Peptides having anti-obesity activity and their uses

Abstract

The present invention relates to a peptide having anti-obesity activity by reducing accumulated fat, and a composition for preventing, improving, or treating obesity, comprising the same. The peptide comprising the amino acid sequence of SEQ ID NO: 1 according to the present invention has activity of reducing intracellular accumulated fat, and therefore can be used as a raw material for pharmaceuticals for preventing, improving, or treating obesity or obesity-related diseases, or cosmetics for body slimming.

Description

Technical Field

[0001] The present invention relates to a peptide having anti-obesity activity and a composition for preventing, improving, or treating obesity containing the peptide as an active ingredient.

Background Art

[0002] Obesity means a state in which adipose tissue is excessively present in the body due to the accumulation of excess energy as body fat when the energy consumption from diet is not balanced. According to the World Health Organization (WHO), more than 1 billion adults worldwide are overweight, and at least 3 million of them are clinically obese, and this is increasing significantly in the United States and Europe. Overweight and obesity increase blood pressure and cholesterol levels, causing various diseases such as heart disease, diabetes, and arthritis, and increasing the incidence of various adult diseases. In addition, overweight and obesity are one of the factors increasing the incidence of various adult diseases such as arteriosclerosis, hypertension, hyperlipidemia, or heart disease not only in adults but also in children and adolescents.

[0003] Currently, representative obesity treatment drugs approved by the US FDA and widely prescribed are drugs that act on the central nervous system and act as appetite suppressants, and orlistat (Xenical), which is an inhibitor of the digestive enzyme lipase secreted from the pancreas. Many drugs such as sibutramine have had their approvals revoked due to cardiovascular and psychiatric side effects of drugs acting on the central nervous system, and orlistat has limitations such as various side effects and variable drug effects depending on the amount of fat intake. On the other hand, as an endocrine peptide target drug, liraglutide, which is a glucagon-like peptide-1 (GLP-1) receptor agonist, has been approved and used (Non-Patent Document 1), but the risk of thyroid cancer has emerged.

[0004] Therefore, there is a demand for the development of a next-generation anti-obesity agent that can overcome the limitations of drugs acting on the central nervous system and gastrointestinal tract. Recently, the development direction of anti-obesity agents has a tendency that the importance is expanding from drugs acting on the central nervous system and gastrointestinal tract to locally acting drugs, and from orally administered low-molecular synthetic compounds to synthetic peptides for subcutaneous or intravenous injection. In addition, new drug development applying new biotechnologies such as therapeutic antibodies, RNAi, and aptamer is also being explored. The development of synthetic peptide therapeutic agents is a promising driving force for the future biopharmaceutical industry, and since it also has great potential for clinical efficacy in the field of anti-obesity agent development, it is a very important situation to discover new drug targets and mechanisms, develop peptide-based drugs, and develop transmission optimization technologies.

Prior Art Documents

Non-Patent Documents

[0005]

Non-Patent Document 1

Summary of the Invention

Problems to be Solved by the Invention

[0006] One object of the present invention is to provide a peptide having anti-obesity activity by promoting lipolysis.

[0007] Another object of the present invention is to provide a pharmaceutical composition for preventing or treating obesity containing the peptide having the above-described activity as an active ingredient.

[0008] Still another object of the present invention is to provide a composition for promoting lipolysis containing the peptide having the above-described activity as an active ingredient.

[0009] Another object of the present invention is to provide a cosmetic composition for preventing or improving obesity, which contains a peptide having the above-described activity as an active ingredient.

Means for Solving the Problems

[0010] In order to achieve the above object, one aspect of the present invention provides a peptide containing the amino acid sequence of SEQ ID NO: 1.

[0011] Another aspect of the present invention provides a pharmaceutical composition for preventing or treating obesity, which contains the peptide as an active ingredient.

[0012] Still another aspect of the present invention provides a composition for promoting lipolysis, which contains the peptide as an active ingredient.

[0013] Still another aspect of the present invention provides a cosmetic composition for preventing or improving obesity, which contains the peptide as an active ingredient.

Effects of the Invention

[0014] The peptide containing the amino acid sequence of SEQ ID NO: 1 according to the present invention has an activity of reducing intracellularly accumulated fat, and thus can be used as a raw material for pharmaceuticals for the purpose of preventing, improving, or treating obesity or obesity-related diseases, or cosmetics for the purpose of body slimming.

[0015] The effects of the present invention are not limited to the effects mentioned above, and other effects not mentioned can be clearly understood by those skilled in the art from the following description.

Brief Description of the Drawings

[0016]

Fig. 1a

Fig. 1b

Fig. 1c

Fig. 1d

Fig. 2a

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Fig. 3

Fig. 4a

Fig. 4b

Fig. 4c

Fig. 4d

Fig. 4e

Fig. 5a

Fig. 5b

Fig. 5c

[0017] Hereinafter, the present invention will be described in detail.

[0018] One aspect of the present invention provides a peptide comprising the amino acid sequence represented by SEQ ID NO: 1.

[0019] As used herein, the term "peptide" means a linear molecule composed of amino acid residues. Specifically, the peptide of the present invention may contain the amino acid sequence represented by SEQ ID NO: 1.

[0020] The "peptide" may be a variant or fragment of an amino acid having a different sequence due to deletion, insertion, substitution, or a combination thereof of amino acid residues within a range that does not affect the function. Amino acid exchanges that do not globally alter the activity of the peptide are known in the art. In some cases, the peptide may be modified by phosphorylation, sulfation, acrylation, glycosylation, methylation, farnesylation, etc. Therefore, the peptide of the present invention includes a peptide having an amino acid sequence substantially identical to the peptide containing the amino acid sequence of SEQ ID NO: 1, its variants, or its active fragments.

[0021] The "substantially identical protein" means an amino acid sequence having a sequence homology of 75% or more, preferably 80% or more, for example, 85% or more, 90% or more, 95% or more, 98% or more, or 99% or more with the amino acid sequence of SEQ ID NO: 1, but is not limited thereto. If it has a homology of 75% or more of the amino acid sequence and has the same activity, it is included in the scope of the present invention.

[0022] In addition, the peptide of the present invention may further include a targeting sequence, a tag, a labeled residue, a half-life, or an amino acid sequence produced for a specific purpose of increasing peptide stability.

[0023] In addition, in order to obtain better chemical stability, enhanced pharmacological properties (half-life, water absorption, titer, efficacy, etc.), altered specificity (for example, a wide range of biological activity spectra), and reduced antigenicity, a protecting group may be attached to the N-terminus or C-terminus of the peptide of the present invention. The "stability" is used to mean not only the in vivo stability that protects the peptide of the present invention from the attack of proteolytic enzymes in the living body, but also storage stability (for example, room temperature storage stability).

[0024] The N-terminal modification may be one in which a protecting group selected from the group consisting of an acetyl group, a fluoreonylmethoxycarbonyl group, a formyl group, a palmitoyl group, a myristyl group, a stearyl group, and polyethylene glycol (PEG) is attached to the N-terminus of the peptide, but is not limited thereto.

[0025] The C-terminal modification is a hydroxyl group (-OH), an amino group (-NH 2 ) at the C-terminus of the peptide, an azide (-NHNH 2) may be a combination of such, but is not limited thereto.

[0026] The synthesis of the peptide of the present invention may be carried out, for example, using equipment or genetic engineering techniques. When synthesizing using equipment, the desired peptide may be synthesized by the Fmoc solid-phase method with an automatic peptide synthesizer.

[0027] In a specific embodiment of the present invention, the peptide containing the amino acid sequence of SEQ ID NO: 1 of the present invention was synthesized and identified using the Fmoc solid-phase method, and the efficacy of the identified peptide was verified and selected.

[0028] In a specific embodiment of the present invention, in order to verify the efficacy of the identified peptide, a peptide having anti-obesity activity was selected through verification of effectiveness against lipolytic effect.

[0029] In a specific embodiment of the present invention, when the peptide of the present invention was treated with adipocytes, it was confirmed that the accumulated fat was decomposed and the glycerol release amount was increased.

[0030] In a specific embodiment of the present invention, when the peptide of the present invention was treated with adipocytes, it was confirmed that the expression of Perilipin, HSL (Hormone sensitive lipase), or ATGL (Adipose triglyceride lipase), which are lipolytic factors, was increased.

[0031] In a specific embodiment of the present invention, when the peptide of the present invention is treated on adipocytes, it was confirmed that the expression of PGC1α (peroxisome proliferator-activated receptor gamma coactivator 1-alpha), TBx1 (T-box transcription factor), FGF21 (Fibroblast growth factor 21), UCP1 (Uncoupling protein 1), or PRDM16 (PRDI-BF1 and RIZ homology domain containing 16), which are factors related to the conversion of brown adipocytes, was increased.

[0032] Such results mean that the peptide of the present invention has the activity of reducing intracellularly accumulated fat.

[0033] Another aspect of the present invention provides a pharmaceutical composition for preventing or treating obesity, comprising a peptide containing the amino acid sequence of SEQ ID NO: 1 as an active ingredient.

[0034] As used herein, "obesity" means a state (condition) or disease in which body fat is excessively accumulated in the body due to an energy imbalance. Specifically, "obesity" may mean fat deposition, and may further be used to mean including "obesity-related diseases" that can be induced by a state of having excessive body fat due to an energy imbalance.

[0035] Said "fat deposition" may mean a state selected from the group consisting of, for example, obesity, lipodystrophy syndrome, lower eyelid fat hernia, lipoma, Dercum's disease, lipodystrophy, visceral adiposity, gynecomastia, morbid obesity, flank and diffuse body fat around the arm, and fat deposition related to cellulite.

[0036] The "obesity-related disease" may be hypertension, diabetes, increased plasma insulin concentration, insulin resistance, hyperlipidemia, metabolic syndrome, insulin resistance syndrome, obesity-related gastroesophageal reflux, arteriosclerosis, hypercholesterolemia, hyperuricemia, low back pain, cardiac hypertrophy, and left ventricular hypertrophy, lipodystrophy, non-alcoholic fatty hepatitis, cardiovascular disease, polycystic ovary syndrome, etc.

[0037] In this specification, the term "prevention" means all acts that suppress or delay the onset of obesity, and the term "improvement or treatment" includes all acts that favorably modify obesity, such as reducing, alleviating, or eliminating the symptoms of obesity, or retarding, interrupting, or reversing the progression of obesity.

[0038] The peptide of the present invention can prevent, improve, or treat the obesity by not only significantly increasing the expression of the lipolysis-promoting gene and the protein encoded by the gene, but also increasing the expression of the brown adipocyte conversion-related gene (thermogenesis-related gene) and the protein to regulate the energy imbalance or deviate from the state of having excessive body fat.

[0039] In a specific embodiment of the present invention, the peptide can prevent, improve, or treat obesity by increasing the expression of Perilipin1, HSL, and / or ATGL in differentiated adipose progenitor cells.

[0040] In a specific embodiment of the present invention, the peptide can prevent, improve, or treat obesity by increasing the expression of PGC1α, TBx1, FGF21, UCP1, and / or PRDM.

[0041] The pharmaceutical composition for preventing or treating obesity of the present invention may further contain a pharmaceutically acceptable carrier.

[0042] Pharmaceutically acceptable carriers may further include, for example, carriers for oral administration or carriers for parenteral administration. Carriers for oral administration may include lactose, starch, cellulose derivatives, magnesium stearate, stearic acid, and the like. In addition, carriers for parenteral administration may include water, suitable oils, saline, aqueous glucose, and glycols. Stabilizers and preservatives may also be further included. Suitable stabilizers include antioxidants such as sodium bisulfite, sodium sulfite, or ascorbic acid. Suitable preservatives include benzalkonium chloride, methyl- or propyl-paraben, and chlorobutanol. Reference can be made to what is described in Remington’s Pharmaceutical Sciences (19th ed., Mack Publishing Company, Easton, PA, 1995) for suitable pharmaceutically acceptable carriers.

[0043] The pharmaceutical composition of the present invention may be administered to mammals including humans by any method. For example, it may be administered orally or parenterally, and parenteral administration methods include, but are not limited to, intravenous, intramuscular, intraarterial, intramedullary, intradural, intracardiac, transdermal, subcutaneous, intraperitoneal, intranasal, enteral, topical, sublingual, or rectal administration.

[0044] The pharmaceutical composition of the present invention may be formulated into a preparation for oral administration or a preparation for parenteral administration according to the administration route described above. Specifically, the preparation for parenteral administration may be formulated and used in the form of an injection preparation or a topical preparation.

[0045] When the pharmaceutical composition of the present invention is used as an injection preparation, it may be used for the purpose of removing localized fat deposits.

[0046] The localized fat may be localized in a target site selected from the group consisting of the abdomen, under the eyes, under the chin, under the arms, buttocks, thighs, calves, back, thighs, and bra line.

[0047] The pharmaceutical composition of the present invention is administered in a pharmaceutically effective amount. In the present invention, "pharmaceutically effective amount" means an amount sufficient to treat a disease at a reasonable benefit / risk ratio applicable to medical treatment, and the effective dose level may be determined according to factors including the type of the patient's disease, severity, drug activity, sensitivity to the drug, administration time, administration route, and excretion rate, treatment period, factors including co-administered drugs, and other factors well known in the medical field. The pharmaceutical composition may be administered as an individual therapeutic agent or in combination with other anti-obesity agents, and may be administered simultaneously with, separately from, or sequentially to conventional therapeutic agents, and may be administered in single or multiple doses. It is important to administer an amount that can achieve the maximum effect with the minimum amount without side effects considering all of the above factors, and this can be easily determined by those skilled in the art.

[0048] The effective amount of the pharmaceutical composition may vary depending on the patient's age, sex, condition, weight, absorption degree of the active ingredient in the body, inactivation rate, excretion rate, disease type, co-administered drugs, etc., and may increase or decrease according to the administration route, severity of obesity, sex, weight, age, etc. For example, the pharmaceutical composition may be administered at about 0.0001 μg to 500 mg, preferably 0.01 μg to 100 mg, per kg of the patient's body weight per day.

[0049] Yet another aspect of the present invention provides a composition for promoting lipolysis containing, as an active ingredient, a peptide containing the amino acid sequence represented by SEQ ID NO: 1.

[0050] The composition may be for removing localized fat deposits.

[0051] The localized fat may mean fat localized at a target site selected from the group consisting of the abdomen, under the eyes, under the chin, under the arms, buttocks, thighs, calves, back, thighs, and bra line.

[0052] Yet another aspect of the present invention provides a cosmetic composition for preventing or improving obesity containing, as an active ingredient, a peptide containing the amino acid sequence of SEQ ID NO: 1.

[0053] The cosmetic composition may be manufactured into any dosage form that is commonly manufactured in the art, for example, solutions, suspensions, emulsions, pastes, gels, creams, lotions, powders, soaps, surfactant-containing cleansing, oils, powder foundations, emulsion foundations, wax foundations, and sprays, etc., but is not limited thereto.

[0054] The cosmetic composition may be manufactured in various forms such as solutions, sol-gels, emulsions, oils, waxes, aerosols, etc., like softening lotions, nutritive lotions, nutritive creams, massage creams, essences, eye creams, cleansing creams, cleansing foams, cleansing waters, packs, sprays, powders, hair tonics, hair creams, hair lotions, hair shampoos, hair rinses, hair conditioners, hair sprays, hair aerosols, pomades, gels, etc., but is not limited thereto.

[0055] The cosmetic composition of the present invention may contain other additives such as excipients and carriers, and it is possible to apply and blend as many ordinary components as necessary for general skin cosmetics.

[0056] When the dosage form of the cosmetic composition is in the form of a paste, cream, or gel, animal oils, vegetable oils, waxes, paraffin, starches, tragacanth, cellulose derivatives, polyethylene glycols, silicones, bentonite, silica, talc, or zinc oxide, etc. may be used as the carrier component.

[0057] When the dosage form of 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 the carrier component. Particularly in the case of a spray, a propellant such as chlorofluorohydrocarbon, propane / butane, or dimethyl ether may further be included, but is not limited thereto.

[0058] When the dosage form of the cosmetic composition is a solution or an emulsion, a solvent, a solubilizing agent, or an emulsifying agent may be used as the carrier component. For example, water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol oil, glycerol fatty acid ester, polyethylene glycol, or fatty acid ester of sorbitan may be used.

[0059] When the dosage form of the cosmetic composition is a suspension, a liquid diluent such as water, ethanol, or propylene glycol, a suspending agent such as ethoxylated isostearyl alcohol, polyoxyethyl sorbitol ester, and polyoxyethylene sorbitan ester, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar, or tragacanth may be used as the carrier component.

[0060] When the dosage form of the cosmetic composition is surfactant-containing cleansing, an aliphatic alcohol sulfate, an aliphatic alcohol ether sulfate, a sulfosuccinic acid monoester, an isethionate, an imidazolinium derivative, a methyl taurate, a sarcosinate, a fatty acid amide ether sulfate, an alkylamide betaine, an aliphatic alcohol, a fatty acid glyceride, a fatty acid diethanolamide, a vegetable oil, a lanolin derivative, or an ethoxylated glycerol fatty acid ester may be used as the carrier component.

[0061] When the dosage form of the cosmetic composition is a hair shampoo, the present invention of the page Puchi to the pageBase components for composing shampoo, such as thickeners, surfactants, viscosity modifiers, humectants, pH adjusters, preservatives, and essential oils, may be mixed. CDE may be used as the thickener, LES, an anionic surfactant, and coco betaine, an amphoteric surfactant, may be used as the surfactant, polyquaternium may be used as the viscosity modifier, glycerin may be used as the humectant, citric acid and sodium hydroxide may be used as the pH adjuster. Grapefruit extract or the like may be used as the preservative, and in addition, essential oils such as sidalwood, peppermint, and rosemary, silk amino acids, pentaol, or vitamin E may be added.

[0062] The components contained in the cosmetic composition, as active ingredients, the peptides of the present invention record page Puchi page and In addition to the carrier component, the cosmetic composition may further contain components commonly used in cosmetic compositions, such as antioxidants, stabilizers, solubilizers, vitamins, pigments, and ordinary auxiliaries such as fragrances, but is not limited thereto.

[0063] As described above, since the peptide of the present invention has the effect of promoting the decomposition of fat accumulated in cells, it may be used as a raw material for pharmaceuticals for the purpose of preventing, improving, or treating obesity or obesity-related diseases, or cosmetics for the purpose of body slimming.

[0064] Yet another aspect of the present invention provides a method for preventing or treating obesity, comprising administering to a subject a therapeutically effective amount of a peptide comprising the amino acid sequence of SEQ ID NO: 1.

[0065] Yet another aspect of the present invention provides a method for promoting lipolysis, comprising administering to a subject a therapeutically effective amount of a peptide comprising the amino acid sequence of SEQ ID NO: 1.

[0066] Since the peptide containing the amino acid sequence of SEQ ID NO: 1 is the same as the peptide described in the item "Pharmaceutical composition for preventing or treating obesity", the specific description refers to the content described above.

[0067] As used herein, the term "therapeutically effective amount" means the amount of the peptide that achieves the purpose of improving the incidence of the disease during the treatment of the peptide and usually avoids harmful side effects related to other treatment methods.

[0068] Still another aspect of the present invention provides the use of the amino acid sequence of SEQ ID NO: 1 for use in the manufacture of a medicament for preventing or treating obesity.

[0069] Hereinafter, the present invention will be described in detail with reference to Examples and Experimental Examples.

[0070] However, the following Examples and Experimental Examples are only for illustrating the present invention, and the content of the present invention is not limited by the following Examples and Experimental Examples.

[0071] [Synthesis Example 1] Peptide synthesis containing the amino acid sequence of SEQ ID NO: 1 Using an automatic peptide synthesizer (Liberty, CEM Corporation, USA), a peptide having the amino acid sequence of SEQ ID NO: 1 described in Table 1 below was synthesized, and these synthesized peptides were purified by C18 reverse-phase high-performance liquid chromatography (HPLC) (U-3000, Thermo fisher scientific, USA). The column used was Pursuit XRs C18 (250*4.65mm 100Å, Agilent, USA).

Table 1

[0072] [Preparation Example 1]

[0073] Preparation of preadipocytes and induction of differentiation into adipocytes

[0074] First, to differentiate preadipocytes into adipocytes, preadipocytes, 3T3-L1 cells purchased from the American Type Culture Collection (ATCC), were cultured in DMEM containing 2% fetal bovine serum (FBS). To differentiate 3T3-L1 cells into adipocytes, they were dispensed into a 12-well plate at a density of 1x10 5 cells / well, and when they grew to confluence (day0), the culture medium was changed to a DMEM medium containing 10% FBS, 10 μg / ml insulin, 0.5 mM isobutylmethylxanthine (IBMX) (Sigma, USA), and 1 μM dexamethasone (sigma, USA) and cultured for 2 days. Two days later (day2), the culture medium was changed to a DMEM medium containing 10% FBS and 10 μg / ml insulin and cultured. Three days later (day5) from this time, the culture medium was changed to a medium containing only 10% FBS and cultured. Two days later (day7) from this time, they were named 3T3-L1 adipocytes, and the cell culture conditions were 37°C, 5% CO 2 environment and cultured. Adipocytes induced to differentiate for 10 days were used in subsequent experiments.

[0075] [Experimental Example 1]

[0076] Confirmation of changes in the expression of genes related to lipolysis in adipocytes

[0077] In adipocytes, to confirm whether the peptide of the present invention has an active effect on the lipolysis-promoting cell signaling pathway, the mRNA expression levels of genes related to lipolysis promotion, HSL, ATGL, and Perilipin1, were confirmed.

[0078] Specifically, 5 μM and 50 μM of the peptide of SEQ ID NO: 1 and TNF-α, a positive control group, were treated at 20 nM to the adipocytes differentiated through Preparation Example 1 and cultured for 72 hours, and then the cells were collected and total RNA was extracted with Trizol (Trizol, Thermo Fisher Scientific, USA). The extracted RNA was TOPScript TMAfter obtaining DNA complementary to RNA using RT DryMIX (Enzynomics, Korea), TOPsimple TM After performing polymerase chain reaction (PCR) using DryMIX-nTaq (Enzynomics, Korea), the PCR products were loaded onto a 1.5% agarose gel to compare the mRNA expression levels of the genes related to promoting lipolysis. The GAPDH gene was used as an internal control group, and the primers used in the experiment are listed in [Table 2].

Table 2

[0079] As a result, as shown in Figures 1a to 1d, after treatment with the peptide, the expressions of the HSL, ATGL, and Perilipin1 genes related to promoting lipolysis increased. Thus, it can be seen that the peptide of the present invention increases the expression of the genes related to promoting lipolysis so that fat can be decomposed very effectively.

[0080] [Experimental Example 2]

[0081] Confirmation of the change in the expression of lipolysis-related proteins in adipocytes

[0082] In adipocytes, to confirm whether the peptide of the present invention has an active effect on the lipolysis-promoting cell signaling pathway, the expression level of the protein HSL related to promoting lipolysis was confirmed.

[0083] Specifically, the differentiated adipocytes through Preparation Example 1 were treated with 5 μM and 50 μM of the peptide of SEQ ID NO: 1, and the cells were collected after 1 hour, and cell lysis buffer was treated to obtain lysates. Then, the proteins were quantified, and Western blotting was performed using an anti-pHSL antibody (Cell signaling technology (CST), USA).

[0084] As a result, as shown in FIGS. 2a and 2b, after treating the peptide, phosphorylation of the HSL protein related to promoting lipolysis increased.

[0085] Thus, it can be seen that the peptide of the present invention increases the expression of proteins related to promoting lipolysis so that fat can be decomposed very effectively.

[0086] [Experimental Example 3]

[0087] Measurement of the amount of glycerol, a lipolysis product, in adipocytes

[0088] Neutral fat accumulated in adipocytes is decomposed into fatty acids and glycerol for energy production or cell signaling, etc. Therefore, the decomposition effect of neutral fat accumulated in adipocytes was confirmed by measuring the release amount of free glycerol.

[0089] Specifically, the peptide of SEQ ID NO: 1 was treated at concentrations of 5 μM and 50 μM on adipocytes differentiated through Preparation Example 1, and TNFα was treated at 20 nM in the positive control group and cultured for 48 hours. Then, the supernatant was obtained and the amount of glycerol was measured using a glycerol calorimetric assay kit (Cayman chemical, USA).

[0090] As a result, as shown in FIG. 3, when the peptide of SEQ ID NO: 1 was treated, it was confirmed that the amount of glycerol released while the fat was decomposed increased in a concentration-dependent manner of the peptide.

[0091] Thus, it can be seen that the peptide of the present invention has the activity of decomposing neutral fat accumulated in adipocytes.

[0092] [Experimental Example 4]

[0093] Confirmation of increased expression of genes related to thermogenesis by the peptide of the present invention

[0094] To confirm whether the peptide of the present invention has an effect of preventing or treating obesity, the mRNA expression levels of genes related to thermogenesis in brown adipocytes, namely, PGC1α, Tbx1, FGF21, and UCP1, were confirmed.

[0095] Specifically, adipocytes obtained through Preparation Example 1 were treated with 5 μM and 50 μM of the peptide of SEQ ID NO: 1 and 20 nM of CL316,243 (Sigma, C5976), which is a positive control group, and cultured for 72 hours. Then, the cells were collected and total RNA was extracted using Trizol (Thermo Fisher Scientific, USA). The extracted RNA was used to obtain DNA complementary to the RNA using TOPScript TM RT DryMIX (enzynomics, Korea), and then polymerase chain reaction (PCR) was performed using TOPsimple TM DryMIX-nTaq (enzynomics, Korea). After that, the PCR products were loaded onto a 1.5% agarose gel to compare the mRNA expression levels of the genes related to thermogenesis in the brown adipocytes. The GAPDH gene was used as an internal control group, and the primers used in the experiment are shown in [Table 3].

Table 3

[0096] As a result, as shown in FIGS. 4a to 4e, it was confirmed that after treatment with the peptide of SEQ ID NO: 1, the expression of genes PGC1α, Tbx1, FGF21, and UCP1 related to thermogenesis in brown adipocytes increased. Thus, it can be seen that the peptide of the present invention has an effect of preventing or treating obesity by increasing the expression of genes related to thermogenesis that increase the beta-oxidation of free fatty acids.

[0097] [Experimental Example 5]

[0098] Confirmation of increased expression of thermogenesis-related proteins by the peptide of the present invention

[0099] In order to confirm whether the peptide of the present invention has an effect of preventing or treating obesity, the expression levels of PRDM16 and UCP1, which are proteins related to the thermogenesis of brown adipocytes, were confirmed.

[0100] Specifically, the adipocytes obtained through Preparation Example 1 were treated with 5 μM and 50 μM of the peptide of SEQ ID NO: 1, respectively. After 24 hours, the cells were collected, treated with cell lysis buffer to obtain lysates. Then, the proteins were quantified, and Western blotting was performed using an anti-PRDM16 antibody (Cell signaling technology (CST), USA) and an anti-UCP1 (abcam, England) antibody.

[0101] As a result, as shown in FIGS. 5a to 5c, after treatment with the peptide, the expression of thermogenic marker proteins that increase the beta-oxidation of free fatty acids increased.

[0102] From this, it can be seen that the peptide of the present invention increases the expression of proteins related to fatty acidification so that fat can be decomposed very effectively.

[0103] As described above, the present invention has been described in detail only for the described embodiments, but it is obvious to those skilled in the art that various modifications and variations are possible within the scope of the technical idea of the present invention, and such modifications and variations naturally belong to the appended claims.

Claims

1. A peptide consisting of the amino acid sequence represented by SEQ ID NO:

1.

2. The peptide according to claim 1, which has an activity of reducing intracellularly accumulated fat.

3. A pharmaceutical composition for preventing or treating obesity, comprising the peptide according to claim 1 as an active ingredient.

4. The pharmaceutical composition for preventing or treating obesity according to claim 3, wherein the composition increases the expression of perilipin 1 (Perilipin1), HSL (Hormone sensitive lipase), or ATGL (Adipose triglyceride lipase), which are fat decomposition-related factors.

5. The pharmaceutical composition for preventing or treating obesity according to claim 3, wherein the composition increases the expression of PGC1α (peroxisome proliferator-activated receptor gamma coactivator 1-alpha), TBx1 (T-box transcription factor), FGF21 (Fibroblast growth factor 21), UCP1 (Uncoupling protein1), or PRDM16 (PRDI-BF1 and RIZ homology domain containing16), which are heat generation-related factors.

6. The pharmaceutical composition for preventing or treating obesity according to claim 3, wherein the pharmaceutical composition is in the form of an oral preparation, an injectable preparation, or a topical preparation.

7. A composition for promoting lipolysis, comprising the peptide according to claim 1 as an active ingredient.

8. The composition for promoting lipolysis according to claim 7, which is for removing localized fat deposits.

9. The localized fat according to claim 8 is localized at a target site selected from the group consisting of the abdomen, under the eyes, under the chin, under the arms, buttocks, calves, back, thighs, and bra line. The composition for promoting lipolysis.

10. A cosmetic composition for preventing or improving obesity, comprising the peptide according to claim 1 as an active ingredient.

11. The cosmetic composition according to claim 10 for preventing or improving obesity has any one or more dosage forms selected from the group consisting of skin, lotion, cream, essence, emulsion, gel, hand cream, lipstick, cleansing foam, cleansing cream, cleansing water, aerosol, shampoo, conditioner, treatment, body cleanser, soap, pack, massage agent, face powder, compact, foundation, two-way cake, and makeup base.

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