JP6698318B2 - Ceramide synthesis accelerator - Google Patents

Description

  The present invention relates to a ceramide synthesis promoter capable of promoting the expression of serine palmitoyltransferase and promoting ceramide synthesis.

  The skin covers the outermost layer of the living body, forms a boundary with the outside world, and has a barrier function essential for life support such as prevention of invasion of external substances and loss of internal components including water. Ceramide-based lipids form a lamellar structure between keratinocytes and play an important role in exerting a barrier function.

  When ceramide decreases due to internal or external factors such as aging and UV exposure, the barrier function decreases, and various skin disorders such as atopic dermatitis, ichthyosis, acne, skin immunity abnormality, and skin infections. Is known to induce. Conventionally, in order to suppress the skin disorder induced by the decrease of ceramide, a method of externally supplying the decreased ceramide has been attempted. However, the method of supplementing the reduced ceramide from the outside is not sufficient in sustainability of the effect, so that the effect is not satisfactory.

  On the other hand, in the process of keratinization of epidermal cells, ceramide is known to be synthesized through several reactions, starting from the condensation reaction of serine and palmitoyl-CoA catalyzed by serine palmitoyltransferase. It is also known that in ceramide biosynthesis, the condensation reaction by serine palmitoyltransferase is rate-limiting.

  Therefore, in recent years, as a method of suppressing the skin damage induced by the decrease of ceramide, by promoting the expression of serine palmitoyltransferase, a method of promoting the synthesis of ceramide by the cells themselves, has been studied, serine palmitoyl transferase Ingredients that promote expression have also been found. For example, in Patent Document 1, the action of promoting the expression of serine palmitoyl transferase in the extract of ashitaba, the extract of nettle, the extract of Indian mackerel, the extract of Kumazasa, the extract of Gentian, and the extract of tonin is disclosed. It has been reported that there is. Further, Patent Document 2 reports that an extract of Japanese maple tree has an action of promoting the expression of serine palmitoyltransferase. However, in order to meet demands such as diversification of pharmaceutical formulations and further improvement of ceramide synthesis promoting action, development of a new component that promotes expression of serine palmitoyltransferase is desired.

JP, 2009-107965, A JP, 2011-68594, A

  An object of the present invention is to provide a ceramide synthesis promoter capable of promoting the expression of serine palmitoyltransferase and promoting ceramide synthesis.

  The present inventors have conducted extensive studies to solve the above problems, and found that sulfated glucosaminoglycans have an action of promoting the expression of serine palmitoyltransferase, and can promote ceramide synthesis by epidermal cells. .. The present invention has been completed by further studies based on such findings.

That is, the present invention provides the inventions of the following modes.
Item 1. A ceramide synthesis accelerator comprising a sulfated glycosaminoglycan.
Item 2. Item 2. The ceramide synthesis accelerator according to Item 1, wherein the sulfated glycosaminoglycan is a heparin-like substance.
Item 3. Item 3. The ceramide synthesis accelerator according to Item 1 or 2, which is a skin external preparation.
Item 4. Item 4. The ceramide synthesis promoter according to any one of Items 1 to 3, which is used for the prevention or treatment of atopic dermatitis, ichthyosis, acne, skin immune abnormality, or skin infection.
Item 5. A serine palmitoyltransferase expression-enhancing agent comprising a sulfated glycosaminoglycan.

  According to the ceramide synthesis promoter of the present invention, by promoting the expression of serine palmitoyltransferase in epidermal cells, it is possible to promote the synthesis of ceramide, and therefore it is effective for the prevention or treatment of skin disorders induced by the decrease of ceramide. ..

FIG. 3 is a diagram showing the results of culturing normal epidermal model cells in the presence of a heparin-like substance and measuring the expression level of serine palmitoyltransferase mRNA. It is a figure which shows the result of having culture|cultivated the epidermal upper layer model cell in presence of a heparin analog, and measuring the expression level of serine palmitoyl transferase mRNA.

1. Ceramide Synthesis Promoter The ceramide synthesis promoter of the present invention is characterized by containing a sulfated glycosaminoglycan. Hereinafter, the ceramide synthesis accelerator of the present invention will be described in detail.

Sulfated Glucosaminoglycan Sulfated glucosaminoglycan is a mucopolysaccharide containing an amino sugar as one of the constituent units and having a sulfate group.

  The type of sulfated glycosaminoglycan used in the present invention is not particularly limited as long as it is pharmaceutically acceptable, for example, heparin analog, heparan sulfate, chondroitin sulfate, keratan sulfate, heparin, etc. Is mentioned. The sulfated glycosaminoglycan may be in the form of a metal salt such as sodium, potassium, calcium, magnesium, iron and manganese; a salt such as ammonium salt.

  These sulfated glycosaminoglycans may be used alone or in combination of two or more. Among these, heparin-like substances are preferable from the viewpoint of promoting ceramide synthesis more effectively.

  The origin of the sulfated glycosaminoglycan is not particularly limited, for example, those obtained by polysulfating mucopolysaccharides, edible animal tissues (for example, in the case of heparin-like substances , Lungs including tracheal cartilage of cows, pigs, etc.) and the like.

  In addition, for example, when a heparin-like substance is used as the sulfated glucosaminoglycan, the heparin-like substance included in the Japanese Pharmacopoeia standard is suitable.

The blending amount of the sulfated glucosaminoglycan in the ceramide synthesis accelerator of the present invention may be appropriately set according to the formulation form of the ceramide synthesis accelerator, but is, for example, 0.01 to 5% by weight, preferably The amount is 0.05 to 3% by weight, and more preferably 0.1 to 1% by weight.
Other Components In addition to the sulfated glucosaminoglycan, the ceramide synthesis promoter of the present invention may contain other pharmacological components, if necessary. Such pharmacological components include, for example, antihistamines (dipotassium glycyrrhizinate, glycyrrhizic acid, diphenhydramine, diphenhydramine hydrochloride, etc.), local anesthetics (procaine, tetracaine, bupipacaine, mepipacaine, chloroprocaine, proparacaine, meprilcaine or salts thereof. , Orthocaine, oxesazein, oxypolyentoxydecane, roto extract, percamine pase, tesitdecitin, etc.), anti-inflammatory agents (allantoin, indomethacin, felbinac, diclofenac sodium, loxoprofen sodium, etc.), skin protectants (colodion, castor oil, etc.), blood circulation promotion Ingredients (nonyl acid vanillyl amide, nicotinic acid benzyl ester, capsaicin, capsicum extract, etc.), cooling agents (menthol, camphor, etc.), vitamins (vitamin A, etc.), mucopolysaccharides (sodium chondroitin sulfate, glucosamine, etc.), etc. Be done.

  Further, the ceramide synthesis accelerator of the present invention may contain a base or an additive, if necessary, in order to obtain a desired formulation form. Such bases and additives are not particularly limited as long as they are pharmaceutically acceptable. For example, water, lower alcohol (ethanol, isopropanol, etc.), polyhydric alcohol (glycerin, propylene glycol, dipropylene) Aqueous bases such as glycol, 1,3-butylene glycol, etc.; oils (olive oil, safflower oil, soybean oil, camellia oil, corn oil, rapeseed oil, sunflower oil, cottonseed oil, peanut oil, lard, squalane, fish oil, etc. ), mineral oil (liquid paraffin, paraffin, gelled hydrocarbon, petrolatum, etc.), waxes and waxes (beeswax, carnauba wax, candelilla wax, ceresin, rice wax, microcrystalline wax, etc.), ester oil (isopropyl myristate) , Isopropyl adipate, diethyl sebacate, isopropyl sebacate, isopropyl palmitate, cetyl palmitate, ethyl oleate, etc.), fatty acid alkyl esters, fatty acids (stearic acid, oleic acid, palmitic acid, behenic acid, linoleic acid, lanolin, etc.) ), fatty acid ester (cetyl palmitate, isopropyl palmitate, ethyl linoleate, etc.), higher alcohol (stearyl alcohol, cetanol, behenyl alcohol, myristyl alcohol, oleyl alcohol, hexadecyl alcohol, lanolin alcohol, etc.), cholesterol, tri-2-ethyl Oily bases such as glyceryl hexanoate, cetyl 2-ethylhexanoate, silicone oil (dimethylpolysiloxane, cyclic silicone, etc.); POE (10 to 50 mol) phytosterol ether, POE (10 to 50 mol) dihydrocholesterol ether, POE (10-50 mol) 2-octyldodecyl ether, POE (10-50 mol) decyl tetradecyl ether, POE (10-50 mol) oleyl ether, POE (2-50 mol) cetyl ether, POE (5-50 mol) ) Polyoxyethylene such as behenyl ether, POE (5 to 30 mol) polyoxypropylene (5 to 30 mol) 2-decyltetradecyl ether, POE (10 to 50 mol) polyoxypropylene (2 to 30 mol) cetyl ether Alkyl ethers, phosphoric acid/phosphate salts thereof (POE cetyl ether sodium phosphate, etc.), POE (20-60 mol) sorbitan monooleate, POE (10-60 mol) sorbitan monoisostearate, POE (10 80 mol) glyceryl monoisostearate, POE (10 to 30 mol) glyceryl monostearate, POE (20 to 100 mol)/polyoxypropylene modified silicone, POE/alkyl modified silicone, polyethylene glycol monolaurate, polyethylene monopalmitate Glycol, polyethylene glycol monostearate, polyethylene glycol dilaurate, polyethylene glycol dipalmitate, polyethylene glycol distearate, polyethylene glycol dioleate, polyethylene glycol diricinoleate, polyoxyethylene hydrogenated castor oil (5-100), polysorbate (20 ~ 85), glycerin fatty acid ester (glycerin monostearate, etc.), hydrogenated soybean phospholipid, hydrogenated lanolin alcohol, etc.; refreshing agent (menthol, camphor, borneol, peppermint water, peppermint oil, etc.), preservative Agents (methylparaben, propylparaben, benzoic acid, sodium benzoate, sorbic acid, etc.), flavoring agents (citral, 1,8-cionele, citronellal, farnesol, etc.), colorants (tar dyes (brown 201, blue 201) , Yellow No. 4, Yellow No. 403, etc.), cacao dye, chlorophyll, aluminum oxide, etc.), a thickening agent (carboxyvinyl polymer, hypromellose, polyvinylpyrrolidone, sodium alginate, ethyl cellulose, sodium carboxymethyl cellulose, xanthan gum, carrageenan, etc.), pH Adjusting agent (phosphoric acid, hydrochloric acid, citric acid, sodium citrate, succinic acid, tartaric acid, sodium hydroxide, potassium hydroxide, triethanolamine, triisopropanolamine, etc.), wetting agent (dl-pyrrolidonecarboxylate sodium liquid, D -Sorbitol liquid, macrogol, etc.), stabilizers (dibutylhydroxytoluene, butylhydroxyanisole, sodium edetate, sodium metaphosphate, L-arginine, L-aspartic acid, DL-alanine, glycine, sodium erythorbate, gallic acid Propyl, sodium sulfite, sulfur dioxide, chlorogenic acid, catechin, rosemary extract, etc.), antioxidant, UV absorber, chelating agent, adhesive, buffer, solubilizer, solubilizer, preservative, etc. Agents.

Formulation form The ceramide synthesis accelerator of the present invention may be any dosage form such as a skin external preparation and an internal preparation, but from the viewpoint of further effectively promoting ceramide synthesis, a skin external preparation is preferably used. Can be mentioned.

  When the ceramide synthesis accelerator of the present invention is used as an external preparation for skin, its shape is not particularly limited as long as it can be applied transdermally, for example, liquid, solid, semisolid (gel, ointment, Paste form) and the like.

  Further, when the ceramide synthesis accelerator of the present invention is used as a skin external preparation, its formulation form is not particularly limited as long as it can be applied transdermally, for example, skin external medicine, skin external quasi drug, cosmetic And skin cleansing agents. As a formulation form when the ceramide synthesis promoter of the present invention is used as an external preparation for skin, specifically, creams, lotions, gels, emulsions, liquids, patches, aerosols, ointments, packs, etc. External drug for skin; cream, lotion, gel, emulsion, liquid, patch, external quasi-drug such as aerosol, ointment, pack; cream, lotion, gel, emulsion Cosmetics such as liquids, ointments and packs; skin cleansing agents such as body shampoos, hair shampoos and conditioners. Among these formulation forms, skin external medicines are preferable, and creams, lotions, gels, emulsions and packs are more preferable.

Use/Dose Since the ceramide synthesis promoter of the present invention can promote the expression of serine palmitoyltransferase in epidermal cells (particularly, cells existing in the upper layer of epidermis), it is used for promoting ceramide synthesis in epidermis. Further, the promotion of ceramide synthesis in the epidermis is known to be effective in the prevention or treatment of skin disorders such as atopic dermatitis, ichthyosis, acne, skin immune disorders, and skin infections caused by viruses and bacteria. The ceramide synthesis promoter of the present invention can be used for the purpose of preventing or treating these skin disorders.

Further, the dose of the ceramide synthesis accelerator of the present invention may be appropriately set depending on the dosage form, the dosage form, the degree of symptoms to be applied, and the like. For example, when the ceramide synthesis promoter of the present invention is used as a skin external preparation, one example of the dose is about 0.1 to 3 mg of sulfated glucosaminoglycan per 1 cm ^{2 of} skin. The amount may be 1 to several times a day.

2. Serine palmitoyl transferase expression promoter As described above, the sulfated glucosaminoglycan has an action of promoting the expression of serine palmitoyl transferase, and therefore the present invention further comprises a sulfated glucosaminoglycan. A featured serine palmitoyltransferase expression promoter is provided. In the serine palmitoyl transferase expression promoter of the present invention, the sulfated glucosaminoglycan used, other components that can be mixed, the formulation form, the use, the dose, etc. are described in the above "1. Ceramide synthesis promoter" column. As described.

  Hereinafter, the present invention will be described in more detail with reference to Examples, but the present invention is not limited thereto.

Test Example 1: Effect of sulfated glucosaminoglycan on ceramide synthesis The following experiment was performed to evaluate the effect of sulfated glucosaminoglycan on the ceramide synthesis ability of epidermal cells.

1. Experimental material (cell)
In this test example, the following two types of cells were used.
(A) Normal epidermal model cell (HaCaT cell)
As normal epidermal model cells, HaCaT cells (human epidermal keratinocytes (Deutsches Krebsforschungszentrum, Stiftung desofentrichen Rechts (DKFZ), Heidelberg, Germany)) were used.
(B) Epidermal upper layer model cell (HaCaT cell highly expressing syntaxin 4)
A cell model of the upper layer of the epidermis with a certain degree of differentiation was prepared and used for the test. (I) Epidermal cells migrate from the basal layer to the spinous layer/granular layer when keratinized, (ii) secretin 4 is extracellularly secreted as it moves to the upper layer, and (iii) HaCaT cells It is known that keratinization progresses when syntaxin 4 is highly expressed in K. (Kadono N. et al., Mol Med 2015). Therefore, HaCaT cells highly expressing syntaxin 4 were prepared as epidermal model cells and used in this test example. The manufacturing method is as follows.
First, a gene encoding a mouse-derived IL-2 signal peptide consisting of 21 amino acid residues is added to the N-terminus of mouse syntaxin 4 gene in order to forcefully display it extracellularly, and this is added to the multi-chain of pQCXIN retroviral vector. It was inserted into the cloning site. Next, this was introduce|transduced into the packaging cell PT67 using the transfection reagent Lipofectamine PLUS (Thermo Fisher Scientific Co., Ltd. make). A retrovirus was obtained from the culture supernatant of the obtained cells. By infecting the HaCaT cells with the obtained virus, HaCaT cells (epithelial upper layer model cells) in which syntaxin 4 was forcibly expressed extracellularly were prepared.

(Culture medium)
D-MEM/Ham's F-12 (Wako Pure Chemical Industries, Ltd.)/10% FCS addition

(Test substance)
Heparin analogue

2. Experimental Method (1) Expression level of serine palmitoyltransferase mRNA in normal epidermal model cells Model cells were seeded in each well of a 12-well plate at a concentration of 1×10 ⁵ cells/well, and 37° C. under 5% CO ₂ conditions. It was cultured in. The next day, a heparin-like substance was added to each well at 0.5 mg/mL, and the wells were cultured at 37° C. under 5% CO ₂ . On the next day, model cells were collected and total RNA was extracted using RNeasy mini kit (manufactured by Qiagen Co., Ltd.). Then, reverse transcription was performed using ReverseTra Ace (manufactured by Toyobo Co., Ltd.) to synthesize cDNA. Next, serine palmitoyl transferase genes (SPTLC1 and SPTLC3) were amplified by qRT-PCR using Fast Start Essential DNA Green Master on LightCycler Nano system (manufactured by Roche Diagnostics KK). β-actin was used as an internal standard to standardize the expression level of serine palmitoyltransferase mRNA. The expression level of this mRNA was divided by the expression level (control) of mRNA confirmed in the same manner as above except that the heparin-like substance was not added.

(2) Expression level of serine palmitoyltransferase mRNA in upper epidermis model cells Except for using epidermal upper layer model cells in place of normal epidermal model cells, the same procedure as in (1) above was performed to determine the expression level of serine palmitoyltransferase mRNA. It was confirmed and divided by the control (mRNA expression level when no heparin-like substance was added).

  The results obtained are shown in FIGS. As can be seen from FIGS. 1 and 2, an increase in the expression level of serine palmitoyltransferase mRNA was observed in both normal epidermal model cells and epidermal model cells under conditions where heparin-like substances were added. The increase in the expression level was remarkable. That is, from this result, the sulfated glucosaminoglycans such as heparin-like substances were found to express serine palmitoyltransferase mRNA in epidermal cells, particularly cells in the upper layer of the epidermis (cells directly affected by the administered sulfated glucosaminoglycans). It was found that it has the effect of increasing the amount and can promote the synthesis of ceramide. There was no significant difference in the expression level of mRNA between the control in the normal epidermal model cells and the control in the epidermal upper layer model cells.

Prescription examples Creams (prescription examples 1 to 8) having the compositions shown in Tables 1 and 2, lotions (prescription examples 9 to 14) shown in Table 3, gel agents (prescription examples 15 to 19) shown in Table 4, and tables. The emulsions 5 to 6 (formulation examples 20 to 29) were prepared. All of these preparations are expected to have an effect of increasing the expression level of serine palmitoyltransferase mRNA in epidermal cells, as in the case of Test Example 1, and are effective in promoting ceramide synthesis.

Claims (3)

Containing sulfated glucosaminoglycans, skin external agent for use in the prevention or treatment of Niki bi (except. Where used in combination with adapalene), characterized in that a, ceramide synthesis promoter.   The ceramide synthesis promoter according to claim 1, wherein the sulfated glycosaminoglycan is a heparin-like substance. Containing sulfated glucosaminoglycans, skin external agent for use in the prevention or treatment of Niki bi (except. Where used in combination with adapalene) characterized in that it is a serine palmitoyltransferase expression promoting agent .