Deoxycholic acid-peptide conjugate with anti-obesity activity and use thereof

Abstract

The present invention relates to: a deoxycholic acid-peptide conjugate, which has anti-obesity activity by inhibiting lipid production and promoting lipolysis; and a use of the conjugate as an active ingredient for preventing, improving or treating obesity.

Description

Technical Field

[0001] The present invention relates to a deoxycholic acid - peptide conjugate having a structure in which deoxycholic acid and a peptide are chemically conjugated and uses thereof. Background Art

[0002] Obesity refers to a state in which, when the energy consumption from food intake is not balanced, excess energy accumulates as body fat, resulting in an excessive amount of adipose tissue in the body. Obesity is considered an independent disease and is also a risk factor for various diseases such as cardiovascular diseases and diabetes, and local obesity with excessive fat deposition in a specific area such as the abdomen is also closely related to these diseases. Therefore, local obesity has a direct impact on health and quality of life, and aesthetically, local obesity also causes atrophy in individual psychology and social maladjustment.

[0003] The treatment of such local obesity can be mainly divided into surgical treatment and non - surgical treatment, but surgical operations have side effects such as an uneven skin surface and side effects of anesthesia. Non - surgical treatment has used various methods such as lipolysis needles, ultrasound, high - frequency treatment, drug injection methods such as Meso - therapy, massage methods, gas injection methods, etc.

[0004] Meanwhile, deoxycholic acid, as one of the secondary bile acids that are metabolic by - products of intestinal bacteria, has been confirmed to induce an inflammatory effect in adipose tissue to kill adipocytes (see the literature Rotunda et al., Dermatol Surg 30(7):1001 - 8(2004)), and deoxycholic acid has been approved by the FDA and has been used as a component of lipolysis injections. However, when deoxycholic acid is topically administered, side effects such as erythema, edema, ecchymosis, hematoma, and pain occur at the administration site, and due to its low solubility in water, an organic solvent is added to dissolve deoxycholic acid, but this has limitations in causing side effects such as dermatitis and skin dryness.

[0005] In this context, the present inventors conducted research to develop a method for increasing anti - obesity activity while minimizing side effects caused by deoxycholic acid, and found that, compared with using a single substance, a deoxycholic acid - peptide conjugate in which a new peptide synthesized by the present inventors is chemically conjugated with deoxycholic acid has high solubility in water and significantly improved anti - obesity activity, and then completed the present invention. Summary of the Invention

[0006] [Technical Problem]

[0007] An object of the present invention is to provide a deoxycholic acid - peptide conjugate which has improved solubility in water and improved adipogenesis inhibitory activity and lipolysis activity compared with a single substance.

[0008] Another object of the present invention is to provide a composition for inhibiting adipogenesis or promoting lipolysis.

[0009] Another object of the present invention is to provide a method for inhibiting adipogenesis or promoting lipolysis.

[0010] Another object of the present invention is to provide a deoxycholic acid-peptide conjugate for inhibiting adipogenesis or promoting lipolysis.

[0011] Another object of the present invention is to provide a composition for preventing, treating or improving obesity or obesity-related diseases.

[0012] Another object of the present invention is to provide a method for preventing or treating obesity or obesity-related diseases.

[0013] Another object of the present invention is to provide a deoxycholic acid-peptide conjugate for preventing or treating obesity or obesity-related diseases.

[0014] [Technical Solution]

[0015] One aspect of the present invention provides a deoxycholic acid-peptide conjugate, wherein a peptide consisting of the amino acid sequence of SEQ ID NO: 1 is conjugated with deoxycholic acid.

[0016] Another aspect of the present invention provides a composition for inhibiting adipogenesis or promoting lipolysis, which comprises the deoxycholic acid-peptide conjugate as an active ingredient.

[0017] Another aspect of the present invention provides a method for inhibiting adipogenesis or promoting lipolysis, comprising administering the deoxycholic acid-peptide conjugate according to claim 1 to a subject.

[0018] Another aspect of the present invention provides a deoxycholic acid-peptide conjugate for inhibiting adipogenesis or promoting lipolysis.

[0019] Another aspect of the present invention provides a composition for preventing, treating or improving obesity or obesity-related diseases, which comprises the deoxycholic acid-peptide conjugate as an active ingredient.

[0020] Another aspect of the present invention provides a method for preventing or treating obesity or obesity-related diseases, comprising administering the deoxycholic acid-peptide conjugate according to claim 1 to a subject.

[0021] Another object of the present invention is to provide a deoxycholic acid-peptide conjugate for preventing or treating obesity or obesity-related diseases.

[0022] [Advantageous Effects]

[0023] According to the present invention, the deoxycholic acid - peptide conjugate has the effects of inhibiting lipid production and accumulation in adipocytes and decomposing fat, and thus can be used for preventing, improving or treating obesity.

[0024] However, the effects of the present invention are not limited to the above - mentioned effects, and those skilled in the art will clearly understand other effects not mentioned according to the following description. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 The NMR analysis data of the conjugate of the peptide of SEQ ID NO:1 of the present invention and deoxycholic acid are shown.

[0026] Figure 2 The photograph shows the results of confirming the solubility of deoxycholic acid and deoxycholic acid - peptide conjugate in water.

[0027] Figure 3A is a result graph showing the effect of confirming that the deoxycholic acid - peptide conjugate inhibits the differentiation of adipocytes and intracellular lipid accumulation through intracellular fat staining.

[0028] Figure 3B is a result graph showing the effect of confirming that the deoxycholic acid - peptide conjugate enhances the fat accumulation inhibitory effect through intracellular fat staining.

[0029] Figure 4A is a result graph showing the effect of confirming that the deoxycholic acid - peptide conjugate promotes intracellular lipolysis through the release amount of glycerol released from adipocytes.

[0030] Figure 4B is a result graph showing the effect of confirming that the deoxycholic acid - peptide conjugate promotes intracellular lipolysis through the reduction of the size of adipocytes in adipose tissue.

[0031] Figure 4C is a result graph showing the effect of confirming that the deoxycholic acid - peptide conjugate enhances the lipolysis effect through the release amount of glycerol released from adipocytes.

[0032] Figure 5 The figure shows the results of confirming that the deoxycholic acid - peptide conjugate increases the expression of adiponectin and leptin genes related to intracellular fat metabolism.

[0033] Figure 6A is a result graph showing the effect of confirming that the deoxycholic acid - peptide conjugate increases the expression of lipolysis - related cell signaling pathway genes AMPKa1, HSL and PGCLa in adipocytes.

[0034] Figure 6B is a result graph showing the effect of confirming that the deoxycholic acid - peptide conjugate increases the expression of lipolysis - related cell signaling pathway genes HSL and PLIN in adipose tissue.

[0035] Figure 7The figure shows the results of confirming that the deoxycholic acid-peptide conjugate reduces the expression of the adipogenesis-related cellular signaling pathway genes ACCα, FAS, and PPARγ in adipose tissue.

[0036] Figure 8 The figure shows the results of confirming the increased expression of the thermogenic genes UCP1, PRDM, and Cidea in brown adipocytes in adipose tissue.

[0037] Figure 9 The figure shows the results of confirming the increased expression of the IL-15 gene related to the inhibition of lipid accumulation in myocytes. Detailed Description

[0038] Hereinafter, the present invention will be described in detail.

[0039] One aspect of the present invention provides a peptide comprising the amino acid sequence of SEQ ID NO:1 and an active substance-peptide conjugate, wherein the active substance is conjugated to the peptide.

[0040] The peptide of the present invention can be synthesized using, for example, a device or genetic engineering techniques. In the case of synthesis using a device, the desired peptide can be synthesized by the Fmoc solid-phase method in an automatic peptide synthesizer.

[0041] In a specific exemplary embodiment, the peptide of the present invention comprising the amino acid sequence of SEQ ID NO:1 is synthesized and identified using the Fmoc solid-phase method, and is selected by verifying the efficacy of the identified peptide.

[0042] In a specific exemplary embodiment, in order to verify the efficacy of the identified peptide, a peptide having anti-obesity activity is selected by verifying i) the adipogenesis inhibitory effect and ii) the lipolytic effect.

[0043] Furthermore, the active substance-peptide conjugate of the present invention can be prepared by conjugating the peptide selected by this method with an active substance. In a specific exemplary embodiment, the active substance is selected by verifying the efficacy of the active substance-peptide conjugate prepared by chemical conjugation with the peptide.

[0044] In the present specification, the term "peptide" refers to a linear molecule composed of amino acid residues, and specifically, the peptide of the present invention may include the amino acid sequence represented by SEQ ID NO:1.

[0045] "Peptide" can be an amino acid variant or fragment with different sequences obtained by deletion, insertion, substitution of amino acid residues or a combination thereof within a range that does not affect the function. Amino acid exchanges that do not completely change the activity of the peptide are known in the art. In some cases, the amino acid exchanges can be modified by phosphorylation, sulfation, acrylation, glycosylation, methylation, farnesylation, etc. Therefore, the present invention includes peptides having an amino acid sequence substantially the same as the peptide having the amino acid sequence of SEQ ID NO:1, as well as variants or active fragments of the peptide. Substantially identical proteins refer to amino acid sequences having a sequence homology of at least 75%, preferably at least 80%, such as at least 85%, at least 90%, at least 95%, at least 98% or at least 99% with the amino acid sequence of SEQ ID NO:1, but not limited thereto, and amino acid sequences having a homology of more than 75% and the same activity are included within the scope of the present invention. In addition, the peptides of the present invention may also include amino acid sequences prepared for specific purposes such as increasing the targeting sequence, tag, labeled residue, half-life or peptide stability.

[0046] In addition, in order to obtain better chemical stability, enhanced pharmacological properties (half-life, absorbability, potency, efficacy, etc.), modified specificity (e.g., broad bioactivity profile) and reduced antigenicity, a protecting group can be bound to the N-terminus or C-terminus of the peptide. For example, the protecting group can be acetyl, fluorenylmethoxycarbonyl, formyl, palmitoyl, myristoyl, stearyl or polyethylene glycol (PEG), however, the protecting group can include any component that can enhance the modification effect on the peptide, especially any component that can enhance the stability of the peptide, without limitation. "Stability" is used to refer to the following meaning: it includes not only the in vivo stability that protects the peptide of the present invention from the attack of proteolytic enzymes in vivo, but also the storage stability (e.g., room temperature storage stability).

[0047] In the present specification, the "active substance" can be a bile acid. Specifically, bile acids selected from cholic acid, 5β-cholanic acid, chenodeoxycholic acid, glycocholic acid, taurocholic acid, deoxycholic acid, lithocholic acid, 7-oxolithocholic acid, etc. can be preferably used, and more specifically, the active substance can be deoxycholic acid.

[0048] The "deoxycholic acid" of the present invention is a compound with the IUPAC name of 3,12-dihydroxycholan-24-oic acid and a molecular weight of 392.58. Generally, deoxycholic acid is a bile acid that decomposes fat and destroys fat cells, and deoxycholic acid is used in injectables for improving submental fat. The structure of deoxycholic acid is shown in the following Chemical Formula 1.

[0049] [Chemical Formula 1]

[0050]

[0051] In the active substance - peptide conjugate of the present invention, when the active substance is deoxycholic acid, the peptide conjugated with deoxycholic acid is referred to as "deoxycholic acid - peptide conjugate (DA - peptide conjugate)".

[0052] Specifically, the present invention provides a deoxycholic acid - peptide conjugate, wherein deoxycholic acid is conjugated with a peptide consisting of the amino acid sequence of SEQ ID NO:1.

[0053] Deoxycholic acid can be conjugated to the N - terminus, C - terminus or side chain of the peptide. Specifically, the carboxyl group of deoxycholic acid can bind to the N - terminal peptide of the peptide.

[0054] An embodiment of the deoxycholic acid - peptide conjugate can be represented by the following Chemical Formula 2.

[0055] [Chemical Formula 2]

[0056]

[0057] In a specific exemplary embodiment of the present invention, the deoxycholic acid - peptide conjugate is administered to adipocytes and adipose tissue to inhibit the lipogenesis signaling pathway and inhibit fat production and accumulation, thereby preventing, ameliorating or treating obesity.

[0058] In addition, the deoxycholic acid - peptide conjugate of the present invention can activate the lipolysis signaling pathway to promote lipolysis of adipocytes and adipose tissue and reduce the size of adipocytes, thereby preventing, ameliorating or treating obesity.

[0059] In addition, compared with deoxycholic acid or the peptide alone, the deoxycholic acid - peptide conjugate of the present invention has high solubility in water and has a significantly improved effect of inhibiting fat accumulation and lipolysis effect, so as to reduce the usage amount of deoxycholic acid, thereby preventing side effects caused by using high - dose deoxycholic acid.

[0060] Therefore, the deoxycholic acid - peptide conjugate having the above activities can be used as an active ingredient of a composition for preventing, treating or ameliorating obesity or obesity - related diseases.

[0061] Therefore, another aspect of the present invention provides a composition for preventing, treating or ameliorating obesity or obesity - related diseases, which comprises a deoxycholic acid - peptide conjugate, wherein deoxycholic acid is conjugated with a peptide including the amino acid sequence of SEQ ID NO:1.

[0062] Another aspect of the present invention provides a composition for inhibiting lipogenesis or promoting lipolysis, which comprises a deoxycholic acid - peptide conjugate, wherein deoxycholic acid is conjugated with a peptide including the amino acid sequence of SEQ ID NO:1.

[0063] In addition, the present invention provides a method for inhibiting adipogenesis or promoting lipolysis, which includes administering a deoxycholic acid-peptide conjugate to a subject, wherein the deoxycholic acid is conjugated to a peptide comprising the amino acid sequence of SEQ ID NO:1.

[0064] In addition, the present invention provides a deoxycholic acid-peptide conjugate, wherein the deoxycholic acid is conjugated to a peptide comprising the amino acid sequence of SEQ ID NO:1, and the deoxycholic acid-peptide conjugate is used for inhibiting adipogenesis or promoting lipolysis.

[0065] Another aspect of the present invention provides a pharmaceutical composition for preventing or treating obesity or obesity-related diseases, which comprises a deoxycholic acid-peptide conjugate, wherein the deoxycholic acid is conjugated to a peptide comprising the amino acid sequence of SEQ ID NO:1.

[0066] In addition, the present invention provides a method for preventing or treating obesity or obesity-related diseases, which includes administering a deoxycholic acid-peptide conjugate to a subject, wherein the deoxycholic acid is conjugated to a peptide comprising the amino acid sequence of SEQ ID NO:1.

[0067] In addition, the present invention provides a deoxycholic acid-peptide conjugate, wherein the deoxycholic acid is conjugated to a peptide comprising the amino acid sequence of SEQ ID NO:1, and the deoxycholic acid-peptide conjugate is used for preventing or treating obesity or obesity-related diseases.

[0068] In this specification, "obesity" refers to a condition or disease in which body fat accumulates excessively in the body due to energy imbalance. The deoxycholic acid-peptide conjugate of the present invention not only significantly increases the expression of genes promoting fat oxidation and the proteins encoded by these genes, as well as the expression of genes promoting lipolysis and the proteins encoded by these genes, but also reduces the expression of genes involved in adipocyte accumulation, so as to control the imbalance or get out of the state with excessive body fat, thereby preventing or treating obesity.

[0069] In this specification, "preventing or treating obesity" may refer to reducing fat deposition, and "fat deposition" may refer to a condition selected from the group consisting of, for example, obesity, lipodystrophy syndrome, lower eyelid fat hernia, lipoma, Derkham disease, lipodystrophy, visceral adiposity, gynecomastia, morbid obesity, body fat distributed around both sides of the body and the arms, and fat deposition related to cellulite.

[0070] In addition, in this specification, "obesity-related diseases" are diseases caused by conditions of excessive body fat due to energy imbalance, and the symptoms caused by obesity-related diseases can be prevented, alleviated, or improved by preventing or treating obesity. Specifically, obesity-related diseases may include hypertension, diabetes, elevated plasma insulin concentration, insulin resistance, hyperlipidemia, metabolic syndrome, insulin resistance syndrome, obesity-related gastroesophageal reflux, arteriosclerosis, hypercholesterolemia, hyperuricemia, lower back pain, cardiac hypertrophy and left ventricular hypertrophy, lipodystrophy, non-alcoholic fatty liver disease, cardiovascular diseases, polycystic ovary syndrome, etc.

[0071] The composition for preventing or treating obesity or obesity-related diseases of the present invention can be formulated and used in the form of oral preparations such as powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, etc., topical preparations, suppositories, and sterile injection solutions according to conventional methods, and may include suitable carriers, excipients, or diluents commonly used for preparing pharmaceutical compositions for preparations.

[0072] Pharmaceutically acceptable carriers may include lactose, glucose, sucrose, sorbitol, mannitol, starch, acacia gum, calcium phosphate, alginate, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrup, methylcellulose, methylparaben, propylparaben, talc, magnesium stearate, and mineral oil, which are commonly used in preparations.

[0073] In addition to the above components, the pharmaceutical composition may further contain lubricants, wetting agents, sweeteners, flavoring agents, emulsifying agents, suspending agents, preservatives, etc.

[0074] The pharmaceutical composition can be administered orally or parenterally (e.g., intramuscular administration, intravenous administration, intraperitoneal administration, subcutaneous administration, intradermal administration, or topical administration) according to the required method, and its dosage varies according to the condition and weight of the patient, the degree of the disease, the form of the drug, and the administration route and time, and can be appropriately selected by those skilled in the art.

[0075] When the pharmaceutical composition of the present invention is used as an injection preparation, the pharmaceutical composition can be used to remove local fat deposits.

[0076] The local fat may be located in a target area selected from the group consisting of the abdomen, under the eyes, under the chin, under the arms, the buttocks, the thighs, the calves, the back, the thighs, and the bra line.

[0077] The pharmaceutical composition of the present invention is administered in a pharmaceutically effective dose. In the present invention, "pharmaceutically effective dose" means an amount sufficient to treat a disease with a reasonable benefit / risk ratio applicable to medical treatment, and the effective dose level can be determined based on factors including the type and severity of the patient's disease, the activity of the drug, the sensitivity to the drug, the administration time, the administration route, the excretion rate, the treatment duration, and the drugs used concomitantly, as well as other factors well known in the medical field. The pharmaceutical composition can be administered as a single therapeutic agent or can be administered in combination with other therapeutic agents, and the pharmaceutical composition can be administered simultaneously, separately, or sequentially with conventional therapeutic agents, and the pharmaceutical composition can be administered singly or multiple times. Importantly, by considering all elements, an amount that can achieve the maximum effect with the least amount without side effects can be easily determined by those skilled in the art.

[0078] The effective dose of the pharmaceutical composition can vary depending on the age, sex, condition and weight of the patient, the absorption, inactivation rate, and excretion rate of the active ingredient in the body, the type of disease, and the drugs used in combination, and can be increased or decreased depending on the administration route, the severity of obesity, sex, weight, age, etc. For example, the pharmaceutical composition can be administered at about 0.0001 μg to 500 mg, preferably 0.01 μg to 100 mg, per 1 kg of patient body weight per day.

[0079] In a specific exemplary embodiment, compared with the case of administering deoxycholic acid or peptide alone, when the deoxycholic acid-peptide conjugate of the present invention is administered to adipocytes or adipose tissue, it was confirmed that the fat accumulation inhibitory effect and the lipolytic effect were significantly increased. Therefore, when using the deoxycholic acid-peptide conjugate, the amount used can be reduced by a synergistic effect, thereby ensuring safety by using the lowest amount of deoxycholic acid and minimizing the side effects caused by deoxycholic acid. A "subject" may be an object in need of administering the deoxycholic acid-peptide conjugate of the present invention. Subjects in need of administration may include objects diagnosed with obesity or obesity-related diseases, objects who have developed obesity or obesity-related symptoms, and objects who wish to administer to prevent the development of diseases or symptoms or to improve health.

[0080] "Administration" means providing a predetermined substance to a patient by any suitable method, and the administration route of the deoxycholic acid-peptide conjugate of the present invention can be oral or parenteral administration through all common routes, as long as the deoxycholic acid-peptide conjugate can reach the target tissue. In addition, the deoxycholic acid-peptide conjugate can be administered by any device capable of moving the active substance to the target cell.

[0081] In yet another aspect, the present invention provides a quasi-pharmaceutical composition for preventing or improving obesity, which comprises a deoxycholic acid-peptide conjugate, wherein the deoxycholic acid is conjugated to a peptide comprising the amino acid sequence of SEQ ID NO:1.

[0082] Details of the peptide, deoxycholic acid, and obesity are as described above.

[0083] When the composition of the present invention is used as a quasi-drug composition, the deoxycholic acid-peptide conjugate can be added as it is or used together with other quasi-drug ingredients, and can be appropriately used according to conventional methods. The mixing amount of the active ingredient can be appropriately determined according to the purpose of use (prevention, health, or therapeutic treatment). According to an exemplary embodiment of the present invention, the quasi-drug composition of the present invention can be a disinfectant cleaner, toothpaste, shower foam, Garglin, wet wipes, decontamination soap, hand sanitizer, humidifier filler, mask, ointment, or filter filler.

[0084] In yet another aspect, the present invention provides a cosmetic composition for preventing or improving obesity, which comprises a deoxycholic acid-peptide conjugate, wherein the deoxycholic acid is conjugated to a peptide comprising the amino acid sequence of SEQ ID NO: 1.

[0085] Details of the peptide, deoxycholic acid, and obesity are as described above.

[0086] The cosmetic composition of the present invention can be prepared in any formulation commonly prepared in the art, and can be formulated by, for example, solutions, suspensions, emulsions, pastes, gels, creams, lotions, powders, soaps, surfactant-containing cleaning agents, oils, powder foundations, emulsion foundations, wax-based foundations, sprays, etc., but is not limited thereto.

[0087] The cosmetic composition can be prepared in various forms, such as softening lotion, nourishing emulsion, nourishing cream, massage cream, essence, eye cream, cleansing cream, cleansing foam, cleansing lotion, facial mask, spray, powder, hair oil, hair milk, shampoo, shampoo, hair dye, hair conditioner, hair spray, hair aerosol, hair pomade, solutions such as gels, sol-gels, emulsions, oils, waxes, aerosols, etc., but is not limited thereto.

[0088] In addition to the pentapeptide composed of the amino acid sequence of KFLIK, the cosmetic composition of the present invention can also contain other additives, such as excipients and carriers, and can be applied and mixed together with commonly used ingredients mixed in general skin cosmetics as needed.

[0089] When the formulation form of the cosmetic composition is a paste, cream, or gel, animal oil, vegetable oil, wax, paraffin wax, starch, tragacanth, cellulose derivatives, polyethylene glycol, silicone resin, bentonite, silica, talc, zinc oxide, etc. can be used as carrier components.

[0090] When the preparation form of the cosmetic composition is a powder or a spray, lactose, talc, silica, aluminum hydroxide, calcium silicate or polyamide powder can be used as the carrier component. In particular, in the case of a spray, the preparation may further contain a propellant such as chlorofluorocarbon, propane / butane or dimethyl ether, but is not limited thereto.

[0091] When the preparation form of the cosmetic composition is a solution or an emulsion, a solvent, a solubilizer or an emulsifier can be used as the carrier component. For example, the carrier component can be water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butanediol, glycerol fatty acid ester, polyethylene glycol or sorbitol fatty acid ester.

[0092] When the preparation 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, polyoxyethylene sorbitan ester and polyoxyethylene sorbitan polysaccharide ester, microcrystalline cellulose, partial aluminum hydroxide, bentonite, agar and tragacanth can be used as the carrier component.

[0093] When the preparation of the cosmetic composition is a surfactant-containing cleaning agent, fatty alcohol sulfate, fatty alcohol ether sulfate, sulfosuccinic acid monoester, hydroxyethyl sulfonate, imidazolinium derivative, methyl taurate, sarcosinate, fatty acid amide ether sulfate, alkyl amide betaine, fatty alcohol, fatty acid glyceride, fatty acid diethanolamide, vegetable oil, lanolin derivative, ethoxylated glycerol fatty acid ester, etc. can be used as the carrier component.

[0094] When the preparation of the cosmetic composition is a shampoo, the basic components for forming the shampoo such as a thickener, a surfactant, a viscosity regulator, a humectant, a pH regulator, a preservative and an essential oil can be mixed with the deoxycholic acid-peptide conjugate of the present invention. CDE can be used as a thickener, LES as an anionic surfactant and coconut oil betaine as an amphoteric surfactant can be used as surfactants, polyquaternium can be used as a viscosity regulator, glycerol can be used as a humectant, and citric acid and sodium hydroxide can be used as pH regulators. Grapefruit extract, etc. can be used as a preservative, and essential oils such as cedarwood, mint and rosemary, silk amino acid, amyl alcohol or vitamin E can also be added.

[0095] In addition to the deoxycholic acid-peptide conjugate of the present invention as the active ingredient and the carrier component, the components contained in the cosmetic composition can also include the components commonly used in cosmetic compositions, such as conventional auxiliaries such as antioxidants, stabilizers, solubilizers, vitamins, pigments and fragrances.

[0096] As described above, the deoxycholic acid - peptide conjugate of the present invention has the effects of inhibiting the differentiation of preadipocytes into adipocytes, inhibiting lipid accumulation in adipocytes, and promoting lipolysis, and thus can be used as a raw material for pharmaceutical compositions or cosmetic compositions to prevent, improve, or treat obesity or obesity - related diseases.

[0097] Hereinafter, the present invention will be described in detail through examples and experimental examples.

[0098] However, the following examples and experimental examples are only illustrative of the present invention, and the content of the present invention is not limited to the following examples and experimental examples.

[0099] [Example 1]

[0100] Synthesis of the peptide including the amino acid sequence of SEQ ID NO:1

[0101] The peptide of the present invention was synthesized by a known method. The amino acid sequence of the new peptide used in the present invention is "Lys PheLeu Ile Lys (SEQ ID NO:1)", and the molecular weight of the peptide including the amino acid sequence of SEQ ID NO:1 was measured with a molecular weight analyzer. As a result, it was confirmed that the molecular weight corresponded to 647.4 Da.

[0102] [Example 2]

[0103] Preparation of the deoxycholic acid - peptide conjugate

[0104] The peptide of SEQ ID NO:1 (500 mmol) and 1,000 mL of dimethylformamide (DMF) were added to a peptide reactor. 393 g of deoxycholic acid (1,000 mmol, 2 equivalents), 135 g of 1 - HOBt (1000 mmol, 2 equivalents), and 379 g of HBTU (1000 mmol, 2 equivalents) were added together with 500 ml of DMF and reacted at room temperature for 30 minutes. Thereafter, 261 mL (194 g, 1500 mmol, 3 equivalents) of diisopropylethylamine (DIPEA) was further added and reacted at room temperature for 1 hour. After the reaction was completed, the deoxycholic acid - peptide resin was filtered and washed, and then dried in a vacuum oven. Thereafter, 20 L of ether was added to the deoxycholic acid - peptide resin, crystallized, filtered, separated, and then purified to prepare the deoxycholic acid - peptide conjugate.

[0105] The molecular weight of the deoxycholic acid - peptide conjugate was measured with a molecular weight analyzer. As a result, it was confirmed that the molecular weight corresponded to 1022.4 Da. In addition, the result of 1 1H - NMR analysis of the prepared deoxycholic acid - peptide conjugate is shown in Figure 1 . In 1In the H-NMR analysis spectrum, in addition to the peaks of deoxycholic acid, the peaks of the peptide were also shown, clearly confirming the synthesis of the deoxycholic acid-peptide conjugate.

[0106] [Preparation Example 1]

[0107] Preparation of preadipocytes and induction of differentiation into adipocytes

[0108] The 3T3-L1 cells, which are preadipocytes, were purchased from the American Type Culture Collection (ATCC) and used in the experiments. 3T3-L1 is a cell line widely used to study the metabolic processes of adipocytes. When the differentiation of this cell is activated, lipid accumulation in adipocytes is activated, thus inducing obesity. Therefore, when this cell is treated with a substance considered to have an anti-obesity effect, the less the cell differentiation, the greater the anti-obesity effect of the substance can be seen.

[0109] The preadipocytes 3T3-L1 were cultured in DMEM containing 2% fetal bovine serum (FBS) under the conditions of 37 °C and 5% CO 2 . The preadipocytes 3T3-L1 were seeded at a density of 1 × 10 5 cells / well into a 12-well plate and then further maintained for 2 days at 100% confluency point. The preadipocytes were cultured for 2 days in a DMEM medium containing differentiation components (containing 10 μg / ml insulin, 0.1 μM dexamethasone and 0.5 mM IBMX) and 10% FBS (hereinafter referred to as "differentiation medium") to induce differentiation into adipocytes. After 48 hours of culture, the preadipocytes were cultured for 2 days in 10% FBS DMEM containing 1 μg / ml insulin (hereinafter referred to as "medium"). Thereafter, the medium was changed every 2 days with 10% FBS DMEM medium. During the process of inducing adipocyte differentiation, according to the experimental purpose, the preadipocytes were treated or not treated with the deoxycholic acid-peptide conjugate at a concentration of 1 μM or 10 μM in each medium, and at the end of differentiation, on the 7th day, the degree of adipocyte differentiation was observed.

[0110] [Preparation Example 2]

[0111] Collection and culture of adipose tissue

[0112] Adipose tissue was collected from the abdomen of 9-week-old BL / 6 mice, placed in a 100 mm × 15 mm culture dish, and then cultured overnight in the medium. The adipose tissue was divided into equal weights, transferred to a 24-well plate, and then used in the following experimental examples.

[0113] [Experimental Example 1]

[0114] Confirmation of the Solubility of Deoxycholic Acid-Peptide Conjugates

[0115] Visually observe and compare the solubilities of the deoxycholic acid-peptide conjugate and deoxycholic acid prepared in Example 2.

[0116] Specifically, add 90 μL of distilled water to E. tubes containing 1 mg each of the deoxycholic acid-peptide conjugate and deoxycholic acid, and confirm the dissolution status.

[0117] As a result, as Figure 2 shown, contrary to the fact that deoxycholic acid is almost insoluble in water, it was confirmed that the deoxycholic acid-peptide conjugate of the present invention is completely soluble in water.

[0118] From this result, it can be seen that in the deoxycholic acid-peptide conjugate of the present invention, the peptide is conjugated to deoxycholic acid, so that the water solubility is improved compared to the case where deoxycholic acid exists alone.

[0119] [Experimental Example 2]

[0120] Fat Accumulation Inhibitory Effect of Deoxycholic Acid-Peptide Conjugates

[0121] 2-1. Oil Red O Staining and Analysis

[0122] Perform Oil Red O staining to confirm whether the deoxycholic acid-peptide conjugate of the present invention has an effect of inhibiting fat accumulation in adipocytes.

[0123] Specifically, first confirm the degree of adipocyte differentiation induced in Preparation Example 1 by Oil Red O staining under a microscope, and for the staining degree of adipocytes, calculate the degree of fat accumulated in the cells based on the absorbance measured at a wavelength of 510 nm using a spectrophotometer. The groups treated with the β3 adrenergic receptor agonist CL316243 or TNF-α were used as positive control groups.

[0124] As a result, as shown in Figure 3A, when treated with 1 μM of the deoxycholic acid-peptide conjugate, the fat staining in adipocytes decreased to the same level as that of the positive control group.

[0125] From this, it can be seen that the deoxycholic acid-peptide conjugate can inhibit the differentiation of preadipocytes into adipocytes and inhibit lipogenesis, thereby inhibiting lipid accumulation in adipocytes.

[0126] 2-2. Confirmation of the Synergistic Inhibition Effect of Deoxycholic Acid-Peptide on Fat Accumulation

[0127] To compare the effect of inhibiting fat accumulation when treated with deoxycholic acid-peptide conjugate and when treated with deoxycholic acid or the peptide of SEQ ID NO:1 alone, the degree of adipocyte differentiation and the degree of lipid accumulation were measured in the same manner as in Experimental Example 1-1. The group treated with TNF-α was used as the positive control group.

[0128] As a result, as shown in Figure 3B, when treated with the peptide of SEQ ID NO:1 and deoxycholic acid at 1 μM each, the amounts of lipid accumulation were 89% and 93% respectively, but when treated with the deoxycholic acid-peptide conjugate at the same concentration, it was confirmed that the amount of lipid accumulation was 78%.

[0129] From this, it can be seen that compared with the case of treating with the peptide of SEQ ID NO:1 or deoxycholic acid alone, when treated with the deoxycholic acid-peptide conjugate at the same concentration, the degree of differentiation into adipocytes and the degree of inhibition of fat accumulation increased significantly. In addition, it can be seen that even when using a small amount of deoxycholic acid-peptide conjugate, an effect equal to or higher than that of a single compound can be exhibited.

[0130] [Experimental Example 3]

[0131] Lipolytic effect of deoxycholic acid-peptide conjugate

[0132] 3-1. Determination of the Amount of Lipolysis Product Glycerol in Adipose Tissue

[0133] Neutral fat stored in adipose tissue is decomposed into fatty acids and glycerol to generate energy or cell signaling. Therefore, by measuring the amount of free glycerol released, the lipolytic effect of neutral fat accumulated in adipose tissue was confirmed.

[0134] Specifically, fat pads that were white adipose tissue were isolated from female mice fed a high-fat diet (rodent diet with 60% Kcal fat, ResearchDiets, USA), and the adipose tissue was placed in a 12-well plate, with 400 mg of adipose tissue in each well, and then cultured in a medium (serum-free DMEM medium) for 48 hours to stabilize it. During the culture period, treatment was carried out with the peptide conjugate at concentrations of 1 μM, 10 μM, and 100 μM, and 20 nM of TNFα was used as the positive control group for treatment, and cultured for 48 hours to obtain the supernatant, and the amount of glycerol was measured using a glycerol colorimetric assay kit (Cayman chemical, USA).

[0135] As a result, as shown in Figure 4A, it was confirmed that compared with the control group, when treated with the deoxycholic acid-peptide conjugate, the amount of glycerol released during fat decomposition increased in a deoxycholic acid-peptide conjugate concentration-dependent manner.

[0136] As can be seen, the deoxycholic acid-peptide conjugate of the present invention has the activity of decomposing neutral fat accumulated in adipose tissue.

[0137] 3-2. Confirmation of the Reduction in Adipocyte Size in Adipose Tissue

[0138] To confirm the lipolytic effect of the deoxycholic acid-peptide conjugate on adipose tissue, the adipose tissue obtained in Preparation Example 2 was subjected to H&E staining to confirm the size of adipocytes.

[0139] Specifically, the adipose tissue was cut, attached to a paraffin slide, and dried at 50°C. The slides with the dried adipose tissue attached were placed in xylene 1, xylene 2, and xylene 3 for 5 minutes respectively, and the paraffin was removed. Thereafter, the slides were placed in 100% ethanol for 2 minutes, and then successively placed in 90%, 80%, and 70% ethanol for rehydration. The rehydrated slides were stained in hematoxylin solution, washed, and then stained with Eosin Y. Thereafter, the slides were successively placed in 90%, 80%, and 70% ethanol, then placed in xylene 3, xylene 2, and xylene 1, and then sealed with a xylene-based mounting medium to prevent air bubbles from entering the tissue, and then dried. Then, the dried slides were observed under an optical microscope.

[0140] As a result, as shown in Figure 4B, it was confirmed that the size of adipocytes decreased when the adipose tissue was treated with the deoxycholic acid-peptide conjugate compared to the control group.

[0141] As can be seen, the deoxycholic acid-peptide conjugate of the present invention decomposed the fat that had accumulated in the cells.

[0142] 3-3. Confirmation of the Synergistic Lipolysis Effect of Deoxycholic Acid-Peptide Conjugate

[0143] To compare the lipolytic effects in the case of treatment with the deoxycholic acid-peptide conjugate and in the case of treatment with deoxycholic acid or the peptide of SEQ ID NO: 1 alone, the amount of glycerol released from adipose tissue was measured in the same manner as in Experimental Example 3-1. The group treated with TNF-α was used as the positive control group.

[0144] As a result, as shown in Figure 4C, it was confirmed that when treated with the peptide of SEQ ID NO: 1 and deoxycholic acid at 1 μM each, the glycerol release amounts were 106% and 116% respectively, but when treated with the deoxycholic acid-peptide conjugate at the same concentration, the glycerol release amount was 175%.

[0145] It can be seen that, compared with the case of treating with the peptide of SEQ ID NO:1 alone or deoxycholic acid, the amount of glycerol released due to lipolysis increased significantly when treated with the deoxycholic acid-peptide conjugate at the same concentration. In addition, it can be seen that even when using a small amount of the deoxycholic acid-peptide conjugate, an effect equal to or higher than that of the single compound can be exhibited.

[0146] [Experimental Example 4]

[0147] Confirmation of changes in the expression of genes related to adipogenesis and lipolysis

[0148] 4-1. RT-PCT Analysis

[0149] The adipocytes obtained in Preparation Example 1 or the adipose tissue obtained in Preparation Example 2 were treated with the deoxycholic acid-peptide conjugate of the present invention, cultured, then recovered, and RNA was extracted using Trizol (Thermo Fisher Scientific, USA). Using TOPScript TM RT DryMIX (enzynomics, Korea) was used to obtain DNA complementary to the RNA from the extracted RNA, and then TOPsimple TM DryMIX-nTaq (enzynomics, Korea) was used for polymerase chain reaction (PCR). Thereafter, the PCR products were loaded on a 1.5% agarose gel to compare the mRNA expression levels of growth factors. The GAPDH gene was used as an internal control group, and the primers used in the experiment are shown in Table 1.

[0150] [Table 1]

[0151]

[0152]

[0153] 4-2. Confirmation of the Increase in Gene Expression Related to Fat Metabolism Using Adipocytes

[0154] In order to confirm whether the deoxycholic acid-peptide conjugate promotes fat metabolism, the mRNA expression levels of adiponectin and leptin genes related to fat metabolism were confirmed.

[0155] Specifically, the adipocytes obtained in Preparation Example 1 were treated with 1 μM and 10 μM of the deoxycholic acid-peptide conjugate of the present invention and cultured for 1 hour, 3 hours, and 6 hours. After recovering the cells and performing polymerase chain reaction by the method described in Experimental Example 4-1, the expression products were loaded on an agarose gel to confirm the mRNA expression levels of genes related to fat metabolism.

[0156] As a result, as Figure 5As shown, it was confirmed that the expression of proteins related to fat metabolism, namely leptin and adiponectin genes, increased by treatment with deoxycholic acid-peptide conjugates.

[0157] From this, it can be seen that the deoxycholic acid-peptide conjugate of the present invention promotes fat metabolism required for energy production, thereby inhibiting fat accumulation or promoting lipolysis.

[0158] 4-3. Confirmation of the Increase in Lipolysis-Related Gene Expression Using Adipocytes

[0159] In adipocytes, to determine whether the deoxycholic acid-peptide conjugate has an activating effect on the lipolytic cell signaling pathway, the mRNA expression levels of genes related to promoting fatty acid oxidation and promoting lipolysis were confirmed.

[0160] Specifically, the differentiated adipocytes obtained in Preparation Example 1 were treated with 1 μM and 10 μM of the deoxycholic acid-peptide conjugate and then cultured for 24 hours. After recovering the cells and performing polymerase chain reaction by the method described in Experimental Example 4-1, the expression products were loaded on an agarose gel to confirm the mRNA expression levels of genes related to the lipolytic cell signaling pathway.

[0161] As a result, as shown in Figure 6A, after treatment with the deoxycholic acid-peptide conjugate, the expression of the lipolysis marker HSL increased, and the expression of the upstream signaling factor AMPKa1 related to fat metabolism and its downstream factor PGC1a increased.

[0162] From this, it can be seen that the deoxycholic acid-peptide conjugate according to the present invention increases the gene expression related to promoting lipolysis or fatty acid oxidation, and thus can decompose fat very effectively.

[0163] 4-4. Confirmation of the Increase in Lipolysis Gene Expression Using Adipose Tissue

[0164] In adipose tissue, to determine whether the deoxycholic acid-peptide conjugate has an activating effect on the lipolytic cell signaling pathway, the mRNA expression levels of genes related to promoting fatty acid oxidation and promoting lipolysis were confirmed.

[0165] Specifically, the adipose tissue obtained in Preparation Example 2 was treated with 1 μM and 10 μM of the deoxycholic acid-peptide conjugate and 1 μM of CL316243 (Sigma, USA) as a positive control group, and cultured for 72 hours. After recovering the cells and performing polymerase chain reaction by the method described in Experimental Example 4-1, the expression products were loaded on an agarose gel to confirm the mRNA expression levels of genes related to promoting fatty acid oxidation and promoting lipolysis.

[0166] As a result, as shown in Figure 6B, after treatment with the deoxycholic acid-peptide conjugate, the expression of the fat-related genes HSL and PLIN increased in a concentration-dependent manner.

[0167] It can be seen therefrom that the deoxycholic acid-peptide conjugate according to the present invention increases the gene expression related to promoting lipolysis or fatty acid oxidation, thereby being able to decompose fat very effectively.

[0168] 4-5. Confirmation of the Decrease in Adipogenesis Gene Expression Using Adipose Tissue

[0169] In adipose tissue, in order to confirm whether the deoxycholic acid-peptide conjugate has an effect of inhibiting adipogenesis, the mRNA expression levels of genes related to adipogenesis were confirmed.

[0170] Specifically, the adipose tissue obtained in Preparation Example 2 was treated with 1 μM and 10 μM of the deoxycholic acid-peptide conjugate and 1 μM of CL316,243 (Sigma, USA) as a positive control group, and further cultured for 72 hours. After recovering the cells and performing polymerase chain reaction by the method described in Experimental Example 4-1, the expression products were loaded on an agarose gel to confirm the mRNA expression levels of genes related to adipogenesis.

[0171] As a result, as Figure 7 shown, it was confirmed that the expressions of adipogenesis-related genes ACCα, FAS, and PPARγ were decreased after treatment with the deoxycholic acid-peptide conjugate.

[0172] It can be seen therefrom that the deoxycholic acid-peptide conjugate according to the present invention has an effect of preventing or treating obesity by inhibiting adipogenesis.

[0173] [Experimental Example 5]

[0174] Confirmation that the deoxycholic acid-peptide conjugate increases the expression of genes related to thermogenesis in myocytes

[0175] In order to confirm whether the deoxycholic acid-peptide conjugate of the present invention has an effect of preventing or treating obesity, the mRNA expression levels of genes related to thermogenesis in brown adipocytes were confirmed.

[0176] Specifically, the adipose tissue obtained in Preparation Example 2 was treated with 1 μM and 10 μM of the deoxycholic acid-peptide conjugate and 1 μM of CL316,243 (Sigma, USA) as a positive control group, and further cultured for 72 hours. After recovering the cells and performing polymerase chain reaction by the method described in Experimental Example 4-1, the expression products were loaded on an agarose gel to confirm the mRNA expression levels of genes related to adipogenesis. At this time, the GAPDH gene was used as an internal control group, and the primers used in the experiment are shown in Table 2 below.

[0177] [Table 2]

[0178]

[0179] As a result, as Figure 8 shown, it was confirmed that after treatment with the deoxycholic acid-peptide conjugate, the expression of the thermogenesis-related genes UCP1, PRDM, and Cidea in brown adipocytes increased. From this, it can be seen that the deoxycholic acid-peptide conjugate according to the present invention increases the expression of thermogenesis-related genes in brown adipocytes whose expression is increased by exercise, and thus has the effect of preventing or treating obesity.

[0180] [Experimental Example 6]

[0181] Confirmation that the deoxycholic acid-peptide conjugate increases the expression of the IL-15 gene in myocytes

[0182] To confirm whether the deoxycholic acid-peptide conjugate of the present invention has the effect of preventing or treating obesity, the change in the expression of the IL-15 gene in myocytes was confirmed. IL-15 is a myokine whose expression level increases due to muscle contraction during resistance exercise. It is known that IL-15 plays a key role in the interaction between muscle and adipose tissue and has an anti-obesity effect by inhibiting lipid accumulation.

[0183] Specifically, mouse myoblast C2C12 cells were seeded into a 6-well plate at a density of 2×10 5 cells / well and then cultured in DMEM (10% FBS) medium for 3 days. When the cells grew to about 70% to 80% of the plate, the medium was replaced with differentiation medium (DMEM, 5% horse serum) and then cultured for 3 days. Thereafter, the medium was replaced with differentiation medium (DMEM, 2% horse serum) and cultured for 4 days to induce the differentiation of myoblasts into myotubes. Then, the medium was replaced with serum-free medium and cultured for 2 hours, and treated with 1 μM and 10 μM of the deoxycholic acid-peptide conjugate for 6 hours, and cultured in an environment of 37 °C and 5% CO 2 . After the cells were recovered, polymerase chain reaction was performed by the method described in Experimental Example 4-1, and the expression product was loaded on an agarose gel to confirm the mRNA expression level of genes related to adipogenesis. At this time, the GAPDH gene was used as an internal control group, and the primers used in the experiment are shown in Table 3 below.

[0184] [Table 3]

[0185]

[0186] As a result, as Figure 9 shown, it was confirmed that after treatment with the deoxycholic acid-peptide conjugate, the expression of IL-15 with adipose tissue lipolysis function increased.

[0187] As can be seen, the deoxycholic acid-peptide conjugate according to the present invention increases the expression of the IL-15 gene that is expressed by exercise and has a lipolytic function, thereby having an effect of preventing or treating obesity.

[0188] As described above, although the present invention has been described in detail with reference to the described embodiments, it will be apparent to those skilled in the art that various modifications and changes can be made within the technical idea of the present invention, and these modifications and changes naturally also belong to the appended claims. <110> Kelgern Co., Ltd. <120> Deoxycholic Acid-Peptide Conjugate Having Anti-Obesity Activity and Its Use <130> 2021-OPA-5319 <150> KR 10-2020-0071657 <151> 2020-06-12 <160> 31 <170> KoPatentIn 3.0 <210> 1 <211> 5 <212> PRT <213> Artificial Sequence <220> <223> Peptide <400> 1 Lys Phe Leu Ile Lys 1 5 <210> 2 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Forward Primer of Leptin <400> 2 ggatcaggtt ttgtggtgct 20 <210> 3 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Reverse Primer of Leptin <400> 3 ttgtggccca taaagtcctc 20 <210> 4 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Forward primer of adiponectin <400> 4 tcctgctttg gtccctccac 20 <210> 5 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Reverse primer of adiponectin <400> 5 tctccagccc cacactgaac 20 <210> 6 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Forward primer of AMPKα <400> 6 tcaccggaca taaagtggct 20 <210> 7 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Reverse primer of AMPKα <400> 7 tgatgatgtg agggtgcctg 20 <210> 8 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> Forward primer of HSL <400> 8 ggacacacac acacctg 17 <210> 9 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Reverse primer of HSL <400> 9 ccctttcgca gcaactttag 20 <210> 10 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Forward primer of PDE3b <400> 10 tgattgaccc aagccttccc 20 <210> 11 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Reverse primer of PDE3b <400> 11 tcttaccact gctgcgatcc 20 <210> 12 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Forward primer of PGCLa <400> 12 atgtgcagcc aagactctgt a 21 <210> 13 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Reverse primer of PGCLa <400> 13 cgctacacca cttcaatcca c 21 <210> 14 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Forward primer of PLIN <400> 14 aaggatcctg cacctcacac 20 <210> 15 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Reverse primer of PLIN <400> 15 cctctgctga agggttatcg 20 <210> 16 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> Forward primer of ACCα <400> 16 accttactgc catcccatgt gcta 24 <210> 17 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> Reverse primer of ACCα <400> 17 gtgcctgatg atcgcacgaa caaa 24 <210> 18 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Forward primer of FAS <400> 18 tgctggcact acagaatgc 19 <210> 19 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Reverse primer of FAS <400> 19 aacagcctca gagcgacaat 20 <210> 20 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Forward primer of PPARγ <400> 20 atgggtgaaa ctctgggaga 20 <210> 21 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Reverse primer of PPARγ <400> 21 gagctgattc cgaagttggt 20 <210> 22 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Forward primer of GAPDH <400> 22 gtgatggcat ggactgtggt 20 <210> 23 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Reverse primer of GAPDH <400> 23 ggagccaaaa gggtcatcat 20 <210> 24 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Forward primer of UCP1 <400> 24 cctgcctctc tcggaaacaa 20 <210> 25 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Reverse primer of UCP1 <400> 25 gtagcggggt ttatcccat 19 <210> 26 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Forward primer of PRDM <400> 26 ccccacattc cgctgtgat 19 <210> 27 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> Reverse primer of PRDM <400> 27 ctcgcaatcc ttgcactca 19 <210> 28 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Forward primer of Cidea <400> 28 cgggaatagc cagagtcacc 20 <210> 29 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Reverse primer of Cidea <400> 29 tgtgcatcgg atgtcgtagg 20 <210> 30 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Forward primer of IL-15 <400> 30 tgtcttcatt ttgggctgtg 20 <210> 31 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Reverse primer of IL-15 <400> 31 tgcaactggg atgaaagtca 20

Claims

1. A deoxycholic acid-peptide conjugate, wherein deoxycholic acid is conjugated to a peptide consisting of the amino acid sequence of SEQ ID NO: 1; in, The deoxycholic acid-peptide conjugate is represented by the following [Chemical Formula 2]: 2 . The deoxycholic acid-peptide conjugate according to claim 1 , wherein the deoxycholic acid-peptide conjugate inhibits lipogenesis in adipocytes. The deoxycholic acid-peptide conjugate according to claim 1 , wherein the deoxycholic acid-peptide conjugate promotes lipolysis. The deoxycholic acid-peptide conjugate according to claim 1 , wherein the deoxycholic acid-peptide conjugate reduces the size of adipocytes. 5 . A composition for inhibiting lipogenesis or promoting lipolysis, comprising the deoxycholic acid-peptide conjugate according to claim 1 . 6 . A composition for preventing, treating or improving obesity or obesity-related diseases, comprising the deoxycholic acid-peptide conjugate according to claim 1 . 7 . A pharmaceutical composition for preventing or treating obesity, comprising the deoxycholic acid-peptide conjugate according to claim 1 .

8. The pharmaceutical composition according to claim 7, further comprising a pharmaceutically acceptable carrier.

9. The pharmaceutical composition according to claim 7, further comprising an excipient.

10. The pharmaceutical composition according to claim 7, further comprising a diluent.

11. The pharmaceutical composition according to claim 8, wherein the carrier is at least one selected from the group consisting of physiological saline, sterile water, Ringer's solution, buffered physiological saline, glucose solution, maltodextrin solution, glycerol and ethanol.

12. The pharmaceutical composition according to claim 7, wherein the pharmaceutical composition is formulated in the form of an oral preparation, an injection preparation or an external preparation.

13. The pharmaceutical composition according to claim 12, wherein the injection preparation is used to remove local fat deposits.

14. The pharmaceutical composition according to claim 13, wherein the localized fat is located in a target area selected from the group consisting of abdomen, under the eyes, under the chin, under the arms, buttocks, thighs, calves, back, and bra line. 15 . A cosmetic composition for preventing or improving obesity, comprising the deoxycholic acid-peptide conjugate according to claim 1 . 16 . The cosmetic composition according to claim 15 , wherein the cosmetic composition has at least one dosage form selected from the group consisting of a cream, an emulsion, a gel, and a spray.

17. The cosmetic composition according to claim 15, wherein the dosage form of the cosmetic composition is lotion. The cosmetic composition according to claim 15 , wherein the dosage form of the cosmetic composition is skin softener.

19. The cosmetic composition according to claim 15, wherein the dosage form of the cosmetic composition is an essence.

20. The cosmetic composition according to claim 15, wherein the dosage form of the cosmetic composition is a hand cream.

21. The cosmetic composition according to claim 15, wherein the dosage form of the cosmetic composition is a cleansing foam.

22. The cosmetic composition according to claim 15, wherein the dosage form of the cosmetic composition is a cleansing cream.

23. The cosmetic composition according to claim 15, wherein the dosage form of the cosmetic composition is clean water.

24. The cosmetic composition according to claim 15, wherein the cosmetic composition is in the form of a shampoo.

25. The cosmetic composition according to claim 15, wherein the dosage form of the cosmetic composition is a hair dye.

26. The cosmetic composition according to claim 15, wherein the dosage form of the cosmetic composition is a care agent.

27. The cosmetic composition according to claim 15, wherein the cosmetic composition is in the form of a body cleanser.

28. The cosmetic composition according to claim 15, wherein the dosage form of the cosmetic composition is soap.

29. The cosmetic composition according to claim 15, wherein the dosage form of the cosmetic composition is a dressing.

30. The cosmetic composition according to claim 15, wherein the dosage form of the cosmetic composition is a massage agent.

31. The cosmetic composition according to claim 15, wherein the dosage form of the cosmetic composition is a face powder.

32. The cosmetic composition according to claim 15, wherein the dosage form of the cosmetic composition is a powder box.

33. The cosmetic composition according to claim 15, wherein the dosage form of the cosmetic composition is a foundation.

34. The cosmetic composition according to claim 15, wherein the dosage form of the cosmetic composition is a dual-purpose pressed powder.

35. The cosmetic composition according to claim 15, wherein the dosage form of the cosmetic composition is a primer.

36. Use of a deoxycholic acid-peptide conjugate in the manufacture of a medicament for inhibiting lipogenesis or promoting lipolysis; in, The deoxycholic acid-peptide conjugate is represented by the following [Chemical Formula 2]:

37. Use of a deoxycholic acid-peptide conjugate in the manufacture of a drug for preventing, treating or improving obesity; in, The deoxycholic acid-peptide conjugate is represented by the following [Chemical Formula 2]:

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