Composition for improving skin condition comprising peptide complex as active ingredient

Abstract

The present invention relates to the use of peptide complexes for improving skin conditions. The peptide complex of the present invention has an activity of reducing skin cell injury by inhibiting various skin injury effects induced by an advanced glycosylation end product or a precursor thereof in skin cells, i.e., activation of inflammatory responses, promotion of apoptosis, increase in expression of MMP-1 and MMP-2, increase in expression of RAGE, and decrease in expression of glyoxalase.

Description

Cross-references to related applications

[0001] This application claims the benefit of priority based on Korean Patent Application No. 10-2023-0098514, filed on July 27, 2023, and all disclosures in the relevant Korean patent application are included as part of this specification. Technical Field

[0002] This invention relates to the use of peptide complexes for improving skin condition. Background Technology

[0003] The skin is the organ that covers all organs and tissues of the human body, protecting the body from external harm and playing an important role in maintaining homeostasis by retaining moisture and regulating body temperature. Skin aging occurs naturally with age, but it can also be induced by external factors such as ultraviolet radiation and environmental pollution.

[0004] Simultaneously, under hyperglycemic conditions, advanced glycation end products (AGEs) are formed through non-enzymatic reactions between reducing sugars (such as glucose) and proteins. Specifically, the carbonyl group of the reducing sugar reacts with the free amino group of a protein (such as lysine or arginine) to form an initial glycosylation product called a Schiff base. As the reaction proceeds, Amadori products are formed, and the resulting compounds subsequently undergo condensation, rearrangement, oxidation, cleavage, and cross-linking processes to produce stable and irreversible compounds known as AGEs.

[0005] It is known that AGEs produced are difficult to degrade in vivo and can damage organs or cells while remaining undegraded. In skin tissue, AGEs bind to collagen and elastin in the dermis, thereby inhibiting protein renewal, inducing tissue hardening, and reducing elasticity, which leads to wrinkle formation. Furthermore, when AGEs bind to receptors for advanced glycation end products (RAGEs) present on the surface of skin cells such as keratinocytes and fibroblasts, they are known to induce skin cell damage by generating reactive oxygen species (ROS), inflammatory responses, and activating apoptosis signaling pathways (Chen CY et al., 2022, Advanced Glycation End Products in the Skin: Molecular Mechanisms, Methods of Measurement, and Inhibitory Pathways, Front. Med. 9:837222. doi:10.3389 / fmed.2022.837222).

[0006] Synthetic agents developed as AGE formation inhibitors include aminoguanidine and pyridoxine (Diabetes, 2002 Sep; 51(9): 2826-32). However, these agents have been reported to cause toxicity with prolonged application, increasing concerns about their safety and efficacy when applied to the skin. Furthermore, various technologies have been developed related to extracts derived from natural products that possess AGE-inhibiting activity and can improve skin damage (WO 2020 / 071630). However, these natural product-derived extracts often fail to exhibit sufficient activity and efficacy, making it difficult to develop them into final products.

[0007] Therefore, there is a need to develop active substances that are safe and exhibit excellent activity against skin damage caused not only by various external factors, but also by advanced glycation end products and their precursors.

[0008] [Primary Technology] [Patent Literature] WO 2020 / 071630 [Non-patent literature] 1. Diabetes 2002 Sep; 51(9):2826-32 2. Front. Med. May 2022, Vol. 9:837222. Summary of the Invention Technical issues The inventors have researched and attempted to develop active substances to improve skin condition by preventing skin damage caused by AGE (advanced glycation end products). Experiments have demonstrated that the peptide complex according to this disclosure inhibits various cellular damage caused by AGE in skin cells, increases the expression of glyoxalase involved in AGE removal, and exhibits activity to improve skin condition.

[0009] Therefore, one aspect of the present invention provides a composition for improving skin condition.

[0010] Another aspect of the present invention provides a pharmaceutical composition for treating or preventing skin damage or skin diseases.

[0011] Another aspect of this disclosure provides a functional food composition for preventing or improving skin damage.

[0012] Technical solution In order to achieve these goals, One aspect of the present invention provides a composition for improving skin condition, comprising a peptide complex as an active ingredient, said peptide complex comprising: (i) a peptide comprising the amino acid sequence of SEQ ID NO.1; and (ii) a peptide comprising the amino acid sequence of SEQ ID NO.2.

[0013] Another aspect of the present invention provides a cosmetic composition for improving skin condition, comprising a peptide complex as an active ingredient.

[0014] Another aspect of the present invention provides a pharmaceutical composition for treating or preventing skin inflammation or skin diseases, comprising a therapeutically effective amount of a peptide complex.

[0015] Another aspect of the present invention provides a functional food composition for improving skin condition, comprising a peptide complex as an active ingredient.

[0016] The details of the invention will be described below.

[0017] Peptide complexes and their activities This invention relates to the use of peptide complexes for improving skin condition, said peptide complexes comprising: (i) a peptide comprising the amino acid sequence of SEQ ID NO.1; and (ii) a peptide comprising the amino acid sequence of SEQ ID NO.2.

[0018] According to the present invention, a peptide can refer to a linear molecule formed by amino acid residues linked together by peptide bonds.

[0019] According to the present invention, peptides comprising the amino acid sequence of SEQ ID NO.1 and peptides comprising the amino acid sequence of SEQ ID NO.2 can be used without modification. However, amino acid variants or fragments with different sequences obtained by deletion, insertion, or substitution of amino acid residues or combinations thereof may also be used, as long as the variant or fragment does not affect the original activity of the peptide, such as the activity of improving skin lesions.

[0020] The peptides according to the present invention can be modified by phosphorylation, sulfation, acrylate, glycosylation, methylation, farnesylation, etc., as long as the activity of the peptide is not changed.

[0021] The peptides according to the invention comprise peptides and variants thereof having substantially the same amino acid sequence as the peptide comprising the amino acid sequence of SEQ ID NO. 1 or the peptide comprising the amino acid sequence of SEQ ID NO. 2. The substantially identical amino acid sequence may refer to an amino acid sequence having at least 75%, for example, at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, or at least 99% sequence identity with the amino acid sequence of SEQ ID NO. 1 or SEQ ID NO. 2, respectively. Furthermore, the peptide may also include amino acid sequences designed for a specific purpose, such as targeting sequences, tags, labeled residues, or sequences that increase the half-life or stability of the peptide.

[0022] The peptides of the present invention can have their activity enhanced by N-terminal and / or C-terminal modification of a selected portion of their amino acid sequence. The stability of the peptides according to the present invention can be significantly improved by N-terminal and / or C-terminal modification. For example, the half-life can be increased when the peptide is administered in vivo. The term "stability" encompasses not only in vivo stability that protects the peptides according to the present invention from attack by protein-cleaving enzymes, but also storage stability (e.g., storage stability at room temperature).

[0023] N-terminal modification can be formed by binding a protecting group selected from the group consisting of acetyl, fluorenylmethoxycarbonyl, formyl, palmitoyl, myristyl, stearoyl, and polyethylene glycol (PEG) to the N-terminus of the peptide. C-terminal modification can be formed by forming a hydroxyl (-OH) or amino (-NH2) group, with an azide bound to the C-terminus of the peptide, but the invention is not limited thereto.

[0024] The peptides according to the invention can be prepared by a variety of methods widely known in the art to which this invention pertains. For example, the peptides according to the invention can be prepared using chemical synthesis methods known in the art, particularly solid-phase synthesis (Merrifield, J.A.Mer. 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 (US Patent No. 5,516,891).

[0025] The peptide complex according to the invention has anti-inflammatory activity in skin cells.

[0026] The peptide complex according to the invention can inhibit the activation of inflammatory responses in skin cells induced by AGE or AGE precursors (methylglyoxal (MGO)).

[0027] The peptide complex according to the present invention can inhibit the expression of inflammatory cytokines (TNF-α, IL-1β or IL-6) and the inflammatory regulator COX-2 in skin cells.

[0028] The peptide complex according to the invention can inhibit the increase in expression levels of TNF-α, IL-1β, IL-6 and COX-2 induced by AGE and AGE precursors.

[0029] The peptide complex according to the present invention has the activity of inhibiting apoptosis in skin cells.

[0030] The peptide complex according to the invention can inhibit apoptosis induced by AGE and AGE precursors.

[0031] The peptide complex according to the invention can inhibit the expression of cleaved PARP-1, cleaved caspase-3, cleaved caspase-9 or apoptosis-mediated protein Bax in skin cells, and can increase the expression of anti-apoptotic protein Bcl-2.

[0032] The peptide complex according to the invention can inhibit the increased expression of cleaved PARP-1, cleaved caspase-3, cleaved caspase-9 and Bax induced by AGE or its precursor in skin cells.

[0033] The peptide complex according to the invention can inhibit the reduction in expression of the anti-apoptotic protein (Bcl-2) induced by AGE and AGE precursor in skin cells.

[0034] The peptide complex according to the invention has the activity of inhibiting the expression of matrix metalloproteinases (MMPs) in skin cells.

[0035] The peptide complex according to the invention can inhibit the increase in matrix metalloproteinase (MMP) expression induced by AGE and AGE precursors in skin cells.

[0036] The peptide complex according to the invention can inhibit the increase in protein expression levels of MMP-1 and MMP-2 induced by AGE and AGE precursors in skin cells.

[0037] MMPs are proteolytic enzymes that specifically target collagen. Representative MMPs include MMP-1 and MMP-2. MMP-1 and MMP-2 degrade collagen, reducing the elasticity of skin tissue and causing wrinkles.

[0038] The peptide complex according to the invention has the activity of inhibiting RAGE expression in skin cells.

[0039] The peptide complex according to the present invention can inhibit the increase in RAGE expression levels induced by AGE and AGE precursors in skin cells.

[0040] AGE binds to AGE receptors on the surface of skin cells, producing ROS, inducing inflammatory responses, and activating apoptosis signaling pathways through AGE receptors, thereby inducing skin cell damage.

[0041] The peptide complex according to the invention inhibits the generation of ROS, the induction of inflammatory responses, or the activation of apoptosis induced by RAGE.

[0042] The peptide complex according to the invention has the activity of increasing the expression of glyoxalase-1 in skin cells.

[0043] The peptide complex according to the invention inhibits the decrease in glyoxalase-1 expression level induced by AGE precursor. Therefore, the peptide complex has the activity of increasing glyoxalase-1 expression.

[0044] Glyoxalase-1 is a protein that has the activity of degrading methylglyoxal, a precursor of AGE, and increasing the expression of glyoxalase-1 can inhibit the production of AGE.

[0045] Compositions for improving skin condition One aspect of the present invention provides a composition for improving skin condition comprising a peptide complex as an active ingredient, said peptide complex comprising: (i) a peptide comprising the amino acid sequence of SEQ ID NO.1; and (ii) a peptide comprising the amino acid sequence of SEQ ID NO.2.

[0046] The peptide complex with the above-described activity according to the present invention can exhibit excellent effects in improving skin condition.

[0047] According to the embodiments, improving skin condition through the composition for improving skin condition can refer to relieving skin inflammation, improving skin aging, promoting skin regeneration, improving skin elasticity, reducing skin wrinkles, improving the skin barrier, or improving skin damage.

[0048] According to embodiments of the present invention, improvement in skin condition can refer to improvement in skin condition caused by AGE and AGE precursor damage.

[0049] In the compositions of the present invention, the weight ratio of the peptide comprising the amino acid sequence of SEQ ID NO.1 to the peptide comprising the amino acid sequence of SEQ ID NO.2 is not particularly limited. For example, the weight ratio may include a weight ratio of 0.5:2 to 2:0.5, and may include a weight ratio of 0.5:1.5 to 1.5:0.5. Specifically, the weight ratio may include a weight ratio of 1:1.

[0050] According to embodiments, the composition for improving skin condition according to the present invention has at least one of the following activities (a) to (e): (a) Inhibit the expression of TNF-α, IL-1β or IL-6, or inhibit the expression of COX-2; (b) Inhibit the expression of cleaved PARP-1, cleaved caspase-3, cleaved caspase-9 or Bax, or increase the expression of Bcl-2. (c) Inhibit the expression of MMP-1 or MMP-2; (d) Inhibit the expression of receptor advanced glycation end products (RAGE); and (e) Increase the expression of glyoxalase-1.

[0051] The compositions according to the present invention have anti-inflammatory activity in skin cells.

[0052] The compositions according to the present invention can inhibit the activation of inflammatory responses in skin cells induced by AGE or AGE precursors (MGO, methylglyoxal).

[0053] The compositions according to the present invention can inhibit the expression of inflammatory cytokines TNF-α, IL-1β or IL-6 and the inflammatory regulator COX-2 in skin cells.

[0054] The compositions according to the present invention can inhibit the increase in expression levels of TNF-α, IL-1β, IL-6 and COX-2 induced by AGE and AGE precursors.

[0055] The composition according to the present invention has the activity of inhibiting apoptosis in skin cells.

[0056] The composition according to the invention can inhibit apoptosis induced by AGE and AGE precursors in skin cells.

[0057] The compositions according to the present invention can inhibit the expression of cleaved PARP-1, cleaved caspase-3, cleaved caspase-9 or apoptosis-mediated protein Bax in skin cells, and can increase the expression of anti-apoptotic protein Bcl-2 in skin cells.

[0058] The composition according to the invention can inhibit the increased expression of cleaved PARP-1, cleaved caspase-3, cleaved caspase-9 and Bax induced by AGE or its precursor in skin cells.

[0059] The composition according to the invention can inhibit the reduction in expression of the anti-apoptotic protein (Bcl-2) induced by AGE and AGE precursor in skin cells.

[0060] The composition according to the invention has the activity of inhibiting the expression of matrix metalloproteinases (MMPs) in skin cells.

[0061] The composition according to the invention can inhibit the increase in matrix metalloproteinase (MMP) expression induced by AGE and AGE precursors in skin cells.

[0062] The composition according to the invention can inhibit the increase in protein expression levels of MMP-1 and MMP-2 induced by AGE and AGE precursors in skin cells.

[0063] MMPs are proteolytic enzymes that specifically target collagen. Representative MMPs include MMP-1 and MMP-2. MMP-1 and MMP-2 degrade collagen, reducing the elasticity of skin tissue and causing wrinkles.

[0064] The composition according to the present invention has the activity of inhibiting RAGE expression in skin cells.

[0065] The composition according to the invention can inhibit the increase in RAGE expression levels induced by AGE and AGE precursors in skin cells.

[0066] AGE binds to AGE receptors on the surface of skin cells, producing ROS, inducing inflammatory responses, activating apoptosis signaling pathways, and thus inducing skin cell damage.

[0067] The peptide complex according to the invention inhibits the generation of ROS, the induction of inflammatory responses, or the activation of apoptosis induced by RAGE.

[0068] The peptide complex according to the invention has the activity of increasing the expression of glyoxalase-1 in skin cells.

[0069] The peptide complex according to the invention inhibits the decrease in glyoxalase-1 expression level induced by AGE precursor. Therefore, the peptide complex has the activity of increasing glyoxalase-1 expression.

[0070] Glyoxalase-1 is a protein that has the activity of degrading methylglyoxal, a precursor of AGE, and increasing the expression of glyoxalase-1 can inhibit the production of AGE.

[0071] The compositions according to the present invention can exhibit effects that improve skin condition.

[0072] According to the present invention, the composition for improving skin condition may comprise a peptide complex in a dose of 0.01 μM to 1000 μM. Specifically, the possible values ​​are 0.01 μM to 1000 μM, 0.05 μM to 800 μM, 0.05 μM to 700 μM, 0.05 μM to 600 μM, 0.05 μM to 500 μM, 0.05 μM to 300 μM, 0.05 μM to 200 μM, 0.1 μM to 800 μM, 0.1 μM to 700 μM, 0.1 μM to 600 μM, 0.1 μM to 500 μM, 0.1 μM to 300 μM, 0.1 μM to 200 μM, 1 μM to 800 μM, 1 μM to 700 μM, 1 μM to 600 μM, 1 μM to 500 μM, 1 μM to 300 μM, 1 μM to 200 μM, 5 μM to 800 μM, and 5 μM to 700 μM. Doses of 5 μM to 600 μM, 5 μM to 500 μM, 5 μM to 300 μM, and 5 μM to 200 μM contain peptide complexes according to the invention, but the invention is not limited thereto.

[0073] According to an embodiment, the composition for improving skin condition can be a cosmetic composition.

[0074] Cosmetic compositions can be formulated into any dosage form commonly prepared in the field to which this invention pertains, and can be topical skin preparations. For example, solutions can be formulated into solutions, suspensions, emulsions, pastes, gels, creams, lotions, powders, soaps, surfactant-containing cleansers, oils, powder foundations, emulsion foundations, wax foundations, sprays, etc., but the invention is not limited thereto.

[0075] Cosmetic compositions can be formulated in various forms, such as solutions (e.g., softening lotions, nourishing lotions, nourishing creams, massage creams, serums, eye creams, cleansing creams, cleansing foams, cleansing waters, masks (packs), sprays, loose powders, hair tonics, hair creams, shampoos, hair dyes, conditioners, spray-type hairsprays, aerosol-type hairsprays, hair waxes, gels, etc.), sol-gels, emulsions, oils, waxes, aerosols, etc., but the present invention is not limited thereto.

[0076] The cosmetic compositions according to the present invention may include other additives, such as excipients or carriers. Therefore, conventional ingredients commonly used in general skin cosmetics can be incorporated in appropriate amounts.

[0077] When the dosage form of the cosmetic composition is a paste, cream or gel, animal oil, vegetable oil, wax, paraffin, starch, tragacanth gum, cellulose derivatives, polyethylene glycol, silicone, bentonite, silicon dioxide, talc or zinc oxide may be used as a carrier component.

[0078] 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. Specifically, when the cosmetic composition is in the form of a spray, a propellant such as chlorofluorocarbon, propane / butane, or dimethyl ether may be additionally included, but the invention is not limited thereto.

[0079] When the dosage form of a cosmetic composition is a solution or emulsion, a solvent, solubilizer, or emulsifier can be used as a carrier component. For example, water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butanediol oil, glycerol fatty acid esters, polyethylene glycol, and fatty acid esters of sorbitol can be used.

[0080] When the cosmetic composition is formulated as a suspension, liquid phase diluents such as water, ethanol or propylene glycol, suspending agents such as ethoxylated isostearyl alcohol, polyoxyethyl sorbitol ester and polyoxyethylene sorbitol ester, microcrystalline cellulose, aluminum hydroxide, bentonite, agar or tragacanth gum can be used as carrier components.

[0081] When the formulation of a cosmetic composition is a surfactant-containing cleanser, fatty alcohol sulfates / esters, fatty alcohol ether sulfates / esters, sulfosuccinate monoesters, hydroxyethyl sulfonates, imidazolinium derivatives, methyl taurate / esters, sarcosinates / esters, fatty acid amide ether sulfates / esters, alkylamide betaine, fatty alcohols, fatty acid glycerides, fatty acid diethanolamides, vegetable oils, lanolin derivatives, ethoxylated glycerol fatty acid esters, etc., can be used as carrier components.

[0082] When the cosmetic composition is formulated as a shampoo, basic components used to form the shampoo can be mixed in, such as thickeners, surfactants, viscosity modifiers, humectants, pH adjusters, preservatives, and essential oils. CDE can be used as a thickener; LES (anionic surfactant) and cocobetaine (ampholyte surfactant) can be used as surfactants; polyquaternium salts can be used as viscosity modifiers; glycerin can be used as a humectant; and citric acid and sodium hydroxide can be used as pH adjusters. Grapefruit extract can be used as a preservative. Additionally, essential oils such as cedarwood, peppermint, and rosemary, as well as serine, pentaol, or vitamin E can be added.

[0083] In addition to the peptide complex and carrier component according to the invention, the components included in the cosmetic composition may also contain components commonly used in cosmetic compositions as active ingredients. For example, the components may further include conventional excipients such as antioxidants, stabilizers, solubilizers, vitamins, pigments, and flavorings, but the invention is not limited thereto.

[0084] Pharmaceutical Composition One aspect of the present invention provides a pharmaceutical composition for treating or preventing skin inflammation or skin diseases, comprising a peptide complex as an active ingredient, said peptide complex comprising: (i) a peptide comprising the amino acid sequence of SEQ ID NO.1; and (ii) a peptide comprising the amino acid sequence of SEQ ID NO.2.

[0085] The peptide complexes with the above-described activities according to the present invention can exhibit excellent effects for treating or preventing skin inflammation or skin diseases.

[0086] According to the present invention, a pharmaceutical composition for treating or preventing skin inflammation or skin diseases has at least one of (a) to (e): (a) Inhibit the expression of TNF-α, IL-1β or IL-6, or inhibit the expression of COX-2; (b) Inhibit the expression of cleaved PARP-1, cleaved caspase-3, cleaved caspase-9 or Bax, or increase the expression of Bcl-2. (c) Inhibit the expression of MMP-1 or MMP-2; (d) Inhibit the expression of receptor advanced glycation end products (RAGE); and (e) Increase the expression of glyoxalase-1.

[0087] The pharmaceutical compositions of the present invention have the same activities related to skin inflammation or skin diseases as those described in the compositions for improving skin conditions described above; therefore, their descriptions are incorporated herein by reference and omitted to avoid redundancy.

[0088] Skin inflammation in the pharmaceutical composition can be skin inflammation caused by AGEs.

[0089] The skin disease for which the drug composition is applied can be an AGE-related skin disease.

[0090] According to the present invention, AGE-related skin diseases refer to skin diseases caused directly or indirectly by AGE.

[0091] AGE-related skin diseases can include bacterial skin infections, skin trauma, diabetic skin trauma, diabetic dermatitis, diabetic foot disease, hyperpigmentation disorders, light-induced hyperpigmentation disorders, psoriasis, or systemic lupus erythematosus; and bacterial skin infections can be Staphylococcus aureus skin infections, and hyperpigmentation disorders can be light-induced hyperpigmentation disorders, but the present invention is not limited thereto.

[0092] The descriptions of AGE-related skin diseases are found in Manli Na et al. The Journal of Infectious Diseases, 218(5): 791-800(2018); Hwan June Kang et al. Journal of Controlled Release 333: 176-187(2021); Jiaqi Fang et al. Journal of Investigative Dermatology, 142 (10): 2591-2602 (2022); Anastasia Papagracoraki et al. Int. J Mol. 18, 2471 (2017); Nowak, A et al. Cells, 10,523(2021); Lee et al. Kor. J Herbal. 32(5) 7-12 (2017); Kim et al. Kor. J Herbal. 32(4) 1717-24 (2017), which are incorporated herein by reference.

[0093] As used herein, the term “prevention” refers to reducing the risk of skin inflammation or skin disease, and refers to all actions taken to suppress or delay the onset of disease by preventing the progression of skin inflammation or skin disease or at least one of its clinical symptoms.

[0094] As used herein, the term “treatment” refers to the relief of skin inflammation or skin disease and includes all actions that reduce or slow the progression of at least one clinical symptom of skin inflammation or skin disease to cure or alter the symptoms of the disease.

[0095] The pharmaceutical compositions according to the present invention may contain a therapeutically effective amount of a peptide complex.

[0096] The term "therapeutic effective amount" refers to an amount sufficient to achieve the activity or efficacy of a peptide complex that is the active ingredient of the pharmaceutical composition of the present invention, and for example, an amount sufficient to achieve the efficacy of treating or preventing skin inflammation or skin diseases.

[0097] The pharmaceutical compositions according to the present invention may contain a pharmaceutically acceptable carrier.

[0098] Examples of pharmaceutically acceptable carriers include lactose, dextrose, sucrose, sorbitol, mannitol, starch, gum arabic, calcium phosphate, alginate, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrup, methylcellulose, methylparaben, propylparaben, talc, magnesium stearate, mineral oil, etc., but the present invention is not limited thereto.

[0099] The pharmaceutical compositions of the present invention can be formulated into unit dosage forms using pharmaceutically acceptable carriers and / or excipients according to methods readily practiced by those skilled in the art, or can be formulated by placing them in multi-dose containers. In this case, the formulation may be in the form of a solution, suspension, or emulsion in an oily or aqueous medium, or may be in the form of an extract, powder, granules, tablets, capsules, or gel (e.g., hydrogel), and may additionally include dispersants or stabilizers.

[0100] In addition to the components described above, the pharmaceutical compositions according to the present invention may also contain lubricants, wetting agents, sweeteners, flavoring agents, emulsifiers, suspending agents or preservatives, but the present invention is not limited thereto.

[0101] Suitable pharmaceutically acceptable carriers and formulations are described in detail in Remington's Pharmaceutical Sciences (19th ed; 1995).

[0102] The pharmaceutical composition according to the invention can be administered by all suitable routes for treating skin inflammation or skin diseases, for example, by oral or parenteral administration, and when administered parenterally, by intravenous injection, subcutaneous injection, intramuscular injection, intraperitoneal injection, topical application, transdermal application, etc.

[0103] The dosage of the pharmaceutical composition may be from 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 daily, but the invention is not limited thereto. The dosage of the pharmaceutical composition may be prescribed differently depending on factors such as the preparation method, administration method, patient age, weight, sex, pathological condition, food, administration time, route of administration, excretion rate, and reaction sensitivity.

[0104] The pharmaceutical compositions according to the invention can be formulated into unit dosage forms using pharmaceutically acceptable carriers and / or excipients according to methods readily practiced by those skilled in the art, or can be formulated by placing them in multi-dose containers. In this case, the formulation can be in the form of a solution, suspension, or emulsion in an oil or aqueous medium, or in the form of an X-dosage agent, powder, granules, tablets, capsules, or gel (e.g., hydrogel), and may additionally include dispersants or stabilizers.

[0105] The pharmaceutical compositions according to the invention can be used alone or in combination with surgery, radiotherapy, hormone therapy, chemotherapy and biological response modifiers to treat or prevent skin inflammation or skin diseases.

[0106] The pharmaceutical compositions according to the present invention can be topical skin preparations. Topical skin preparations are preparations that can be applied externally to the skin. When the pharmaceutical compositions according to the present invention are used as topical skin preparations, the pharmaceutical compositions can be applied to the skin, specifically to skin sites where skin inflammation or skin diseases occur. Topical skin preparations can be creams, gels, ointments, skin emulsifiers, skin suspensions, transdermal patches, medicated bandages, lotions, or combinations thereof. Topical skin preparations can include topical skin preparations used in cosmetics or pharmaceuticals. For example, topical skin preparations can contain aqueous components, oily components, powdered components, alcohols, moisturizers, medium-viscosity agents, ultraviolet absorbers, whitening agents, preservatives, antioxidants, surfactants, fragrances, colorants, various skin nutrients, or combinations thereof, as well as substances formulated by appropriately mixing the above substances. Topical skin preparations can be formulated by appropriately mixing metal chelating agents such as disodium EDTA, trisodium EDTA, sodium citrate, sodium polyphosphate, sodium methanate, or gluconate; hot water extracts such as caffeine, tannins, bellafamil, licorice extract, glabridine, and caline fruits; various herbs; tocopheryl acetate, glycidylic acid, tranexamic acid, tranexamic acid derivatives, or drugs such as tranexamic acid salts; saccharides such as vitamin C, magnesium ascorbate phosphate, ascorbate glucoside, arbutin, or kojic acid; and sugars such as glucose, fructose, and trehalose.

[0107] Food composition Another aspect of this disclosure provides a functional food composition for improving skin condition, comprising a peptide complex as an active ingredient, said peptide complex comprising: (i) a peptide comprising the amino acid sequence of SEQ ID NO.1; and (ii) a peptide comprising the amino acid sequence of SEQ ID NO.2.

[0108] Improved skin condition can refer to relieving skin inflammation, improving skin aging, promoting skin regeneration, improving skin elasticity, reducing skin wrinkles, improving the skin barrier, or improving skin damage.

[0109] The skin-condition-improving activities in the functional food compositions of the present invention have been described in the above-described compositions for improving skin condition. Therefore, the details thereof are incorporated herein by reference and omitted to avoid redundancy.

[0110] In the functional food composition of the present invention, based on the total weight of the composition, an appropriate amount of peptide complex selected in the range of 0.000001 wt% to 10 wt% may be included.

[0111] The functional food composition according to the present invention may contain a food-effective amount of peptide complex and a food-acceptable carrier.

[0112] The functional food compositions according to the present invention include not only peptide complexes as active ingredients, but also components commonly added in food preparation, and may include, for example, proteins, carbohydrates, fats, nutrients, seasonings, and flavorings. The carbohydrates include: monosaccharides, such as glucose and fructose; disaccharides, such as maltose, sucrose, and oligosaccharides; polysaccharides, including typical sugars such as dextrin and cyclodextrin; and sugar alcohols, such as xylitol, sorbitol, and erythritol. Flavorings may include natural flavorings, sematriol and stevia extracts (e.g., rebaudioside A, glycyrrhizin), and synthetic flavorings (e.g., saccharin and aspartame).

[0113] In addition to the above-mentioned ingredients, the functional food compositions according to the present invention may also contain various nutrients, vitamins, minerals (electrolytes), flavoring agents such as synthetic and natural flavorings, coloring agents and enhancers (for cheese or chocolate), pectic acid and its salts, alginic acid and its salts, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohols, and carbonating agents used in carbonated beverages. Furthermore, the functional food compositions may include fruit pulp used in the preparation of natural fruit juices and fruit juice beverages and vegetable beverages. For example, when the functional food compositions according to the present invention are prepared as beverage formulations, in addition to the peptides of the present invention as active ingredients, the functional food compositions may also contain citric acid, liquid fructose, sugar, glucose, acetic acid, malic acid, fruit juice, Eucommia extract, jujube extract, and licorice extract.

[0114] Functional foods can be offered in various types without particular restrictions. Functional foods and foods may include, for example, meats, sausages, bread, chocolate, candy, snacks, biscuits, pizza, ramen, other noodles, chewing gum, ice cream, dairy products, various soups, beverages, tea drinks, alcoholic beverages, vitamin complexes, dairy products and fermented milk, and include all typical functional foods or foods.

[0115] Use of the peptide complex according to the present invention Another aspect of the present invention provides the use of peptide complexes for improving skin condition, treating, preventing or alleviating skin inflammation or skin diseases, said peptide complexes comprising: (i) a peptide comprising the amino acid sequence of SEQ ID NO.1; and (ii) a peptide comprising the amino acid sequence of SEQ ID NO.2.

[0116] Another aspect of the present invention provides the use of peptide complexes in the preparation of medicaments, cosmetics or health functional foods for improving skin condition, treating, preventing or alleviating skin inflammation or skin diseases, said peptide complexes comprising: (i) a peptide comprising the amino acid sequence of SEQ ID NO.1; and (ii) a peptide comprising the amino acid sequence of SEQ ID NO.2.

[0117] Another aspect of the present invention provides a method for improving skin condition, or for treating, preventing or alleviating skin inflammation or skin disease, the method comprising: administering a peptide complex to a subject who requires improved skin condition or a subject who requires treatment, prevention or alleviation of skin inflammation or skin disease, said peptide complex comprising: (i) a peptide comprising the amino acid sequence of SEQ ID NO. 1; and (ii) a peptide comprising the amino acid sequence of SEQ ID NO. 2.

[0118] Subjects may be humans or animals other than humans. Furthermore, subjects may be individuals or animals suffering from skin inflammation or skin diseases.

[0119] The descriptions of the peptide complexes, compositions, cosmetic compositions, pharmaceutical compositions, and functional food compositions according to the present invention are equally applicable to methods for improving skin conditions and for treating, preventing, or alleviating skin inflammation or skin diseases, and repetitions thereof will be omitted to avoid excessive complexity in this specification.

[0120] Beneficial effects The peptide complex according to the invention has inhibitory activity against skin cell damage. The peptide complex according to the invention inhibits various skin damage effects induced by AGE or AGE precursors in skin cells, such as activation of inflammatory responses, promotion of apoptosis, increased expression of MMP-1 and MMP-2, increased expression of RAGE, and decreased expression of glyoxalase, thereby improving damage to skin cells. The effects of the invention are not limited to the above-described effects, and any other effects not mentioned herein will be clearly understood by those skilled in the art from the following description. Attached Figure Description

[0121] Figure 1A shows the results of treatment with peptide complexes leading to a decrease in the expression levels of TNF-α, IL-1β, and IL-6, which are inflammatory response factors, in HaCaT keratinocytes after AGE-BSA treatment. Figure 1B is a graph obtained by quantifying the intensity of the bands in Figure 1A.

[0122] Figure 2A shows the result of reduced expression levels of TNF-α, IL-1β, and IL-6, which are inflammatory response factors, in HaCaT keratinocytes after treatment with the peptide complex via MGO treatment, and Figure 2B is a graph obtained by quantifying the intensity of the bands in Figure 2A.

[0123] Figure 3A shows the results of treatment with peptide complexes leading to a decrease in the expression levels of COX-2, IL-1β, and IL-6, which are inflammatory response factors, in NIH3T3 fibroblasts after AGE-BSA treatment. Figure 3B is a graph obtained by quantifying the intensity of the bands in Figure 3A.

[0124] Figure 4A shows the results of treatment with peptide complexes leading to increased expression levels of cleaved PARP-1, cleaved caspase-3, cleaved caspase-9, and Bax, an apoptosis-mediating protein, in HaCaT keratinocytes after AGE-BSA treatment, and decreased expression levels of these substances. Figure 4B is a graph obtained by quantifying the intensity of the bands in Figure 4A.

[0125] Figure 5A shows the results of MGO treatment leading to decreased expression levels of cleaved caspase-3 and cleaved caspase-9, apoptosis regulators, in HaCaT keratinocytes, as indicated by peptide complex treatment, and increased expression levels of Bcl-2, an anti-apoptotic protein, as indicated by MGO treatment, as indicated by peptide complex treatment. Figure 5B is a graph obtained by quantifying the intensity of the bands in Figure 5A.

[0126] Figure 6A shows the results of treatment with the peptide complex in NIH3T3 fibroblasts with AGE-BSA, which increased the expression levels of cleaved PARP-1, cleaved caspase-3, and cleaved caspase-9 (actually apoptosis regulators) and decreased the expression levels of Bax (an apoptosis-mediating protein). Figure 6B shows the results of treatment with the peptide complex in NIH3T3 fibroblasts with AGE-BSA, which increased the expression levels of Bcl-2 (an anti-apoptotic protein) and decreased the expression levels of Bcl-2 (an anti-apoptotic protein).

[0127] Figure 7A shows the result of reduced expression levels of MMP-1 and MMP-2 in HaCaT keratinocytes after treatment with the peptide complex, which was increased by AGE-BSA treatment. Figure 7B is a graph obtained by quantifying the intensity of the bands in Figure 7A.

[0128] Figure 8A shows the result of reduced expression levels of MMP-1 and MMP-2 in HaCaT keratinocytes after treatment with the peptide complex, which was increased by MGO treatment. Figure 8B is a graph obtained by quantifying the intensity of the bands in Figure 8A.

[0129] Figure 9A shows the result of reduced MMP-1 expression levels in NIH3T3 fibroblasts after treatment with AGE-BSA via peptide complex treatment, and Figure 9B is a graph obtained by quantifying the intensity of the bands in Figure 9A.

[0130] Figure 10A shows the result of reduced RAGE expression levels in HaCaT keratinocytes after treatment with the peptide complex, which was increased by AGE-BSA treatment, and Figure 10B is a graph obtained by quantifying the intensity of the bands in Figure 10A.

[0131] Figure 11A shows the result of reduced RAGE expression levels in HaCaT keratinocytes after treatment with the peptide complex, which was increased by MGO treatment, and Figure 11B is a graph obtained by quantifying the intensity of the bands in Figure 11A.

[0132] Figure 12A shows the result of reduced RAGE expression levels in NIH3T3 fibroblasts after treatment with the peptide complex via AGE-BSA, and Figure 12B is a graph obtained by quantifying the intensity of the bands in Figure 12A.

[0133] Figure 13A shows the result of increased glyoxalase-1 expression levels in HaCaT keratinocytes after treatment with the peptide complex, which was reduced by MGO treatment, and Figure 13B is a graph obtained by quantifying the intensity of the bands in Figure 13A. Detailed Implementation

[0134] The present invention will now be described in detail. However, the following embodiments are provided only to illustrate the invention in more detail, and the invention is not limited to the following description.

[0135] Preparation Example 1: Preparation of Peptide Complexes Peptides having the amino acid sequences of SEQ ID NO. 1 and SEQ ID NO. 2, as shown in Table 1, were synthesized using an automated peptide synthesizer (Milligen 9050; Millipore; USA). The synthesized peptides were purified and separated using C18 reversed-phase high-performance liquid chromatography (HPLC) (Waters Associates, USA). The column used was an ACQUITY UPLC BEH300 C18 (2.1 mm × 100 mm; 1.7 μM; Waters Co., USA).

[0136] [Table 1]

[0137] Then, peptide complexes were prepared by mixing equal amounts of the peptide with the amino acid sequence of SEQ ID NO. 1 and the peptide with the amino acid sequence of SEQ ID NO. 2, and the effects of the prepared peptide complexes were evaluated in the following experimental examples.

[0138] Experimental Example 1: Measurement of the expression levels of inflammatory cytokines in skin cells The effect of the peptide complex prepared in Example 1 on the expression levels of inflammatory cytokines induced by AGE or its precursor methylglyoxal (MGO) in skin cells was evaluated. HaCaT human keratinocytes and NIH3T3 fibroblasts were cultured at 2 × 10⁶ cells per well. 5Cells were seeded at a density of 100 μM in 6-well plates and cultured for 24 hours. Then, cells were pretreated for 1 hour with the peptide complex prepared in Preparation Example 1 at concentrations of 50 μM, 100 μM, and 200 μM. The pretreated cells were then treated with 200 μg / mL AGE-BSA or 200 μM methylglyoxal (MGO) and cultured in a CO2 incubator at 37°C for 48 hours. In the AGE-BSA treatment experiment, the control wells were treated with 200 μg / mL BSA. After treatment, cells were washed twice with PBS and treated with easy-BLUE. TM Total RNA was isolated using an iNtRON kit (catalog number: 17061, Korea). The isolated total RNA was quantified, and cDNA was synthesized using an RT kit (enzynomics, catalog number: RT200, Korea). PCR was then performed using a PCR kit (enzynomics, catalog number: P581T, Korea). The primers used are shown in Table 2 below.

[0139] [Table 2]

[0140] The mRNA expression levels of inflammatory cytokines TNF-α, IL-1β, and IL-6, which are markers of AGE-BSA-induced inflammatory responses, were measured in HaCaT keratinocytes. The results are shown in Figures 1A and 1B. As shown in Figures 1A and 1B, compared with the control group, treatment with AGE-BSA increased the mRNA expression levels of TNF-α, IL-1β, and IL-6 in the negative control group. Treatment with the peptide complex of the present invention decreased the mRNA expression levels of TNF-α, IL-1β, and IL-6 compared with the negative control group. Furthermore, the mRNA expression levels of inflammatory cytokines TNF-α, IL-1β, and IL-6, which are markers of methylglyoxal (MGO)-induced inflammatory responses, were measured in HaCaT keratinocytes. The results are shown in Figures 2A and 2B. As shown in Figures 2A and 2B, compared with the control group, treatment with MGO increased the mRNA expression levels of TNF-α, IL-1β, and IL-6 in the negative control group. Compared with the negative control group, treatment with the peptide complex of the present invention reduced the mRNA expression levels of TNF-α, IL-1β and IL-6.

[0141] The mRNA expression levels of COX-2 (an inflammatory mediator), IL-1β, and IL-6 (an inflammatory cytokine), markers of AGE-BSA-induced inflammation, were measured in CENIH3T3 fibroblasts. The results are shown in Figures 3A and 3B. As shown in Figures 3A and 3B, the mRNA expression levels of COX-2 and IL-1β were significantly increased in the negative control group treated with AGE-BSA compared to the control group. When treated with the peptide complex according to the invention, the mRNA expression levels of COX-2 and IL-6 were decreased compared to the negative control group. For IL-1β, the mRNA expression level was slightly higher when treated with a 50 μM dose of the peptide complex compared to the negative control group. However, the mRNA expression level decreased when treated with 100 μM and 200 μM doses of the peptide complex.

[0142] Experimental Example 2: Measurement of the expression levels of apoptosis regulators MMP and RAGE in skin cells 1. Experimental Methods To evaluate the skin damage improvement effect of the peptide complex prepared in Preparation Example 1, the expression levels of apoptosis regulators, matrix metalloproteinases (MMPs), and receptor for advanced glycation end products (RAGE) in skin cells were detected by Western blotting.

[0143] Specifically, HaCaT human skin keratinocytes and NIH3T3 fibroblasts were mixed at a ratio of 2 × 10⁶ cells per well. 5Cells were seeded at a density of 100 μM in 6-well plates and cultured for 24 hours. Then, cells were pretreated for 1 hour with the peptide complex prepared in Preparation Example 1 at concentrations of 50 μM, 100 μM, and 200 μM. The treated cells were then treated with 200 μg / ml AGE-BSA or 200 μM methylglyoxal (MGO) and cultured in a CO2 incubator at 37°C for 48 hours. In the AGE-BSA treatment experiments, control wells were treated with 200 μg / ml BSA. After treatment, cells were washed twice with PBS and lysed by adding cell lysis buffer. After treatment with 5X sample buffer, SDS-PAGE was performed using a 10% SDS-PAGE gel, and the proteins separated by SDS-PAGE were transferred to a PVDF membrane. The membrane was blocked with 5% skim milk at room temperature for 1 hour. Each primary antibody listed in Table 3 was diluted 1:1000 in 5% skim milk and incubated with the membrane for 2 hours, followed by washing three times for 10 minutes with 0.1% PBS-T (0.1% Tween-20 in PBS). Then, each secondary antibody corresponding to the primary antibody listed in Table 3 was diluted 1:3000 in 5% skim milk and incubated with the membrane for 2 hours. After treatment with ECL solution (GE Healthcare, catalog number: RPN2232, USA), images were acquired using an ImageQuant 800 system (Cytiva, catalog number: 29-3994-81, USA).

[0144] [Table 3]

[0145] 2. Observation of the effect of inhibiting skin cell apoptosis To evaluate the inhibitory effect of the peptide complex on skin cell apoptosis, the expression levels of apoptosis regulators were measured. The results are shown in Figures 4A, 4B, 5A, 5B, 6A, and 6B. Cleaved PARP-1, cleaved caspase-3, and cleaved caspase-9 are characteristic marker proteins of apoptosis, Bax is a pro-apoptotic mediator, and Bcl-2 is an anti-apoptotic protein.

[0146] First, in HaCaT keratinocytes, the expression levels of cleaved PARP-1, cleaved caspase-3, cleaved caspase-9, and Bax, which are apoptosis regulators, induced by AGE-BSA treatment were measured, and the results are shown in Figures 4A and 4B. As shown in the negative control groups of Figures 4A and 4B, the expression levels of cleaved caspase-3, cleaved caspase-9, and Bax increased after AGE-BSA treatment, and the increased expression levels of these apoptosis regulators decreased after treatment with a peptide complex.

[0147] In addition, the expression levels of cleaved caspase-3, cleaved caspase-9, and Bcl-2, which are apoptosis regulators, were measured in HaCaT keratinocytes after treatment with MGO. The results are shown in Figures 5A and 5B. As shown in the negative control groups of Figures 5A and 5B, MGO treatment increased the expression level of cleaved caspase-3, did not change the expression level of cleaved caspase-9, and decreased the expression level of the anti-apoptotic factor Bcl-2. However, as shown in the experimental groups, treatment with the peptide complex decreased the expression levels of cleaved caspase-3 and cleaved caspase-9. Furthermore, the expression level of Bcl-2, which was decreased by MGO treatment, was increased by treatment with the peptide complex.

[0148] In NIH3T3 fibroblasts, the expression levels of cleaved PARP-1, cleaved caspase-3, cleaved caspase-9, Bax, and Bcl-2, which are apoptosis regulators, were measured after AGE-BSA treatment. The results are shown in Figures 6A and 6B. As shown in the negative control groups of Figures 6A and 6B, AGE-BSA treatment increased the expression levels of cleaved PARP-1, cleaved caspase-3, cleaved caspase-9, and Bax, while decreasing the expression level of the anti-apoptotic factor Bcl-2. However, as shown in the experimental groups, the increased expression levels of cleaved PARP-1, cleaved caspase-3, and Bax after AGE-BSA treatment were reduced by treatment with the peptide complex. For Bcl-2, the decreased expression level in the negative control group was increased in a dose-dependent manner by treatment with the peptide complex.

[0149] Therefore, these results confirm that the peptide complex of the present invention inhibits the expression of pro-apoptotic factors induced by AGE-BSA or MGO, promotes the expression of anti-apoptotic factors in HaCaT keratinocytes and NIH3T3 fibroblasts, thereby inhibiting cell apoptosis and improving skin cell damage.

[0150] 3. Observation of the effect of inhibiting MMP expression Matrix metalloproteinases (MMPs) are proteolytic enzymes that specifically target collagen. MMP-1 and MMP-2 degrade collagen, leading to decreased skin elasticity and wrinkle formation. Therefore, this study investigated whether treatment of skin cells with the peptide complex of the present invention reduced MMP expression levels, and the results are shown in Figures 7A, 7B, 8A, 8B, 9A, and 9B.

[0151] First, in HaCaT keratinocytes, the expression levels of MMP-1 and MMP-2 induced by AGE-BSA treatment and the changes in MMP-1 and MMP-2 expression levels after treatment with a peptide complex were investigated, and the results are shown in Figures 7A and 7B. As shown in Figures 7A and 7B, in the negative control group, AGE-BSA treatment increased the expression levels of MMP-1 and MMP-2. However, as shown in the experimental group, treatment with the peptide complex decreased the protein expression levels of MMP-1 and MMP-2.

[0152] Furthermore, in HaCaT keratinocytes, the increased expression levels of MMP-1 and MMP-2 after MGO treatment and the changes in MMP-1 and MMP-2 expression levels after peptide complex treatment were investigated, and the results are shown in Figures 8A and 8B. As shown in Figures 8A and 8B, in the negative control group, MGO treatment increased the expression levels of MMP-1 and MMP-2. However, as shown in the experimental group, compared with the negative control group, peptide complex treatment decreased the protein expression levels of MMP-1 and MMP-2.

[0153] In NIH3T3 fibroblasts, the increased expression level of MMP-1 after AGE-BSA treatment and the changes in MMP-1 expression level after treatment with a peptide complex were investigated, and the results are shown in Figures 9A and 9B. As shown in Figures 9A and 9B, in the negative control group, AGE-BSA treatment increased the expression level of MMP-1. However, as shown in the experimental group, treatment with the peptide complex decreased the protein expression level of MMP-1 compared with the negative control group.

[0154] Therefore, these results confirm that the peptide complex of the present invention inhibits the expression of MMP-1 and MMP-2, which are increased by AGE-BSA or MGO treatment in HaCaT keratinocytes and NIH3T3 fibroblasts. Through this inhibition, the peptide complex reduces collagen degradation, enhances the elasticity of skin tissue, prevents wrinkle formation, and thus improves skin damage.

[0155] 4. Observation of the effect of inhibiting RAGE expression The receptor for advanced glycation end products (RAGE) is a transmembrane receptor protein located on the cell surface that binds to advanced glycation end products (AGEs). When AGEs bind to RAGEs in skin cells such as keratinocytes and fibroblasts, it is known that they can induce skin cell damage through the generation of reactive oxygen species (ROS), the induction of inflammatory responses, and the activation of apoptosis signaling pathways. Therefore, this study investigated whether treatment of skin cells with the peptide complex of the present invention reduced the expression level of RAGEs. The results are shown in Figures 10A, 10B, 11A, 11B, 12A, and 12B.

[0156] First, in HaCaT keratinocytes, the changes in RAGE expression levels induced by AGE-BSA treatment and after treatment with a peptide complex were investigated. As shown in Figures 10A and 10B, in the negative control group, RAGE expression levels increased after AGE-BSA treatment. However, as in the experimental group, treatment with the peptide complex decreased RAGE protein expression levels in a dose-dependent manner.

[0157] Furthermore, in HaCaT keratinocytes, the changes in RAGE expression levels induced by MGO treatment and after treatment with the peptide complex were investigated. As shown in Figures 11A and 11B, in the negative control group, RAGE expression levels increased with MGO treatment. However, as shown in the experimental group, treatment with the peptide complex decreased RAGE protein expression levels compared to the negative control group at peptide complex concentrations of 100 μM or higher.

[0158] In NIH3T3 fibroblasts, the changes in RAGE expression levels induced by AGE-BSA treatment and after treatment with the peptide complex were examined. As shown in Figures 12A and 12B, in the negative control group, RAGE expression levels increased after AGE-BSA treatment. However, as shown in the experimental group, RAGE expression levels decreased after treatment with the peptide complex compared to the negative control group.

[0159] Therefore, these results demonstrate that treatment with the peptide complex of the present invention reduces the increased RAGE expression levels induced by AGE-BSA or MGO in HaCaT keratinocytes and NIH3T3 fibroblasts. Through this reduction, the peptide complex decreases AGE binding, inhibits ROS production, inflammatory responses, and activation of apoptosis signaling pathways, thereby improving skin damage.

[0160] Example 3: Evaluation of glyoxalase-1 expression level for inhibiting the formation of advanced glycation end products (AGEs) Reactive dicarbonyl compounds such as glyoxal, methylglyoxal, and 3-deoxyglucosone can cross-link with proteins to produce advanced glycation end products (AGEs). Glyoxalase-1 is a detoxification enzyme that converts methylglyoxal to lactate, thereby eliminating methylglyoxal.

[0161] The effect of the peptide complex prepared in Example 1 on the expression level of glyoxalase-1 in HaCaT keratinocytes was evaluated by RT-PCR. First, human HaCaT keratinocytes were cultured in wells at a density of 2 × 10⁶ cells / well. 5Cells were seeded at a density of 100 μM in 6-well plates and cultured for 24 hours. Then, the cells were pretreated for 1 hour with the peptide complex prepared in Preparation Example 1 at concentrations of 50 μM, 100 μM, and 200 μM. The pretreated cells were then treated with 200 μM methylglyoxal (MGO) and cultured in a CO2 incubator at 37°C for 24 hours. After treatment, the cells were washed twice with PBS and then treated with easy-BLUE. TM Total RNA was isolated using (iNtRON, catalog number: 17061, Korea). The isolated total RNA was quantified, and cDNA was synthesized using an RT kit (enzynomics, catalog number: RT200, Korea). PCR was then performed using a PCR kit (enzynomics, catalog number: P581T, Korea). The primers used are shown in Table 4 below.

[0162] [Table 4]

[0163] As shown in Figures 13A and 13B, in the negative control group, MGO treatment reduced the expression level of glyoxalase-1. However, as in the experimental group, treatment with the peptide complex increased the expression level of glyoxalase-1. Therefore, the peptide complex increased the expression of glyoxalase-1, which was reduced by MGO treatment, thereby enabling the degradation of methylglyoxal, a precursor of AGE. Through this mechanism, AGE formation was inhibited, thus improving skin lesions.

[0164] Therefore, these results indicate that treatment of skin cells with the peptide complex of the present invention reduces the expression of AGE-induced inflammatory factors, thereby exhibiting an anti-inflammatory effect, and reduces the expression of apoptosis-related factors, thereby inhibiting apoptosis. Furthermore, the expression of collagen-degrading enzymes is reduced, thereby enhancing the elasticity of skin tissue and preventing wrinkle formation. In addition, treatment with the peptide complex reduces the expression of RAGE, which is the receptor for AGE, thereby reducing AGE binding and inhibiting ROS generation, inflammatory responses, and activation of apoptosis signaling pathways, thereby improving skin damage. Furthermore, the peptide complex increases the expression level of glyoxalase-1, which is reduced by MGO treatment, thereby promoting the degradation of methylglyoxal, the precursor of AGE, and inhibiting AGE formation, thereby improving skin damage.

[0165] As described above, embodiments of the present invention have been provided for illustrative purposes, but the scope of the invention is not limited to the specific embodiments, specific preparation embodiments, and specific experimental examples. In other words, the present invention can be appropriately modified by those skilled in the art within the scope of the appended claims.

Claims

1. A composition for improving skin condition comprising a peptide complex as an active ingredient, said peptide complex comprising: (i) a peptide comprising the amino acid sequence of SEQ ID NO.1; and (ii) a peptide comprising the amino acid sequence of SEQ ID NO.

2.

2. The composition according to claim 1, wherein, The improvement in skin condition is selected from at least one of the following groups: reducing skin inflammation, improving skin aging, promoting skin regeneration, improving skin elasticity, reducing skin wrinkles, improving the skin barrier, and improving skin damage.

3. The composition according to claim 1, wherein, The composition exhibits at least one activity selected from (a) to (e) below, (a) Inhibit the expression of TNF-α, IL-1β or IL-6, or inhibit the expression of COX-2; (b) Inhibit the expression of cleaved PARP-1, cleaved caspase-3, cleaved caspase-9 or Bax, or increase the expression of Bcl-2. (c) Inhibit the expression of MMP-1 or MMP-2; (d) Inhibit the expression of receptor advanced glycation end products (RAGE); and (e) Increase the expression of glyoxalase-1.

4. The composition according to claim 1, wherein, The composition is a cosmetic composition.

5. The composition according to claim 4, wherein, The cosmetic composition comprises any one of the following formulations selected from the group consisting of solutions, suspensions, emulsions, pastes, gels, creams, lotions, powders, soaps, surfactant-containing detergents, oils, powder foundations, emulsion foundations, wax foundations, and sprays.

6. A pharmaceutical composition for treating or preventing skin inflammation or skin diseases, comprising a therapeutically effective amount of a peptide complex, said peptide complex comprising: (i) a peptide comprising the amino acid sequence of SEQ ID NO.1; and (ii) a peptide comprising the amino acid sequence of SEQ ID NO.

2.

7. The composition according to claim 6, wherein, The skin diseases mentioned are selected from bacterial skin infections, skin trauma, diabetic skin trauma, diabetic dermatitis, diabetic foot disease, hyperpigmentation, photoinduced hyperpigmentation, psoriasis, and systemic lupus erythematosus.

8. A functional food composition for improving skin condition, comprising a peptide complex as an active ingredient, said peptide complex comprising: (i) a peptide comprising the amino acid sequence of SEQ ID NO.1; and (ii) a peptide comprising the amino acid sequence of SEQ ID NO.

2.

9. The composition according to claim 8, wherein, The improvement in skin condition is selected from at least one of the following groups: reducing skin inflammation, improving skin aging, promoting skin regeneration, improving skin elasticity, reducing skin wrinkles, improving the skin barrier, and improving skin damage.

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