor. Here, "heat" refers to the heat generated when the skin is exposed to sunlight, especially infrared rays. The amount of infrared radiation (irradiation energy) that causes skin damage cannot be determined uniquely, but is preferably 20 to 800 J / cm. 2 and more preferably 30 to 400 J / cm 2 , and more preferably 40 to 120 J / cm 2 is.

[0012] In this specification, skin damage caused by infrared rays includes, for example, an increase in the amount of melanin accumulated, deterioration of the stratum corneum state, the occurrence of inflammation, and changes in skin properties caused by these, such as a decrease in skin brightness and an increase in skin redness. Furthermore, "preventing or improving skin damage" means alleviating, suppressing, or improving such changes in skin properties.

[0013] As used herein, "skin brightness" refers to the amount of pigmentation, such as blemishes and dullness, and "skin redness" refers to the amount of discoloration due to the state of the stratum corneum or the occurrence of inflammation. Skin brightness and skin redness can be determined by measurements of brightness and hue, such as the L*a*b* color system, or by the spectral characteristics of skin. For example, skin brightness can be expressed by the L* value, and skin redness can be expressed by the a* value.

[0014] When skin is exposed to ultraviolet rays, melanin synthesis is promoted and distributed throughout the epidermis through the following process. Melanocytes receive information from other skin cells, such as keratinocytes, and synthesize melanin. Keratinocytes produce signaling substances that promote melanin synthesis in response to stimuli such as ultraviolet rays. When these substances act on melanocytes, they become activated and melanin synthesis is promoted. In melanocytes, melanin is synthesized using the amino acid tyrosine as a substrate through the action of enzymes such as tyrosinase and dopa oxidase. The melanin synthesized in melanocytes is transferred to surrounding keratinocytes through the melanocyte dendrites. When melanin transferred to keratinocytes accumulates, it can cause spots and freckles. As demonstrated in the Examples below, UV irradiation increases dopa oxidase activity, which correlates with the amount of melanin produced, but dopa oxidase activity does not increase solely due to an increase in temperature.

[0015] The present inventors therefore investigated the mechanism by which heat-induced skin damage occurs, and found that heat-induced skin damage occurs via a mechanism in which dermal fibroblasts are involved in heat-induced skin damage, and that signaling substances derived from dermal fibroblasts act on epidermal keratinocytes and melanocytes to induce inflammation or promote melanin synthesis in melanocytes. As used herein, "dermal fibroblasts" refers to cells that produce and maintain the proteins that make up the dermis (collagen, hyaluronic acid, elastin, etc.). The production of these proteins helps maintain skin firmness and moisture.

[0016] Osteopontin is a dermal fibroblast-derived factor involved in heat-induced skin damage. As used herein, "osteopontin" refers to a highly phosphorylated extracellular matrix protein. This factor acts on epidermal keratinocytes and melanocytes to induce inflammation or promote melanin synthesis in melanocytes, thereby affecting skin brightness and redness.

[0017] As used herein, "prevention" refers to preventing or delaying the onset of symptoms in an individual, or reducing the risk of an individual developing symptoms. Furthermore, "improvement" refers to improvement of a symptom or condition, prevention or delay of worsening of a symptom or condition, or reversal, prevention or delay of progression of a symptom.

[0018] As used herein, "TRPV1 antagonists" are also referred to as "TRPV1 inhibitors" and may have competitive or non-competitive antagonistic effects. TRPV1 antagonists include small molecular weight compounds, aptamers composed of oligonucleotides or peptides, and biological preparations such as neutralizing antibodies.

[0019] The TRPV1 antagonist used in the present invention is a component that inhibits TRPV1 activity, and specifically, a component that can inhibit the TRPV1 activity of 30 nM capsaicin by preferably 50% or more, more preferably 65% ​​or more, at a content of 0.001% by mass. The inhibition rate (%) of TRPV1 activity can be determined by the method described in JP 2014-111592 A.

[0020] Examples of the TRPV1 antagonist used in the present invention include trans-4-tert-butylcyclohexanol, 4-(1-methylethyl)benzenepropanal, cis-3-hexenyl anthranilate, isobutyl salicylate, N-[2-(4-chlorophenyl)ethyl]-1,3,4,5-tetrahydro-7,8-dihydroxy-2H-2-benzazepine-2-carbothioamide, N-(4-tert-butylbenzyl)-N'-[3-fluoro-4-(methylsulfonylamino)benzyl]thiourea, 4-(3 -chloro-2-pyridinyl)-N-[4-(1,1-dimethylethyl)phenyl]-1-piperazinecarboxamide, 1-isoquinolin-5-yl-3-(4-trifluoromethyl-benzyl)-urea, N-(2-bromophenyl)-N'-[2-[ethyl(3-methylphenyl)amino]ethyl]-urea, N-(3-methoxyphenyl)-4-chlorocinamide, (E)-3-(4-t-butylphenyl)-N-(2,3-dihydrobenzo[b][1,4]dioxin-6-yl)acrylamide, (E)-N-(7-hydroxy- 5,6,7,8-Tetrahydronaphthalen-1-yl)-3-(2-(piperidin-1-yl)6-(trifluoromethyl)pyridin-3-yl)acrylamide, 6-(4-fluorophenyl)-2-methyl-N-(2-methylbenzothiazol-5-yl)nicotinamide, N-(4-[6-(4-trifluoromethyl-phenyl)-pyrimidin-4-yloxy]-benzothiazol-2-yl)-acetamide, 1-[3-(trifluoromethyl)pyridin-2-yl]-N-[4-(trifluoromethylsulfonyl)phenyl ]-1,2,3,6-tetrahydropyridine-4-carboxamide, N-1H-indazol-4-yl-N'-[(1R)-5-piperidin-1-yl-2,3-dihydro-1H-inden-1-yl]urea, N-(8-tert-butyl-3,4-dihydro-2H-1-benzopyran-4-yl)-N'-(1H-indazol-4-yl)urea, N-(4-[6-(4-trifluoromethyl-phenyl)-pyrimidin-4-yloxy]-benzothiazol-2-yl)acetamide, (R)-(5-tert-butyl-2,3-dihydro-1H-inden-1-yl)-3-(1H-indazol-4-yl)-urea, N-(2-bromophenyl)-N'-[((R)-1-(5-trifluoromethyl-2-pyridyl)pyrrolidin-3-yl)]urea, (E)-N-[(R)-1-(3,5-difluoro-4-methanesulfonylamino-phenyl)-ethyl]-3-(2-propyl-6-trifluoromethyl-pyridin-3-yl)-acrylamide, etc. These may be used alone or in combination of two or more. Among them, from the viewpoints of the effect of preventing or improving heat-induced skin damage and the production costs of the agent, one or more selected from trans-4-tert-butylcyclohexanol, 4-(1-methylethyl)benzenepropanal, cis-3-hexenyl anthranilate, and isobutyl salicylate are preferred, and trans-4-tert-butylcyclohexanol is more preferred.

[0021] The TRPV1 antagonist used in the present invention may be synthesized by conventional methods or may be commercially available. Commercially available products containing trans-4-tert-butylcyclohexanol include SymSitive 1609 and SymSitive DPG (both manufactured by Symrise). A mixture of cis and trans isomers of 4-tert-butylcyclohexanol, such as PT-BUTYL CYCLOHEXANOL (manufactured by Takasago International Corporation), can also be used. Commercially available products containing 4-(1-methylethyl)benzenepropanal include CYCLEMAX (manufactured by IFF Japan Inc.), cis-3-hexenyl anthranilate include cis-3-HEXENYL ANTHRANIRATE (manufactured by Shin-Etsu Chemical Co., Ltd.), and isobutyl salicylate include iso-BUTYL SALICYLATE (manufactured by Toyotama Fragrance Co., Ltd.).

[0022] The agent for preventing or improving heat-induced skin damage of the present invention may itself be a drug, quasi-drug, or cosmetic for preventing or improving heat-induced skin damage, or may be a material or active ingredient to be incorporated into such a drug, quasi-drug, or cosmetic.

[0023] The agent for preventing or ameliorating heat-induced skin damage of the present invention may be applied directly to the skin, impregnated into a nonwoven fabric or the like and applied or stuck to the skin, or formed into a film and stuck to the skin. From the viewpoint of ease of application to the skin, the agent for preventing or ameliorating heat-induced skin damage of the present invention may be in the form of an ointment, cream, gel, lotion, foam, spray, stick (e.g., oil gel stick), oil (e.g., essential oil), sheet, etc. To produce these agents for preventing or ameliorating heat-induced skin damage, a TRPV1 antagonist may be incorporated into a base appropriate for the desired form and produced by a conventional method.

[0024] The pharmaceutical (including quasi-drugs, the same applies hereinafter) contains a TRPV1 antagonist as an active ingredient for preventing or improving heat-induced skin damage. Furthermore, the pharmaceutical may contain a pharmaceutically acceptable carrier or other active ingredient or medicinal ingredient as needed, as long as the function of the active ingredient is not impaired. The dosage form of a pharmaceutical containing a TRPV1 antagonist may be any, including oral or parenteral administration. Examples of dosage forms for oral administration include tablets, capsules, granules, powders, and syrups. Examples of dosage forms for parenteral administration include topical, transdermal, transmucosal, nasal, rectal, injection, suppository, inhalation, and patch formulations. For parenteral administration, a suitable dosage form is a topical formulation, specifically, an ointment, emulsion, cream, milky lotion, lotion, gel, aerosol, and the like.

[0025] The cosmetic contains a TRPV1 antagonist as an active ingredient for preventing or improving heat-induced skin damage. Furthermore, the cosmetic may contain a cosmetically acceptable carrier, other active ingredients, cosmetic ingredients, etc., as needed, as long as the function of the active ingredient is not impaired. Preferred examples of cosmetics containing a TRPV1 antagonist include face and body cosmetics (for example, lotions, gels, creams, packs, etc.), makeup cosmetics, and face or body cleansers.

[0026] Specific bases for the agent for preventing or improving heat-induced skin damage of the present invention include lower alcohols such as ethanol; higher alcohols such as myristyl alcohol, cetanol, stearyl alcohol, isostearyl alcohol, and behenyl alcohol; fatty acid esters such as isopropyl myristate, diisopropyl adipate, and neopentyl glycol dicaprate; alkyl glyceryl ethers, polyethylene glycol fatty acid esters, polyoxyethylene alkyl ethers, glycerin fatty acid esters, polyoxyethylene glycerin fatty acid esters, sorbitan fatty acid esters, and polyoxyethylene sorbitan fatty acid esters. surfactants such as esters, polyoxyethylene sorbitol fatty acid esters, polyglycerin fatty acid esters, polyoxyethylene hydrogenated castor oil, fatty acid salts, and N-acylamino acid salts; polyhydric alcohols such as glycerin, propylene glycol, 1,3-butylene glycol, pentylene glycol, dipropylene glycol, polyethylene glycol, erythritol, sorbitol, xylitol, and polyglycerin; neutralizing agents such as sodium hydroxide, potassium hydroxide, and triethanolamine, pH adjusters such as sodium citrate and sodium lactate, and preservatives such as methylparaben and propylparaben; and water.

[0027] The content of the TRPV1 antagonist in the above-mentioned pharmaceutical or cosmetic formulations cannot be generalized because it varies depending on the type of TRPV1 antagonist and the form of the formulation, but from the viewpoint of the preventive or ameliorative effect on heat-induced skin damage and production costs, for example, based on the total amount of the formulation, it is preferably 0.001% by mass or more, more preferably 0.01% by mass or more, even more preferably 0.05% by mass or more, and preferably 1% by mass or less, more preferably 0.5% by mass or less, and even more preferably 0.3% by mass or less. Also, 0.001 to 1% by mass is preferred, 0.01 to 0.5% by mass is more preferred, and 0.05 to 0.3% by mass is even more preferred.

[0028] The dosage or use amount of the TRPV1 antagonist can be an amount that can achieve the effects of the present invention. The dosage or use amount can vary depending on the type of TRPV1 antagonist, the subject's species, body weight, sex, age, condition, or other factors. In the case of parenteral administration such as topical skin preparations, the dosage or use amount is preferably 0.00002 mg / cm per adult (60 kg) once. 2 Above the skin surface, preferably 0.0002 mg / cm 2 above the skin surface, even more preferably 0.002 mg / cm 2 above the skin surface, preferably 0.02 mg / cm 2 Below the skin surface, preferably 0.01 mg / cm 2 below the skin surface, even more preferably 0.004 mg / cm 2 Below the skin surface. 2 Skin surface ~0.02mg / cm 2 The skin surface is preferred, 0.0002 mg / cm 2 Skin surface ~0.01mg / cm 2 The skin surface is more preferably 0.002 mg / cm 2 Skin surface ~0.004mg / cm 2 In the present invention, such an amount can be administered or used repeatedly or continuously, once or in divided doses per day, for one day or more, preferably for 14 days or more, more preferably for 28 days or more, and even more preferably for 49 days or more.

[0029] As shown in the Examples below, TRPV1 is an important skin color control factor involved in heat-induced pigmentation enhancement and inflammation induction, and has been shown to be a target for suppressing heat-induced skin damage. Therefore, TRPV1 antagonists have the effect of suppressing heat-induced skin damage, and can be used as or to produce agents for preventing or improving heat-induced skin damage. Furthermore, TRPV1 antagonists can be applied to animals, including humans, to prevent or improve heat-induced skin damage.

[0030] As used herein, "use" may refer to use in humans or non-human animals, and may be therapeutic or non-therapeutic use. "Non-therapeutic" is a concept that does not include medical procedures, i.e., methods of surgery, therapy, or diagnosis on humans, and more specifically, methods of surgery, therapy, or diagnosis on humans by physicians or those under the direction of physicians.

[0031] The agent for preventing or improving heat-induced skin damage of the present invention can be administered to or used by humans or non-human animals who need or desire to prevent or improve heat-induced skin damage. Specific examples include humans and non-human animals that have or are suffering from heat-induced skin damage. Non-human animals include non-human mammals such as apes, other primates, and felines.

[0032] The site to which the agent for preventing or improving heat-induced skin damage of the present invention is administered or used is not particularly limited as long as it is a site that has been or may be damaged by heat, and specific examples include the skin of the arms, backs of the hands, fingers, legs, neck, back, face, ears, scalp, etc.

[0033] In relation to the above-mentioned embodiments, the present invention further discloses the following agent for preventing or improving skin damage or use thereof, TRPV1 antagonist, use, and method.

[0034] <1> An agent for preventing or improving heat-induced skin damage, containing a TRPV1 antagonist as an active ingredient.

[0035] <2> The heat-induced skin damage is skin damage caused by heat generated when exposed to sunlight, particularly infrared rays. <1> Item 1. A preventive or ameliorating agent for heat-induced skin damage. <3> The heat-induced skin damage is an increase in the amount of melanin accumulated, a deterioration in the condition of the stratum corneum, or the occurrence of inflammation. <1> or <2> Item 1. A preventive or ameliorating agent for heat-induced skin damage. <4> The heat-induced skin damage is a change in skin properties selected from a decrease in skin brightness and an increase in skin redness. <1> ~ <3> 10. The agent for preventing or improving heat-induced skin damage according to any one of claims 1 to 9. <5> The above-mentioned heat-induced skin damage involves dermal fibroblasts, <1> ~ <4> 10. The agent for preventing or improving heat-induced skin damage according to any one of claims 1 to 9. <6> The signaling substance derived from the dermal fibroblasts acts on the keratinocytes and melanocytes of the epidermis, causing skin damage due to heat. <5> Item 1. A preventive or ameliorating agent for heat-induced skin damage. <7> The dermal fibroblast-derived factor acting on keratinocytes and melanocytes is osteopontin. <6> Item 1. A preventive or ameliorating agent for heat-induced skin damage. <8> The TRPV1 antagonist can inhibit the TRPV1 activity of 30 nM capsaicin by 50% or more, preferably 65% ​​or more, at a content of 0.001% by mass. <1> ~ <7> 10. The agent for preventing or improving heat-induced skin damage according to any one of claims 1 to 9. <9> The TRPV1 antagonist may be selected from the group consisting of trans-4-tert-butylcyclohexanol, 4-(1-methylethyl)benzenepropanal, cis-3-hexenyl anthranilate, isobutyl salicylate, N-[2-(4-chlorophenyl)ethyl]-1,3,4,5-tetrahydro-7,8-dihydroxy-2H-2-benzazepine-2-carbothioamide, N-(4-tert-butylbenzyl)-N'-[3-fluoro-4-(methylsulfonylamino)benzyl]thiourea, 4-(3-chloro-2-pyridinyl )-N-[4-(1,1-dimethylethyl)phenyl]-1-piperazinecarboxamide, 1-isoquinolin-5-yl-3-(4-trifluoromethyl-benzyl)-urea, N-(2-bromophenyl)-N'-[2-[ethyl(3-methylphenyl)amino]ethyl]-urea, N-(3-methoxyphenyl)-4-chlorocinamide, (E)-3-(4-t-butylphenyl)-N-(2,3-dihydrobenzo[b][1,4]dioxin-6-yl)acrylamide, (E)-N-(7-hydroxy-5,6,7,8- Tetrahydronaphthalen-1-yl)-3-(2-(piperidin-1-yl)6-(trifluoromethyl)pyridin-3-yl)acrylamide, 6-(4-fluorophenyl)-2-methyl-N-(2-methylbenzothiazol-5-yl)nicotinamide, N-(4-[6-(4-trifluoromethyl-phenyl)-pyrimidin-4-yloxy]-benzothiazol-2-yl)-acetamide, 1-[3-(trifluoromethyl)pyridin-2-yl]-N-[4-(trifluoromethylsulfonyl)phenyl]-1, 2,3,6-Tetrahydropyridine-4-carboxamide, N-1H-indazol-4-yl-N'-[(1R)-5-piperidin-1-yl-2,3-dihydro-1H-inden-1-yl]urea, N-(8-tert-butyl-3,4-dihydro-2H-1-benzopyran-4-yl)-N'-(1H-indazol-4-yl)urea, N-(4-[6-(4-trifluoromethyl-phenyl)-pyrimidin-4-yloxy]-benzothiazol-2-yl)acetamide, (R)-(5-tert-butyl-2,the said amine is one or more selected from the group consisting of 3-dihydro-1H-inden-1-yl)-3-(1H-indazol-4-yl)-urea, N-(2-bromophenyl)-N'-[((R)-1-(5-trifluoromethyl-2-pyridyl)pyrrolidin-3-yl)]urea, and (E)-N-[(R)-1-(3,5-difluoro-4-methanesulfonylamino-phenyl)-ethyl]-3-(2-propyl-6-trifluoromethyl-pyridin-3-yl)-acrylamide, preferably one or more selected from the group consisting of trans-4-tert-butylcyclohexanol, 4-(1-methylethyl)benzenepropanal, cis-3-hexenyl anthranilate, and isobutyl salicylate; <1> ~ <8> The agent for preventing or improving heat-induced skin damage according to any one of the above. <10> The content of the TRPV1 antagonist in the formulation is preferably 0.001% by mass or more, more preferably 0.01% by mass or more, even more preferably 0.05% by mass or more, based on the total amount of the formulation, and is preferably 1% by mass or less, more preferably 0.5% by mass or less, even more preferably 0.3% by mass or less, and is preferably 0.001 to 1% by mass, more preferably 0.01 to 0.5% by mass, even more preferably 0.05 to 0.3% by mass. <1> ~ <9> 10. The agent for preventing or improving heat-induced skin damage according to any one of the preceding items. <11> The dose or use amount of the TRPV1 antagonist, when administered parenterally, is preferably 0.00002 mg / cm per adult (60 kg). 2 Above the skin surface, preferably 0.0002 mg / cm 2 above the skin surface, even more preferably 0.002 mg / cm 2 above the skin surface, and preferably 0.02 mg / cm 2 Below the skin surface, preferably 0.01 mg / cm 2 below the skin surface, even more preferably 0.004 mg / cm 2 below the skin surface, and preferably 0.00002 mg / cm 2 Skin surface ~0.02mg / cm 2On the skin surface, more preferably 0.0002 mg / cm 2 Skin surface ~0.01mg / cm 2 skin surface, even more preferably 0.002 mg / cm 2 Skin surface ~0.004mg / cm 2 The skin surface <1> ~ <10> 10. The agent for preventing or improving heat-induced skin damage according to any one of claims 1 to 9. <12> The dosage form is a topical agent for skin. <1> ~ <11> 10. The agent for preventing or improving heat-induced skin damage according to any one of claims 1 to 9.

[0036] <13> Use of a TRPV1 antagonist as an agent for preventing or improving heat-induced skin damage. <14> Use of a TRPV1 antagonist for manufacturing an agent for preventing or improving heat-induced skin damage. <15> A method for using a TRPV1 antagonist as an agent for preventing or improving heat-induced skin damage. <16> A TRPV1 antagonist for use in a method for preventing or improving heat-induced skin damage. <17> Use of a TRPV1 antagonist for manufacturing a drug for preventing or improving heat-induced skin damage. <18> Non-therapeutic use of a TRPV1 antagonist to prevent or ameliorate heat-induced skin damage. <19> A method for preventing or improving heat-induced skin damage by administering or applying a TRPV1 antagonist to a subject. <20> The TRPV1 antagonist is administered or applied to the arm, the back of the hand, the finger, the leg, the neck, the back, the face, the ear, or the scalp. <18> or <19> Item 1. A method for preventing or improving skin damage caused by heat. <21> The dose or use amount of the TRPV1 antagonist, when administered parenterally, is preferably 0.00002 mg / cm per adult (60 kg). 2 Above the skin surface, preferably 0.0002 mg / cm 2 above the skin surface, even more preferably 0.002 mg / cm 2 above the skin surface, and preferably 0.02 mg / cm 2Below the skin surface, preferably 0.01 mg / cm 2 below the skin surface, even more preferably 0.004 mg / cm 2 below the skin surface, and preferably 0.00002 mg / cm 2 Skin surface ~0.02mg / cm 2 On the skin surface, more preferably 0.0002 mg / cm 2 Skin surface ~0.01mg / cm 2 skin surface, even more preferably 0.002 mg / cm 2 Skin surface ~0.004mg / cm 2 The skin surface <18> ~ <20> 10. A method for preventing or improving heat-induced skin damage according to any one of claims 1 to 9. <22> The TRPV1 antagonist is administered or used repeatedly or continuously once or multiple times a day for at least 1 day, preferably at least 14 days, more preferably at least 28 days, and even more preferably at least 49 days. <18> ~ <21> 10. A method for preventing or improving heat-induced skin damage according to any one of claims 1 to 9.

[0037] <23> The heat-induced skin damage is skin damage caused by heat generated when exposed to sunlight, particularly infrared rays. <13> ~ <22> The TRPV1 antagonist or its use, or method described in any one of the above. <24> The heat-induced skin damage is an increase in the amount of melanin accumulated, a deterioration in the condition of the stratum corneum, or the occurrence of inflammation. <13> ~ <23> The TRPV1 antagonist or its use, or method described in any one of the above. <25> The heat-induced skin damage is a change in skin properties selected from a decrease in skin brightness and an increase in skin redness. <13> ~ <24> The TRPV1 antagonist or its use, or method described in any one of the above. <26> The above-mentioned heat-induced skin damage involves dermal fibroblasts, <13> ~ <25> The TRPV1 antagonist or its use, or method described in any one of the above. <27> The signaling substance derived from the dermal fibroblasts acts on the keratinocytes and melanocytes of the epidermis, causing skin damage due to heat. <26> Item 1. A preventive or ameliorating agent for heat-induced skin damage. <28> The dermal fibroblast-derived factor acting on keratinocytes and melanocytes is osteopontin. <27> Item 1. A preventive or ameliorating agent for heat-induced skin damage. <29> The TRPV1 antagonist can inhibit the TRPV1 activity of 30 nM capsaicin by 50% or more, preferably 65% ​​or more, at a content of 0.001% by mass. <13> ~ <28> The TRPV1 antagonist or its use, or method described in any one of the above. <30> The TRPV1 antagonist may be selected from the group consisting of trans-4-tert-butylcyclohexanol, 4-(1-methylethyl)benzenepropanal, cis-3-hexenyl anthranilate, isobutyl salicylate, N-[2-(4-chlorophenyl)ethyl]-1,3,4,5-tetrahydro-7,8-dihydroxy-2H-2-benzazepine-2-carbothioamide, N-(4-tert-butylbenzyl)-N'-[3-fluoro-4-(methylsulfonylamino)benzyl]thiourea, 4-(3-chloro-2-pyridinyl )-N-[4-(1,1-dimethylethyl)phenyl]-1-piperazinecarboxamide, 1-isoquinolin-5-yl-3-(4-trifluoromethyl-benzyl)-urea, N-(2-bromophenyl)-N'-[2-[ethyl(3-methylphenyl)amino]ethyl]-urea, N-(3-methoxyphenyl)-4-chlorocinamide, (E)-3-(4-t-butylphenyl)-N-(2,3-dihydrobenzo[b][1,4]dioxin-6-yl)acrylamide, (E)-N-(7-hydroxy-5,6,7,8- Tetrahydronaphthalen-1-yl)-3-(2-(piperidin-1-yl)6-(trifluoromethyl)pyridin-3-yl)acrylamide, 6-(4-fluorophenyl)-2-methyl-N-(2-methylbenzothiazol-5-yl)nicotinamide, N-(4-[6-(4-trifluoromethyl-phenyl)-pyrimidin-4-yloxy]-benzothiazol-2-yl)-acetamide, 1-[3-(trifluoromethyl)pyridin-2-yl]-N-[4-(trifluoromethylsulfonyl)phenyl]-1, 2,3,6-Tetrahydropyridine-4-carboxamide, N-1H-indazol-4-yl-N'-[(1R)-5-piperidin-1-yl-2,3-dihydro-1H-inden-1-yl]urea, N-(8-tert-butyl-3,4-dihydro-2H-1-benzopyran-4-yl)-N'-(1H-indazol-4-yl)urea, N-(4-[6-(4-trifluoromethyl-phenyl)-pyrimidin-4-yloxy]-benzothiazol-2-yl)acetamide, (R)-(5-tert-butyl-2,the compound is one or more selected from the group consisting of 3-dihydro-1H-inden-1-yl)-3-(1H-indazol-4-yl)-urea, N-(2-bromophenyl)-N'-[((R)-1-(5-trifluoromethyl-2-pyridyl)pyrrolidin-3-yl)]urea, and (E)-N-[(R)-1-(3,5-difluoro-4-methanesulfonylamino-phenyl)-ethyl]-3-(2-propyl-6-trifluoromethyl-pyridin-3-yl)-acrylamide, preferably one or more selected from trans-4-tert-butylcyclohexanol, 4-(1-methylethyl)benzenepropanal, cis-3-hexenyl anthranilate, and isobutyl salicylate; <13> ~ <29> The TRPV1 antagonist or its use, or method according to any one of the preceding claims. <31> The content of the TRPV1 antagonist in the formulation is preferably 0.001% by mass or more, more preferably 0.01% by mass or more, even more preferably 0.05% by mass or more, based on the total amount of the formulation, and is preferably 1% by mass or less, more preferably 0.5% by mass or less, even more preferably 0.3% by mass or less, and is preferably 0.001 to 1% by mass, more preferably 0.01 to 0.5% by mass, even more preferably 0.05 to 0.3% by mass. <13> ~ <30> The TRPV1 antagonist or its use, or method described in any one of the above. [Example]

[0038] The present invention will be described in more detail below based on examples, but the present invention is not limited to these examples.

[0039] Test Example 1 Improves skin tone Sunscreens (SPF50+, PA++++) having the compositions and blending amounts (active ingredient, mass %) shown in Table 1 were produced by the following method, and their effects were evaluated.

[0040] (Manufacturing method) All ingredients shown in Table 1 except water were blended and mixed uniformly using a disper. Purified water was then added to the resulting mixture, and the mixture was mixed uniformly using a homogenizer to produce topical skin preparations (placebo sunscreen, TRPV1 antagonist-containing sunscreen).

[0041] [Table 1]

[0042] (Evaluation method) Thirty healthy women were used as subjects to evaluate the effect of continuous application for eight weeks on improving skin color. The subjects were divided into two groups of 15 each. One group applied a placebo sunscreen to the entire face, while the other group applied a TRPV1 antagonist-containing sunscreen to the entire face, approximately 0.68g per application, once daily. Skin color was measured before and after 8 weeks of application, and changes in skin color were evaluated. Changes in skin color were evaluated based on changes in lightness (L* value) and redness (a* value). Lightness (L* value) and redness (a* value) were measured using a spectrophotometer CM-2600d (Konica Minolta). Measurements were taken three times on the cheek, and the average value was calculated. The results are shown in Tables 2 and 3.

[0043] [Table 2]

[0044] [Table 3]

[0045] The results in Table 2 show that the TRPV1 antagonist-containing sunscreen significantly increased lightness (L* value) and significantly reduced redness (a* value) at week 8 compared to week 0. On the other hand, the placebo sunscreen, which does not contain a TRPV1 antagonist, did not show any significant changes in lightness or redness at week 8.

[0046] The results in Table 3 show that the change in lightness (ΔL*) from week 0 to week 8 was greater with the TRPV1 antagonist-containing sunscreen than with the placebo sunscreen. The change in redness (Δa*) from week 0 to week 8 showed a tendency for the TRPV1 antagonist-containing sunscreen to suppress an increase in skin redness compared to the placebo sunscreen.

[0047] Test Example 2 Normal human epidermal melanocytes (melanocytes; manufactured by Kurabo) were cultured in Medium 254 (Thermo Fisher Scientific) containing a growth additive (HMGS; manufactured by Kurabo) at 37°C in a 5% CO2 atmosphere according to standard methods. Normal human neonatal foreskin-derived epidermal keratinocytes (Keratinocytes; Kurabo Industries, Ltd.) were cultured in a standard epidermal keratinocyte growth medium (EpiLife Medium, with 60 μM calcium; Thermo Fisher Scientific) containing a growth additive (HKGS; Thermo Fisher Scientific) at 37°C and 5% CO2. For various evaluations, EpiLife Medium with 60 μM calcium supplemented with the HKGS Kit (BPE- and EGF-free; Thermo Fisher Scientific; hereafter referred to as "EpiLife Evaluation Medium (BPE- and EGF-free)") was used. Foreskin-derived normal human dermal fibroblasts (Kurabo) were cultured in Dulbecco's modified Eagle's medium (DMEM; Sigma-Aldrich) containing 10% (v / v) fetal bovine serum (FBS; Sigma-Aldrich) and 1% (v / v) penicillin-streptomycin (Thermo Fisher Scientific) at 37°C in a 5% CO environment according to standard methods.

[0048] A 300W xenon light source (MAX-301, manufactured by Asahi Spectroscopy) equipped with a xenon lamp was used as the light source, and a WG-320 filter (1 mm thick, manufactured by Shibuya Optical Co., Ltd.) was attached to it to create an ultraviolet irradiation device (ultraviolet A rays (UVA):ultraviolet B rays (UVB) = 10.8:1 (irradiation energy ratio)).

[0049] (1) UV irradiation and dopa oxidase activity measurement in a co-culture system of melanocytes and keratinocytes Keratinocytes were cultured at 1.0 × 10 cells per well in a 96-well plate (collagen-coated) using EpiLife evaluation medium (containing no BPE or EGF). 4 cells / well, melanocytes 2.0 × 10 4 The next day, the medium was replaced with HBSS (Thermo Fisher Scientific), and the cells were exposed to UV light (2.5 J / cm) for 15 minutes on a heat block (set to the same temperature as the incubator) in an incubator set to 37°C or 43°C. 2 ) was performed (UV irradiation group). The control group was not UV-irradiated and was kept on a heat block (set to the same temperature as the incubator) for 15 minutes. After irradiation, the medium was replaced with EpiLife evaluation medium and cultured at 37°C and 5% CO2. Dopa oxidase activity was measured after 3 days. Specifically, the culture plate was washed with PBS, and 20 μL / well of extraction buffer (0.1 M Tris-HCl (pH 7.2), 1% NP-40, 0.01% SDS) and 20 μL / well of assay buffer (4% dimethylformamide, 100 mM sodium phosphate buffer (pH 7.1)) were added. Cells were solubilized at 4°C for 2 hours. To each well of the solubilized cell solution, 80 μL of the above-mentioned Assay Buffer, 60 μL of 20.7 mM MBTH (3-methyl-2-benzothiazolinone hydrazone) solution, and 40 μL of 5 mM L-DOPA (L-dihydroxyphenylalanine) solution were added and incubated for 30 minutes at 37°C. After incubation, the absorbance at 505 nm was measured and used as an index of dopa oxidase activity. The control group, maintained on a heat block at 37°C, was used as the control, and the absorbance was set at 100 to evaluate dopa oxidase activity relative to the control. The results are shown in Table 4.

[0050] [Table 4]

[0051] As shown in Table 4, when UV irradiation was performed, dopa oxidase activity significantly increased at both 37°C and 43°C. However, there was no significant difference between 37°C and 43°C, and no increase in dopa oxidase activity due to temperature increase was observed. Similarly, no significant difference was observed between 37°C and 43°C in the control group.

[0052] (2) Evaluation using three types of co-culture systems Keratinocytes were cultured at a density of 5.0 × 10 cells in a multi-cell culture dish (VWR, 60 mm diameter dish with 18 8 mm diameter cells placed) using EpiLife evaluation medium (containing no BPE or EGF). 5 cells / dish, melanocytes 5.0 × 10 5 Dermal fibroblasts were seeded at a density of 5.0 × 10 cells / dish in a 24-well plate. 4 The cells were seeded at a density of 100 cells / well in Dulbecco's modified Eagle's medium (containing FBS and penicillin-streptomycin), and the next day, they were replaced with EpiLife evaluation medium (containing no BPE or EGF) and cultured for another day. The medium was replaced with HBSS (Thermo Fisher Scientific), and the cells were exposed to UV light (2.5 J / cm) for 15 minutes on a heat block (set to the same temperature as the incubator) in an incubator set at 37°C or 43°C. 2) were performed separately. The control group was not exposed to UV light and was maintained on a heat block (37°C or 43°C) for 15 minutes. After irradiation, one cell from the multi-cell culture dish was transferred to a 24-well plate, and three types of co-cultures were initiated in EpiLife evaluation medium (without BPE or EGF). Dopa oxidase activity was measured six days later. Specifically, one cell from the multi-cell culture dish was transferred to another 24-well plate, and 100 μL / well of extraction buffer (0.1 M Tris-HCl (pH 7.2), 1% NP-40, 0.01% SDS) and 100 μL / well of assay buffer (4% dimethylformamide, 100 mM sodium phosphate buffer (pH 7.1)) were added. Cells were solubilized at 4°C for 18 hours, and 40 μL of the solubilized cell solution was added to a 96-well plate. This was then incubated at 37°C for 30 minutes, after which the absorbance at 505 nm was measured and used as an index of dopa oxidase activity. The control group, which was maintained on a heat block at 37°C, was used as the control, and the absorbance was set at 100 to evaluate dopa oxidase activity. The results are shown in Table 5.

[0053] [Table 5]

[0054] As shown in Table 5, in the triple co-culture system, it was confirmed that dopa oxidase activity was significantly elevated and melanin synthesis ability was increased by applying heat stimulation in addition to UV irradiation. These results suggest that dermal fibroblasts are involved in the heat-induced enhancement of pigmentation.

[0055] (3) Effect of TRPV1 antagonist on heat-induced increase in dopa oxidase activity tBCH (trans-4-tert-Butylcyclohexanol; Tokyo Chemical Industry Co., Ltd.) was used as a TRPV1 antagonist. The effects of TRPV1 antagonists were evaluated using three types of co-culture systems under the same culture conditions, UV irradiation conditions, and temperature conditions as in (2). tBCH was added to HBSS (Thermo Fisher Scientific), the same solution used for UV irradiation of dermal fibroblasts, to a final concentration of 30 μM. As in (2), the three types of co-culture were initiated after UV irradiation, and dopa oxidase activity was measured six days later. The dopa oxidase activity was evaluated relative to the value of the control group, which was not UV-irradiated and kept on a heat block at 37°C without tBCH. The results are shown in Table 6.

[0056] [Table 6]

[0057] As shown in Table 6, in the triple co-culture system, the increase in dopa oxidase activity that occurred when UV irradiation and heat stimulation were combined was significantly suppressed by adding the TRPV1 antagonist tBCH to dermal fibroblasts. These results indicate that the heat-induced enhancement of pigmentation (increased dopa oxidase activity) is related to the activation of TRPV1 in dermal fibroblasts, and suggest that inhibiting TRPV1 activity in dermal fibroblasts is important for preventing or improving skin damage.

[0058] Test Example 3 (1) Evaluation of proteins secreted by dermal fibroblasts 5.0 × 10 dermal fibroblasts were placed in a 24-well plate. 4The cells were seeded at a density of 1000 cells / well in Dulbecco's modified Eagle's medium (containing FBS and penicillin-streptomycin), and the next day, they were replaced with EpiLife evaluation medium (containing no BPE or EGF) and cultured for another day. The next day, the medium was replaced with HBSS (Thermo Fisher Scientific), and the cells were exposed to ultraviolet (UV) radiation (2.5 J / cm) for 15 minutes on a heat block (37°C or 43°C) in an incubator set at 37°C or 43°C. 2 ) was performed. The same equipment as in Test Example 2 was used for UV irradiation. As a control, UV irradiation was not performed and the cells were kept on a heat block (37°C or 43°C) for 15 minutes. After UV irradiation, the medium was replaced with EpiLife evaluation medium and culture was continued. After 6 days, the culture supernatant was collected and the proteins secreted by the dermal fibroblasts were quantified using a Proteome Profiler Human XL Cytokine Array Kit (R&D Systems). The measurement data was evaluated relative to the protein amount, with the value obtained at 37°C without UV irradiation being set to 1. Table 7 shows the results for proteins whose secretion amount increased by two-fold or more after UV irradiation at 43°C.

[0059] [Table 7]

[0060] As shown in Table 7, it was confirmed that UV irradiation at 43°C increased the expression levels of many factors derived from dermal fibroblasts.

[0061] (2) Measurement of dopa oxidase activity and melanin quantification in a co-culture system of melanocytes and keratinocytes supplemented with secretory proteins from dermal fibroblasts 2.0 × 10 keratinocytes were cultured in a 6-well plate (collagen-coated) using EpiLife evaluation medium (containing no BPE or EGF). 5 cells / well, melanocytes 2.0 × 10 5 The next day, the medium was replaced with HBSS (Thermo Fisher Scientific), and the cells were exposed to UV light (2.5 J / cm) for 15 minutes on a heat block in an incubator set at 37°C.2 After irradiation, the medium was replaced with EpiLife evaluation medium supplemented with various secreted proteins, and culturing continued. A control group was cultured under the same conditions without the addition of secreted proteins. After 6 days, the dopa oxidase culture plates were washed with PBS, and the cells were harvested using a cell detachment enzyme (Accumax; Nacalai Tesque). The number of viable cells was counted, followed by measurement of dopa oxidase activity and melanin quantification. Dopa oxidase activity was measured by adding 50 μL / well of extraction buffer (0.1 M Tris-HCl (pH 7.2), 1% NP-40, 0.01% SDS) and 50 μL / well of assay buffer (4% dimethylformamide, 100 mM sodium phosphate buffer (pH 7.1)) to the cells and solubilizing them at 4°C for 2 hours. Then, 40 μL of the solubilized cell solution, 80 μL of the assay buffer, 60 μL of 20.7 mM MBTH (3-methyl-2-benzothiazolinone hydrazone) solution, and 40 μL of 5 mM L-DOPA (L-dihydroxyphenylalanine) solution as a substrate were added to a 96-well plate and incubated at 37°C for 30 minutes. The absorbance at 505 nm was measured, and the absorbance per viable cell count was used as an index of dopa oxidase activity. To quantify melanin, cells were dissolved in 2N aqueous NaOH, and absorbance (405 nm) was measured using a plate reader. The amount of melanin per viable cell was calculated from a calibration curve prepared using synthetic melanin (SIGMA). Measurement and quantitative data were evaluated relative to the value of 100 for the control group exposed to UV light at 37°C. The secretory proteins added and their final concentrations at the time of addition are as follows. SDF-1α (10 nM; Fujifilm Wako Pure Chemical Industries, Ltd.) Osteopontin (10 nM; Fujifilm Wako Pure Chemical Industries, Ltd.) MCP-1 (100 ng / mL; Fujifilm Wako Pure Chemical Industries, Ltd.) VEGF (1 ng / mL; Fujifilm Wako Pure Chemical Industries, Ltd.) The results are shown in Table 8.

[0062] [Table 8]

[0063] As shown in Table 8, when osteopontin was added, UV irradiation tended to increase dopa oxidase activity, and the amount of melanin produced also increased statistically significantly. These results suggest that osteopontin has the effect of promoting UV-induced melanogenesis.

[0064] As shown above, it has been demonstrated that TRPV1 antagonists that effectively suppress TRPV1 activity are effective in preventing or improving skin damage caused by heat, such as sunburn.

Claims

1. A preventative or ameliorative agent for heat-induced skin damage, containing a TRPV1 (transient receptor potential vanilloid type 1) antagonist as its active ingredient.

2. 2. The agent for preventing or improving heat-induced skin damage according to claim 1, wherein the heat-induced skin damage is skin damage caused by heat generated when irradiated with infrared rays.

3. 3. The agent for preventing or improving heat-induced skin damage according to claim 1 or 2, wherein the heat-induced skin damage is an increase in the amount of melanin accumulated, a deterioration in the condition of the stratum corneum, or the occurrence of inflammation.

4. 3. The agent for preventing or improving heat-induced skin damage according to claim 1, wherein the heat-induced skin damage is a change in skin properties selected from a decrease in skin brightness and an increase in skin redness.

5. The agent for preventing or improving heat-induced skin damage according to claim 1 or 2, wherein the heat-induced skin damage involves dermal fibroblasts.

6. The agent for preventing or improving heat-induced skin damage according to claim 5, wherein the dermal fibroblast-derived signaling substance acts on epidermal keratinocytes and melanocytes, thereby causing heat-induced skin damage.

7. The agent for preventing or improving heat-induced skin damage according to claim 6, wherein the factor derived from dermal fibroblasts that acts on keratinocytes and melanocytes is osteopontin.

8. 3. The agent for preventing or improving heat-induced skin damage according to claim 1 or 2, wherein the TRPV1 antagonist is one or more selected from the group consisting of trans-4-tert-butylcyclohexanol, 4-(1-methylethyl)benzenepropanal, cis-3-hexenyl anthranilate, and isobutyl salicylate.

9. The agent for preventing or improving heat-induced skin damage according to claim 1 or 2, which is in the form of an external preparation for skin.

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