Peptides with hair growth promoting and hair loss inhibiting activity and their uses

Abstract

The present invention relates to a peptide having the activity of promoting hair growth or preventing or suppressing alopecia. The peptide of the present invention promotes the proliferation of hair follicle dermal papilla cells (HFDPC) and human outer root sheath cells (HHORSC), upregulates the expression of growth factors in hair follicle dermal papilla cells (HFDPC), and downregulates the expression level of DKK-1 (Dickkopf-1), a hair loss protein.

Description

[Technical Field]

[0001] The present invention relates to a peptide having hair growth promoting and hair loss inhibiting activity and uses thereof. [Background technology]

[0002] Alopecia is a phenomenon in which excessive hair loss occurs due to a longer than normal resting phase in the hair growth cycle or an increase in the number of hair follicles in the resting phase. It refers to excessive hair loss that does not recover due to stress, nutritional deficiencies, local blood flow disorders, hormones, genetic factors, etc.

[0003] As hair loss treatment drugs, 5α-reductase inhibitors, such as finasteride and dutasteride, which act to convert the male hormone testosterone into the androgenic hormone dihydrotestosterone (DHT), have been developed and approved for sale. Minoxidil, a drug that stimulates hair follicles, increases blood flow, and promotes hair growth, has also been approved and is in use.

[0004] However, 5α-reductase inhibitors are known to frequently cause side effects such as sexual dysfunction, depression, anxiety, and breast enlargement, while minoxidil is known to cause cardiovascular side effects such as electrocardiogram abnormalities, tachycardia, and endocarditis. Furthermore, minoxidil is known to cause hair loss again if the drug is discontinued.

[0005] Peptides have high biocompatibility, and when used as a single active ingredient in drugs or cosmetics, they have fewer side effects than extracts containing various ingredients. After they have a localized hair loss inhibitory effect, they can be easily broken down and removed by degradative proteins in the body, so there is less risk of side effects compared to existing treatments.

[0006] PCT International Patent WO2005-082395 discloses a peptide that promotes hair growth by accelerating the progression from the resting stage to the growth stage in the hair growth cycle, and Korean Patent Registration No. 10-2387764 discloses a dimeric peptide derivative that promotes the proliferation of hair follicle cells. [Prior art documents] [Patent documents]

[0007] [Patent Document 1] WO2005-082395 [Patent Document 1] Korean Patent Registration No. 10-2387764 Summary of the Invention [Problem to be solved by the invention]

[0008] The present inventors have conducted research efforts to develop peptides with improved activity that are effective in promoting hair growth and inhibiting hair loss with fewer side effects. As a result, the inventors have experimentally demonstrated that the novel peptides they have developed promote the proliferation of human hair follicle dermal papilla cells (HHFDPCs) and human outer root sheath cells (HHORSCs), increase the expression levels of growth factors bFGF, KGF, and VEGF in dermal papilla cells, and inhibit the expression of the hair loss protein DKK-1 induced by dihydrotestosterone (DHT), thereby completing the present invention.

[0009] Therefore, an object of the present invention is to provide a novel peptide having hair growth promoting and hair loss inhibiting activities.

[0010] Another object of the present invention is to provide a composition for promoting hair growth or suppressing, preventing or ameliorating alopecia, which contains a peptide having the above-mentioned activity as an active ingredient.

[0011] It is yet another object of the present invention to provide a pharmaceutical composition for preventing or treating alopecia.

[0012] A further object of the present invention is to provide a cosmetic composition for promoting hair growth or preventing or ameliorating alopecia. [Means for solving the problem]

[0013] In order to achieve the above purpose, One aspect of the present invention provides a peptide comprising the amino acid sequence of SEQ ID NO:1.

[0014] Another aspect of the present invention provides a composition for promoting hair growth or suppressing, preventing, or ameliorating alopecia, comprising the peptide as an active ingredient.

[0015] Another aspect of the present invention provides a pharmaceutical composition for preventing or treating alopecia, comprising the peptide as an active ingredient.

[0016] Yet another aspect of the present invention provides a cosmetic composition for promoting hair growth or preventing or ameliorating alopecia, which contains the peptide as an active ingredient.

[0017] The present invention will be described in detail below.

[0018] 1. Peptides and their activities According to one aspect of the present invention, there is provided a peptide comprising the amino acid sequence disclosed in SEQ ID NO:1.

[0019] As used herein, the term "peptide" refers to a linear molecule formed by amino acid residues joined together by peptide bonds.

[0020] The peptide of the present invention comprising the amino acid sequence of SEQ ID NO: 1 may be used without modification, but amino acid variants or fragments having different sequences due to deletion, insertion, substitution, or a combination thereof of amino acid residues may also be used, as long as the original activity of the peptide, such as the activity of promoting hair growth or inhibiting alopecia, is not affected.

[0021] The peptides of the present invention may be modified by phosphorylation, sulfation, acrylation, glycosylation, methylation, farnesylation, etc., within the range that does not change their activity.

[0022] The peptides of the present invention include peptides comprising an amino acid sequence substantially identical to that of a peptide comprising the amino acid sequence of SEQ ID NO: 1, as well as variants thereof or active fragments thereof. The term "substantially identical amino acid sequence" refers to an amino acid sequence having 75% or more, for example, 80% or more, 85% or more, 90% or more, 95% or more, or 97% or more sequence identity with the amino acid sequence of SEQ ID NO: 1. The peptides may additionally include a targeting sequence, a tag, a labeled residue, or an amino acid sequence specifically engineered to increase half-life or peptide stability.

[0023] The peptides of the present invention may be modified at the N- and / or C-terminus by selecting a portion of the amino acid sequence to increase their activity. Such N- and / or C-terminal modifications can significantly improve the stability of the peptides of the present invention, for example, increasing the half-life of the peptides when administered in vivo. The term "stability" refers not only to in vivo stability, which protects the peptides of the present invention from attack by in vivo protease enzymes, but also to storage stability (e.g., storage stability at room temperature).

[0024] The N-terminal modification may be a protecting group selected from the group consisting of an acetyl group, a fluorenylmethoxycarbonyl group, a formyl group, a palmitoyl group, a myristyl group, a stearyl group, and polyethylene glycol (PEG) attached to the N-terminus of the peptide. The C-terminal modification may be a protecting group such as a hydroxyl group (-OH), an amino group (-NH), or an azide (-NHNH) attached to the C-terminus of the peptide, but is not limited thereto.

[0025] The peptides of the present invention can be prepared by various methods well known in the art, including chemical synthesis techniques well known in the art, particularly solid-phase synthesis techniques (Merrifield, J. Amer. Chem. Soc. 85:2149-54 (1963); Stewart, et al., Solid Phase Peptide Synthesis, 2nd ed., Pierce Chem. Co.: Rockford, 111 (1984)) or liquid-phase synthesis techniques (U.S. Patent No. 5,516,891).

[0026] The peptide of the present invention has an activity of promoting hair growth or preventing or suppressing alopecia.

[0027] In one embodiment, the peptide of the present invention promotes proliferation of hair follicle dermal papilla cells (HFDPC).

[0028] In one embodiment, the peptide of the present invention promotes the proliferation of human hair outer root sheath cells (HHORSCs).

[0029] In one embodiment, the peptide of the present invention upregulates the expression of a growth factor in hair follicle dermal papilla cells (HFDPCs), which may be one or more selected from the group consisting of basic fibroblast growth factor (bFGF), keratinocyte growth factor (KGF), and vascular endothelial growth factor (VEGF).

[0030] In one embodiment, the peptide of the present invention downregulates the expression level of DKK-1 (Dickkopf-1), a hair loss protein, in hair follicle dermal papilla cells (HFDPCs). Dihydrotestosterone (DHT) in hair follicles promotes the expression of DKK-1 (Dickkopf-1), and DHT-induced DKK-1 is known to inhibit the proliferation of hair follicle keratinocytes and induce their apoptosis, promoting hair loss (J of Invest Dermatol. 2008, Feb., 128(2), pp. 262-269). The expression level of the hair loss protein DKK-1 is increased by dihydrotestosterone (DHT), and the peptide of the present invention can downregulate the DHT-induced increase in DKK-1 expression.

[0031] The peptide of the present invention described above has the above-mentioned activity, and thus can exhibit the effect of promoting hair growth or suppressing, preventing, or ameliorating alopecia.

[0032] 2. Composition for promoting hair growth or preventing, treating or improving alopecia In another aspect of the present invention, there is provided a composition for promoting hair growth or preventing, treating or ameliorating alopecia, which comprises a peptide comprising the amino acid sequence of SEQ ID NO: 1 as an active ingredient.

[0033] As described above, the peptide of the present invention comprising the amino acid sequence of SEQ ID NO: 1 has the activity of promoting hair growth or suppressing or preventing alopecia.

[0034] In another aspect of the present invention, the composition of the present invention may be a pharmaceutical composition for preventing or treating alopecia, which comprises a peptide comprising the amino acid sequence of SEQ ID NO: 1 as an active ingredient.

[0035] As used herein, the term "hair loss" or "alopecia" refers to a condition in which abnormal hair loss results in the absence of hair in areas where hair normally exists, or the number of hairs is reduced compared to normal. In one embodiment, hair loss or symptoms of hair loss in the present invention include hair loss on the scalp.

[0036] In the present invention, hair loss includes, but is not limited to, hereditary androgenetic alopecia (male pattern hair loss), female pattern hair loss, telogen effluvium, anagen effluvium, alopecia areata, tinea capitis due to fungal infection, hair loss due to hypothyroidism or hyperthyroidism, hair loss due to inflammation (seborrheic scalp), hair loss due to malnutrition, hair loss due to diabetes, hair loss due to lupus, hair loss due to hair production disorders, hair loss due to drugs (anticoagulants, antidepressants, discontinuation of oral contraceptives, chemotherapy), toxic folliculitis, lichen planus, hair loss due to burns or trauma, hair loss due to ultraviolet rays, and hair loss due to contact with airborne fine particulate matter.

[0037] In one embodiment, a pharmaceutical composition comprising the peptide of the present invention as an active ingredient promotes the proliferation of hair follicle dermal papilla cells (HFDPC).

[0038] In one embodiment, a pharmaceutical composition comprising the peptide of the present invention as an active ingredient promotes the proliferation of human hair outer root sheath cells (HHORSCs).

[0039] In one embodiment, a pharmaceutical composition comprising the peptide of the present invention as an active ingredient upregulates the expression of growth factors in hair follicle dermal papilla cells (HFDPCs), which may be one or more selected from the group consisting of basic fibroblast growth factor (bFGF), keratinocyte growth factor (KGF), and vascular endothelial growth factor (VEGF).

[0040] In one embodiment, a pharmaceutical composition containing the peptide of the present invention as an active ingredient down-regulates the expression level of DKK-1 (Dickkopf-1), a hair loss protein, in hair follicle dermal papilla cells (HFDPCs). The expression level of DKK-1, a hair loss protein, is increased by dihydrotestosterone (DHT), and the peptide of the present invention can down-regulate the DHT-induced increased DKK-1 expression level.

[0041] The pharmaceutical compositions of the present invention may comprise a therapeutically effective amount of the peptides of the present invention.

[0042] The term "therapeutically effective amount" means an amount sufficient for the peptide, which is the active ingredient of the pharmaceutical composition of the present invention, to achieve its activity or efficacy, for example, an amount sufficient for achieving efficacy in treating or preventing alopecia.

[0043] As used herein, the term "prevention" means reducing the risk of contracting a disease or disorder, and refers to any action that inhibits or delays the onset of a disease by preventing the disease or one or more of its clinical symptoms from progressing.

[0044] As used herein, the term "treatment" means alleviating a disease or disorder and includes any action that prevents or reduces the progression of a disease or one or more of its clinical symptoms, thereby improving or modifying the symptoms of the disease.

[0045] In the present invention, the prevention or treatment of hair loss may be achieved by removing the cause of hair loss or inhibiting the progression of hair loss, or by inhibiting hair loss or promoting hair formation to promote hair growth.

[0046] The pharmaceutical composition of the present invention may comprise a pharmaceutically acceptable carrier.

[0047] The pharmaceutically acceptable carriers are those commonly used in formulations, and include, but are not limited to, lactose, dextrose, sucrose, sorbitol, mannitol, starch, gum arabic, calcium phosphate, alginate, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrup, methylcellulose, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and mineral oil.

[0048] The pharmaceutical composition of the present invention may further contain, in addition to the above ingredients, lubricants, wetting agents, sweeteners, flavoring agents, emulsifiers, suspending agents, preservatives and the like, but is not limited to these.

[0049] Suitable pharmaceutically acceptable carriers and formulations are described in detail in Remington: The Science and Practice of Pharmacy, (19th ed., 1995, Williams & Wilkins).

[0050] The pharmaceutical composition of the present invention may be administered by any suitable route for treating alopecia, for example, orally or parenterally, and in the case of parenteral administration, it may be administered by intravenous injection, subcutaneous injection, intramuscular injection, intraperitoneal injection, topical administration, transdermal administration, etc. Since the pharmaceutical composition of the present invention has an activity for preventing or treating alopecia, it may be applied by topical administration, such as by application to the scalp.

[0051] The dosage of the pharmaceutical composition may be, but is not limited to, 0.0001 μg to 100 mg, 0.001 μg to 100 mg, 0.01 μg to 100 mg, 0.1 μg to 100 mg, or 1.0 μg to 1000 mg per day, and may be variously prescribed depending on factors such as formulation method, administration method, age, weight, sex, pathological condition, diet of the patient, administration time, administration route, excretion rate, and reaction sensitivity.

[0052] The pharmaceutical composition of the present invention may be prepared in a unit dose form or in a multi-dose container by formulating it with pharmaceutically acceptable carriers and / or excipients by a method easily performed by a person skilled in the art to which the invention pertains. In this case, the dosage form may be a solution, suspension, or emulsion in an oil or aqueous medium, or may be in the form of an extract, powder, granules, tablets, or capsules, and may further contain a dispersing agent or stabilizer.

[0053] The pharmaceutical composition of the present invention may be a topical skin preparation. The topical skin preparation is a formulation that can be applied externally to the skin. When the pharmaceutical composition of the present invention is used as a topical skin preparation, it may be applied to the scalp, specifically, to the scalp at a site where hair loss has occurred or the scalp at a site where hair growth is desired to be promoted. The topical skin preparation may be a cream, gel, ointment, skin emulsifier, skin suspension, transdermal delivery patch, drug-containing bandage, lotion, or a combination thereof. The topical skin preparation may be appropriately blended with ingredients commonly used in topical skin preparations such as cosmetics and pharmaceuticals, for example, aqueous ingredients, oily ingredients, powder ingredients, alcohols, moisturizers, thickeners, UV absorbers, whitening agents, preservatives, antioxidants, surfactants, fragrances, colorants, various skin nutrients, or combinations thereof, as needed. The topical skin preparation may also contain, as appropriate, sequestering agents such as disodium edetate, trisodium edetate, sodium citrate, sodium polyphosphate, sodium metaphosphate, and gluconic acid; drugs such as caffeine, tannin, licorice extract, glabridin, various herbal medicines, tocopherol acetate, glycyrrhizin, tranexamic acid, and derivatives or salts thereof; vitamin C, magnesium ascorbyl phosphate, ascorbyl glucoside, arbutin, kojic acid, and sugars such as glucose, fructose, and trehalose.

[0054] In another aspect of the present invention, the composition of the present invention may be a cosmetic composition for promoting hair growth or preventing or ameliorating alopecia, which comprises a peptide comprising the amino acid sequence of SEQ ID NO: 1 as an active ingredient.

[0055] As used herein, the term "amelioration" refers to any act that makes the symptoms of a disease or disorder better or less severe.

[0056] In one embodiment, a cosmetic composition containing the peptide of the present invention as an active ingredient promotes the proliferation of hair follicle dermal papilla cells (HFDPC).

[0057] In one embodiment, a cosmetic composition containing the peptide of the present invention as an active ingredient promotes the proliferation of human hair outer root sheath cells (HHORSCs).

[0058] In one embodiment, a cosmetic composition containing the peptide of the present invention as an active ingredient upregulates the expression of growth factors in hair follicle dermal papilla cells (HFDPCs), which may be selected from the group consisting of basic fibroblast growth factor (bFGF), keratinocyte growth factor (KGF), and vascular endothelial growth factor (VEGF).

[0059] In one embodiment, a cosmetic composition containing the peptide of the present invention as an active ingredient down-regulates the expression level of DKK-1 (Dickkopf-1), a hair loss protein, in hair follicle dermal papilla cells (HFDPCs). The expression level of DKK-1, a hair loss protein, is increased by dihydrotestosterone (DHT), and the peptide of the present invention can down-regulate the DHT-induced increased DKK-1 expression level.

[0060] The cosmetic composition may be prepared in any form commonly used in the technical field to which the present invention pertains, and may be a topical skin preparation, such as, but not limited to, a solution, suspension, emulsion, paste, gel, cream, lotion, powder, soap, surfactant-containing cleanser, oil, powder foundation, emulsion foundation, wax foundation, spray, etc.

[0061] The cosmetic composition may be prepared in various forms such as a solution, sol-gel, emulsion, oil, wax, aerosol, etc., including, but not limited to, a softening lotion, a nourishing lotion, a nourishing cream, a massage cream, an essence, an eye cream, a cleansing cream, a cleansing foam, a cleansing water, a pack, a spray, a powder, a hair tonic, a hair cream, a hair lotion, a hair shampoo, a hair rinse, a hair conditioner, a hair spray, a hair aerosol, a pomade, a gel, etc.

[0062] The cosmetic composition of the present invention may contain other additives such as excipients and carriers, and it is possible to apply and blend in the required amounts of common ingredients blended in general skin cosmetics.

[0063] When the cosmetic composition is in the form of a paste, cream, or gel, the carrier component may be animal oil, vegetable oil, wax, paraffin, starch, tragacanth, cellulose derivatives, polyethylene glycol, silicone, bentonite, silica, talc, zinc oxide, or the like.

[0064] When the cosmetic composition is in the form of a powder or spray, lactose, talc, silica, aluminum hydroxide, calcium silicate, or polyamide powder may be used as a carrier component, and particularly when the cosmetic composition is in the form of a spray, a propellant such as chlorofluorohydrocarbon, propane / butane, or dimethyl ether may be further included, but is not limited thereto.

[0065] When the cosmetic composition is in the form of a solution or emulsion, a solvent, solubilizer, or emulsifier may be used as a carrier component, and examples of such solvents include water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butyl glycol oil, glycerol aliphatic esters, polyethylene glycol, and sorbitan fatty acid esters.

[0066] When the cosmetic composition is in the form of a suspension, a liquid diluent such as water, ethanol, or propylene glycol, a suspending agent such as ethoxylated isostearyl alcohol, polyoxyethyl sorbitol ester, or polyoxyethylene sorbitan ester, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar, or tragacanth may be used as a carrier component.

[0067] When the cosmetic composition is in the form of a surfactant-containing cleanser, the carrier component may be a fatty alcohol sulfate, a fatty alcohol ether sulfate, a sulfosuccinic acid monoester, isethionic acid, an imidazolinium derivative, methyl taurate, sarcosinate, a fatty acid amide ether sulfate, an alkylamidobetaine, a fatty alcohol, a fatty acid glyceride, a fatty acid diethanolamide, a vegetable oil, a lanolin derivative, or an ethoxylated glycerol fatty acid ester.

[0068] When the cosmetic composition is in the form of a hair shampoo, the peptide of the present invention may be mixed with base ingredients for forming a shampoo, such as a thickener, surfactant, viscosity modifier, moisturizer, pH adjuster, preservative, essential oil, etc. The thickener may be CDE, the surfactant may be an anionic surfactant LES or an amphoteric surfactant cocobetaine, the viscosity modifier may be polyquaternary, the moisturizer may be glycerin, and the pH adjuster may be citric acid or sodium hydroxide. The preservative may be grapefruit extract, and in addition, essential oils such as cedarwood, peppermint, and rosemary, silk amino acids, pentaol, or vitamin E may be added.

[0069] The components contained in the cosmetic composition may further include, in addition to the peptide of the present invention as an active ingredient and a carrier component, components commonly used in cosmetic compositions, such as, for example, common adjuvants such as antioxidants, stabilizers, solubilizers, vitamins, pigments, and fragrances, but are not limited thereto.

[0070] The peptide of the present invention may be contained in the above-mentioned composition at a concentration of 0.01 uM to 1000 uM. Specifically, the peptide of the present invention may be contained in a concentration of 0.01 uM to 1000 uM; 0.05 uM to 800 uM, 0.05 uM to 700 uM, 0.05 uM to 600 uM, 0.05 uM to 500 uM, 0.05 uM to 300 uM, 0.05 uM to 200 uM; 0.1 uM to 800 uM, 0.1 uM to 700 uM, 0.1 uM to 600 uM , 0.1 uM to 500 uM, 0.1 uM to 300 uM, 0.1 uM to 200 uM; 1 uM to 800 uM, 1 uM to 700 uM, 1 uM to 600 uM, 1 uM to 500 uM, 1 uM to 300 uM, 1 uM to 200 uM; 5 uM to 800 uM, 5 uM to 700 uM, 5 uM to 600 uM, 5 uM to 500 uM, 5 uM to 300 uM, or 5 uM to 200 uM, but is not limited to these.

[0071] 3. Uses of the peptides of the present invention In another aspect of the present invention, there is provided a use of a peptide comprising the amino acid sequence of SEQ ID NO: 1 for promoting hair growth or suppressing, preventing, treating or ameliorating alopecia.

[0072] In another aspect of the present invention, there is provided a method for promoting hair growth or suppressing, preventing, treating or ameliorating alopecia, comprising administering a peptide comprising the amino acid sequence of SEQ ID NO: 1 or a composition comprising said peptide to a subject in need thereof.

[0073] In another aspect of the present invention, there is provided the use of a peptide comprising the amino acid sequence of SEQ ID NO: 1 for the manufacture of a medicament or cosmetic for promoting hair growth or preventing, treating or ameliorating alopecia. [Effects of the Invention]

[0074] The peptide of the present invention has the activity of promoting hair growth or preventing, suppressing or treating alopecia. The peptide of the present invention promotes the proliferation of hair follicle dermal papilla cells (HFDPC) and human outer root sheath cells (HHORSC), upregulates the expression of growth factors such as bFGF, KGF and VEGF in hair follicle dermal papilla cells (HFDPC), and downregulates the expression level of DKK-1 (Dickkopf-1), a hair loss protein.

[0075] However, the effects of the present invention are not limited to those mentioned above, and other effects not mentioned will be clearly understood by those skilled in the art from the following description. [Brief explanation of the drawings]

[0076] [Figure 1] 1 shows experimental results showing that the proliferation of HHFDPC cells is promoted when the peptide of the present invention is treated at 10 uM and 100 uM. [Figure 2] 1 shows experimental results showing that the proliferation of HHORSCs is promoted when HHORSC cells are treated with the peptide of the present invention at 50 μM and 100 μM. [Figure 3a] These are experimental results showing that when HHFDPC cells were treated with the peptide of the present invention at 10 μM, 50 μM, and 100 μM, the activity of HHFDPC increased, resulting in increased expression of growth factors such as bFGF, KGF, and VEGF. [Figure 3b] 3b is a graph showing the relative intensity values ​​of the bands in FIG. 3a. [Figure 4a] These are experimental results showing that when HHFDPC cells were treated with the peptide of the present invention at 10 μM, 50 μM, and 100 μM, the expression of DKK-1, a hair loss protein induced by DHT treatment, was reduced in a concentration-dependent manner. [Figure 4b] 4b is a graph showing the relative intensity values ​​of the bands in FIG. 4a. DETAILED DESCRIPTION OF THE INVENTION

[0077] The present invention will be described in more detail below with reference to examples. However, the following examples are provided to specifically illustrate the present invention, and the scope of the present invention is not limited to the following examples.

[0078] Production Example 1: Production of peptides and peptide complexes Peptides having the amino acid sequence of SEQ ID NO: 1 listed in Table 1 below were synthesized using an automated peptide synthesizer (Milligen 9050, Millipore, USA), and the synthesized peptides were purified and separated using C18 reverse-phase high-performance liquid chromatography (HPLC) (Waters Associates, USA). The column used was an ACQUITY UPLC BEH300 C18 (2.1 mm x 100 mm, 1.7 μm, Waters Co., USA).

[0079] [Table 1]

[0080] The efficacy of the prepared peptide of SEQ ID NO: 1 was evaluated through the following experiment.

[0081] Experimental Example 1: Promoting the proliferation of human dermal papilla cells (HHFDPC) The effect of the peptide prepared in Preparation Example 1 on the proliferation of human hair follicle dermal papilla cells (HHFDPC) was examined.

[0082] HHFDPC 4x10 3The cells were seeded into a 96-well cell culture plate at a density of 10 ... Finally, the absorbance was measured at a wavelength of 540 nm using a spectrophotometer (SpectraMax iD3, Molecular Devices (USA, CA)).

[0083] As a result of the experiment, as shown in FIG. 1, it was confirmed that when HHFDPCs were treated with the peptide of Preparation Example 1 at 10 μM and 100 μM, the proliferation of HHFDPCs was promoted compared to the negative control group (con).

[0084] Experimental Example 2: Promoting the proliferation of human outer root sheath cells (HHORSCs) The effect of the peptide prepared in Preparation Example 1 on the proliferation of human hair outer root sheath cells (HHORSCs) was examined.

[0085] 4x10 HHORSC 3The cells were seeded into a 96-well cell culture plate at a density of 1000 cells / well and cultured in mesenchymal stem cell complete media for 24 hours. The medium was then replaced with serum-free mesenchymal stem cell medium and cultured for an additional 24 hours. The cultured cells were treated with 50 μM or 100 μM of the peptides prepared in Preparation Example 1, 100 nM of EGF (epidermal growth factor) as a positive control, or an untreated negative control (con), and cultured for an additional 72 hours at 37°C. Next, 10 μL of 5 mg / ml MTT solution was added and incubated in a CO2 incubator at 37°C for 4 hours. After removing the medium, the cells were treated with 100 μL of DMSO and shaken for 10 minutes. Finally, the absorbance was measured at a wavelength of 540 nm using a spectrophotometer (SpectraMax iD3, Molecular Devices (USA, CA)).

[0086] As a result of the experiment, as shown in FIG. 2, it was confirmed that when HHORSCs were treated with the peptide of Preparation Example 1 at 50 μM and 100 μM, the proliferation of HHORSCs was promoted compared to the negative control group (con).

[0087] Experimental Example 3: Promoting growth factor expression in human dermal papilla cells The effect of the peptide prepared in Preparation Example 1 on the expression level of growth factors in human hair follicle dermal papilla cells (HHFDPC) was confirmed by RT-PCR.

[0088] HHFDPC 4x10 5The cells were seeded into 6-well cell culture plates at a density of 10 ... The PCR reaction products were electrophoresed on a 1.5% agarose gel, and bands were detected using a Bio-Rad Gel Image System.

[0089] [Table 2]

[0090] As a result of the experiment, as shown in Figures 3a and 3b, it was confirmed that when HHFDPC was treated with the peptide of Preparation Example 1 at 10uM, 50uM, and 100uM, the activity of HHFDPC increased and the expression of growth factors such as bFGF, KGF, and VEGF was promoted.

[0091] Experimental Example 4: Inhibition of the expression of hair loss protein DKK-1 in human dermal papilla cells Dihydrotestosterone (DHT) in hair follicles promotes the expression of DKK-1 (Dickkopf-1), and the DKK-1 produced by DHT is known to suppress the proliferation of keratinocytes in hair follicles, induce their apoptosis, and promote hair loss (J of Invest Dermatolo. 2008, Feb., 128(2), pp. 262-269.).

[0092] The effect of the peptide prepared in Preparation Example 1 on the expression level of DKK-1, a hair loss-inducing protein, in human dermal papilla cells (HHFDPC) was confirmed by Western blot.

[0093] HHFDPC 4x10 5The cells were seeded into 6-well cell culture plates at a density of 1000 cells / well and cultured in mesenchymal stem cell complete media for 24 hours. The medium was then replaced with serum-free mesenchymal stem cell media and cultured for an additional 24 hours. The cultured cell cultures were treated with 10 μM, 50 μM, or 100 μM of the peptides prepared in Preparation Example 1. A positive control group was treated with 5 μM finasteride (Fina), and an untreated group served as a negative control (con). DHT, an inducer of the hair loss protein DKK-1, was also treated at a concentration of 100 nM. The cell cultures treated with the peptides were cultured for an additional 24 hours at 37°C. The cultures were washed with PBS and then treated with a lysis buffer to lyse the cells and obtain cell lysates. The cell lysates were subjected to BCA quantification to prepare equal amounts of protein samples. The prepared protein samples were electrophoresed using 10% SDS-PAGE. Proteins separated by SDS-PAGE were transferred to a PVDF membrane and blocked with 5% skim milk for 30 minutes at room temperature. Antibody against DKK-1 (Cell Signaling, USA) was diluted 1:1000 with 3% BSA and incubated with the membrane for 16 hours at 4°C. The membrane was washed three times for 15 minutes each with 0.1% PBS-T (0.1% Tween-20 in PBS). A secondary antibody was added, diluted 1:2000 with 5% skim milk, and incubated for 1 hour at room temperature. The membrane was washed three times for 15 minutes each with 0.1% PBS-T (0.1% Tween-20 in PBS). After treatment with ECL solution (GE Healthcare, USA), the membrane was visualized on a film.

[0094] As a result of the experiment, as shown in Figures 4a and 4b, when HHFDPC was treated with the peptide of Preparation Example 1 at 10uM, 50uM, and 100uM, it was confirmed that the expression of DKK-1, a hair loss protein induced by DHT treatment, was reduced in a concentration-dependent manner.

[0095] Preparation Example 2: Preparation of pharmaceutical composition 2-1. Manufacturing of powders 2g of the peptide of the present invention Lactose 1g The ingredients were mixed and filled into an airtight bag to prepare a powder.

[0096] 2-2. Tablet manufacturing 100mg of the peptide of the present invention Corn starch 100mg Lactose 100mg Magnesium stearate 2mg The above ingredients were mixed and then compressed into tablets according to a conventional tablet manufacturing method.

[0097] 2-3. Capsule manufacturing 100mg of the peptide of the present invention Corn starch 100mg Lactose 100mg Magnesium stearate 2mg The above ingredients were mixed and then filled into gelatin capsules according to a conventional capsule manufacturing method to prepare capsules.

[0098] 2-4.Making of rounds Peptide of the present invention 1g Lactose 1.5g 1g glycerin Xylitol 0.5g The ingredients were mixed and then prepared in a conventional manner so that each pill weighed 4 g.

[0099] 2-5. Granule manufacturing 150mg of the peptide of the present invention Soybean extract 50mg Glucose 200mg Starch 600mg After mixing the ingredients, 100 mg of 30% ethanol was added and dried at 60°C to form granules, which were then filled into sachets.

[0100] Production Example 3: Production of cosmetic composition 3-1. Cream production Peptide of the present invention 4.6 parts by weight Cetostearyl alcohol 2.8 parts by weight Beeswax 2.6 parts by weight Stearic acid 1.4 parts by weight Lipophilic glycerin monostearate 2 parts by weight PEG-100 stearate 1 part by weight Sorbital sesquioleate 1.4 parts by weight Jojoba oil 4 parts by weight Squalene 3.8 parts by weight Polysorbate 60 1.1 parts by weight Macadamia oil 2 parts by weight Tocopherol acetate 0.2 parts by weight Methylpolysiloxane 0.4 parts by weight Ethylparaben 0.1 parts by weight Propylparaben 0.1 parts by weight Euxyl K-400 0.1 parts by weight 1,3-butylene glycol 7 parts by weight Methylparaben 0.05 parts by weight Glycerin 6 parts by weight d-Panthenol 0.2 parts by weight Triethanolamine 0.2 parts by weight pt41891 0.2 parts by weight p-H2O 46.05 parts by weight 3-2. Lotion manufacturing Peptide of the present invention 3.5 parts by weight Cetostearyl alcohol 1.6 parts by weight Stearic acid 1.4 parts by weight Lipophilic glycerin monostearate 1.8 parts by weight PEG-100 stearate 2.6 parts by weight Sorbital sesquioleate 0.6 parts by weight Squalene 4.8 parts by weight Macadamia oil 2 parts by weight Jojoba oil 2 parts by weight Tocopherol acetate 0.4 parts by weight Methylpolysiloxane 0.2 parts by weight Ethylparaben 0.1 parts by weight Propylparaben 0.1 parts by weight 1,3-butylene glycol 4 parts by weight Methylparaben 0.1 parts by weight Xanthan gum 0.1 parts by weight Glycerin 4 parts by weight d-Panthenol 0.15 parts by weight Allantoin 0.1 parts by weight Calcium carbonate (2% aq. Sol) 4 parts by weight Triethanolamine 0.15 parts by weight Ethanol 3 parts by weight pt41891 0.1 parts by weight p-H20 48.3 parts by weight 3-3. Manufacturing softening lotion Peptide of the present invention 0.2% by weight Ethanol 10.0% by weight Polyoxyethylene sorbitan polylaurate 1.0% by weight Methyl parahydroxybenzoate 0.2% by weight Glycerin 5.0% by weight 1,3-butyl glycol 6.0% by weight Incense (appropriate amount) Dye appropriate amount Purified water (appropriate amount) Total 100 3-4. Manufacturing of nutritional lotion Peptide of the present invention 0.1% by weight Vaseline 2.0% by weight Sorbitan sesquioleate 0.8% by weight Polyoxyethylene oleyl ethyl 1.2% by weight Methyl parahydroxybenzoate (appropriate amount) Propylene glycol 5.0% by weight Ethanol 3.2% by weight Carboxyvinyl polymer 18.0% by weight Dye appropriate amount Incense (appropriate amount) Purified water (appropriate amount) Total 100 3-5. Essence production Peptide of the present invention 5.0% by weight Propylene glycol 10.0% by weight Glycerin 10.0% by weight Sodium hyaluronate aqueous solution (1%) 5.0% by weight Ethanol 3.2% by weight Polyoxyethylene hydrogenated castor oil 1.0% by weight Methyl parahydroxybenzoate 0.1% by weight Incense (appropriate amount) Purified water (appropriate amount) Total 100 3-6. Pack manufacturing Peptide of the present invention 0.5% by weight Glycerin 5.0% by weight Propylene glycol 4.0% by weight Polyvinyl alcohol 15.0% by weight Ethanol 8.0% by weight Polyoxyethylene oleyl ethyl 1.0% by weight Methyl parahydroxybenzoate 0.2% by weight Incense (appropriate amount) Dye appropriate amount Purified water (appropriate amount) Total 100 The composition ratio is a mixture of suitable ingredients in a preferred embodiment, but the ingredients or the blending ratio may be arbitrarily modified depending on the demand class, demand country, use, regional and ethnic preferences, etc.

[0101] The above describes exemplary embodiments of the present application, but the scope of the present application is not limited to the specific embodiments described above, and a person with ordinary knowledge in the field can make appropriate modifications within the scope of the claims of the present application.

Claims

1. A peptide comprising the amino acid sequence of SEQ ID NO:

1.

2. A composition for promoting hair growth or suppressing, preventing, or improving alopecia, comprising the peptide according to claim 1 as an active ingredient.

3. A pharmaceutical composition for preventing or treating alopecia, comprising the peptide of claim 1 as an active ingredient.

4. The pharmaceutical composition for preventing or treating alopecia according to claim 3, wherein the peptide promotes proliferation of hair follicle dermal papilla cells (HFDPC) or hair outer root sheath cells (HORSC).

5. The peptide induces the growth of hair follicle dermal papilla cells (HFDPCs) by (i) upregulating the expression of one or more growth factors selected from the group consisting of bFGF, KGF, and VEGF; or (ii) The pharmaceutical composition for preventing or treating alopecia according to claim 3, which downregulates the expression level of DKK-1 (Dickkopf-1), a hair loss protein.

6. The pharmaceutical composition for preventing or treating alopecia according to claim 3, wherein the pharmaceutical composition is an external preparation for skin.

7. A cosmetic composition for promoting hair growth or preventing or ameliorating alopecia, comprising the peptide according to claim 1 as an active ingredient.

8. The cosmetic composition for promoting hair growth or preventing or improving alopecia according to claim 7, wherein the peptide promotes the proliferation of hair follicle dermal papilla cells (HFDPC) or hair outer root sheath cells (HORSC).

9. The peptide induces the growth of hair follicle dermal papilla cells (HFDPCs) by (i) upregulating the expression of one or more growth factors selected from the group consisting of bFGF, KGF, and VEGF; or (ii) The cosmetic composition for promoting hair growth or preventing or ameliorating alopecia according to claim 7, which downregulates the expression level of DKK-1 (Dickkopf-1), a hair loss protein.

10. The cosmetic composition for promoting hair growth or preventing or ameliorating alopecia according to claim 7 , wherein the cosmetic composition is a topical skin preparation.

11. 8. The cosmetic composition for promoting hair growth or preventing or improving alopecia according to claim 7, wherein the cosmetic composition is in one or more dosage forms selected from the group consisting of a solution, a suspension, an emulsion, a paste, a gel, a cream, a lotion, a powder, a soap, a surfactant-containing cleanser, an oil, a powder foundation, an emulsion foundation, a wax foundation, and a spray.

12. 8. The cosmetic composition for promoting hair growth or preventing or improving alopecia according to claim 7, wherein the cosmetic composition is in the form of one or more selected from the group consisting of hair tonic, hair cream, hair lotion, hair shampoo, hair rinse, hair conditioner, hair spray, hair aerosol, pomade, and gel.

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