Composition for preventing or treating obesity or liver diseases, comprising thioredoxin-binding protein-derived peptide

Abstract

The present invention relates to a composition for preventing, treating, or alleviating obesity or liver diseases, comprising a thioredoxin-interacting protein (TXNIP)-derived peptide as an active ingredient. The TXNIP-derived peptide according to the present invention inhibits adipogenesis, exhibits an anti-obesity effect, and exhibits the effects of inhibiting metabolic dysfunction-associated fatty liver development and liver fibrosis, and thus can be used for the prevention or treatment of obesity or liver diseases.

Description

Composition for preventing or treating obesity or liver disease comprising a peptide derived from thioredoxin binding protein

[0001] The present invention relates to a composition for preventing, treating or improving obesity or liver disease, comprising a peptide derived from thioredoxin binding protein as an active ingredient.

[0002] Changes in modern eating habits and lifestyles are steadily increasing obesity rates. Obesity is the most common metabolic disease, resulting from a high-calorie diet and reduced physical activity. The average obesity rate across OECD countries is 58.7%, a very serious problem. The prevalence of obesity reached 13% of all adults in 2016 and is projected to reach 20% of adults worldwide by 2025. This increase in obesity leads to metabolic disorders, including chronic inflammation, fatty liver disease, and various immune dysfunctions. These disorders can lead to cardiovascular disease and diabetes, which are major complications of obesity. In severe cases, it can even lead to cancer and pose a life-threatening threat. Therefore, the development of treatments for obesity and metabolic fatty liver disease, which can cause these serious diseases, is urgently needed.

[0003] MAPKs (mitogen-activated protein kinases) are Ser / Thr kinases that transmit extracellular stimuli to the cell interior and are involved in regulating various cellular functions. Among them, p38 MAPK (p38) is strongly activated by various environmental stresses, including inflammatory cytokines and oxidative stress, and contributes to cellular homeostasis by regulating cell differentiation, proliferation, survival, and apoptosis. It also plays a crucial role in regulating transcription factors and cytokine expression levels in immune and inflammatory responses, with the p38α form being primarily involved in this process. Pharmacological inhibition of p38α is known to have positive effects on improving neurodegenerative diseases, heart failure-related diseases, obesity, and some metabolic diseases such as fatty liver, in addition to its anti-inflammatory effects. Regarding the association between obesity and p38, it was found that p38α knockout mice exhibited elevated body temperature and enhanced fat burning due to increased expression of Uncoupling Protein 1 (UCP1), a gene associated with thermogenesis. This increase in energy expenditure effectively suppresses diet-induced obesity. Furthermore, p38α is upregulated in hepatocytes from patients with metabolic fatty liver disease, suggesting that inhibition of p38α may improve this condition.

[0004] TN13 is a 13-amino acid peptide derived from a protein called thioredoxin-interacting protein (TXNIP). TXNIP is known to bind to TRX and regulate its activity, thereby regulating oxidative stress and contributing to the establishment of biological balance. Furthermore, thioredoxin-interacting protein (TXNIP) is a thioredoxin inhibitor that inhibits cell cycle progression and is therefore also known as a tumor suppressor. It also increases resistance to reactive oxygen species, making it a useful treatment for reactive oxygen-related diseases. However, the anti-obesity effects, inhibition of fatty liver formation, and therapeutic effects on liver disease of thioredoxin-interacting protein-derived peptides have not been disclosed.

[0005] Against this backdrop, the present invention was completed by confirming the anti-obesity effect, the inhibition of fatty liver formation, and the treatment effect of liver disease of a peptide derived from thioredoxin binding protein.

[0006] The present invention provides a pharmaceutical composition for preventing or treating obesity or liver disease, comprising a peptide derived from a thioredoxin binding protein having an amino acid sequence of sequence number 1.

[0007] In addition, the present invention provides a food composition for preventing or improving obesity or liver disease, comprising a peptide derived from a thioredoxin binding protein having an amino acid sequence of sequence number 1.

[0008] In addition, the present invention provides a method for preventing or treating obesity or liver disease, comprising administering to a subject in need thereof a composition comprising a peptide derived from a thioredoxin binding protein comprising an amino acid sequence of SEQ ID NO: 1.

[0009] In addition, the present invention seeks to provide a use of a peptide derived from a thioredoxin binding protein comprising an amino acid sequence of SEQ ID NO: 1 for the manufacture of a drug for preventing or treating obesity or liver disease.

[0010] In addition, the present invention provides a pharmaceutical composition comprising a peptide derived from a thioredoxin binding protein comprising an amino acid sequence of SEQ ID NO: 1 for use in the prevention or treatment of obesity or liver disease.

[0011] The terminology used in this application is merely used to describe specific embodiments and is not intended to limit the present invention. The singular expression includes the plural expression unless the context clearly indicates otherwise. In this application, the terms “comprise” or “have” should be understood to indicate the presence of a feature, step, structure, or combination thereof described above, but do not preclude the possibility of the presence or addition of one or more other features, steps, structures, or combinations thereof.

[0012] Unless otherwise defined, all terms used herein, including technical or scientific terms, have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. Terms defined in commonly used dictionaries should be interpreted as having a meaning consistent with their meaning in the context of the relevant technology, and will not be interpreted in an idealized or overly formal sense unless explicitly defined herein.

[0013] In addition, terms and abbreviations used in this specification have their original meanings unless otherwise defined.

[0014] In order to solve the above problem, the inventor of the present invention discovered that the TN13 peptide derived from thioredoxin binding protein has a therapeutic effect on obesity and metabolic liver disease, thereby completing the present invention.

[0015] Hereinafter, a pharmaceutical composition and a food composition for preventing or treating obesity or liver disease, each comprising a peptide derived from a thioredoxin binding protein of the present invention, and a method for preventing or treating obesity or liver disease using the same are described in detail.

[0016] Pharmaceutical composition and treatment method for preventing or treating obesity or liver disease comprising a peptide derived from a thioredoxin binding protein according to the present invention

[0017] In one aspect of the present invention, a pharmaceutical composition for preventing or treating obesity or liver disease is provided, comprising a peptide derived from a thioredoxin binding protein having an amino acid sequence of sequence number 1.

[0018] A thioredoxin-binding protein-derived peptide comprising the amino acid sequence of SEQ ID NO: 1 may be, specifically, a thioredoxin-binding protein-derived peptide consisting essentially of the amino acid sequence of SEQ ID NO: 1. Specifically, it may be a thioredoxin-binding protein-derived peptide consisting of the amino acid sequence of SEQ ID NO: 1.

[0019] It was confirmed that the peptide derived from thioredoxin binding protein (TXNIP) according to the present invention directly binds to p38, which induces oxidative stress, and inhibits its activity. A short peptide having a 13 amino acid sequence derived from TXNIP that binds to the docking motif of p38 was prepared and named TN13. p38 is composed of four isomers with similar base sequences, p38α, p38β, p38γ, and p38δ, and TN13 specifically binds to the docking motif of p38α among them and acts. To enable TN13 to efficiently penetrate cells, a cell-penetrating peptide (CPP) derived from the human immunodeficiency virus (HIV) trans-activator protein (TAT) sequence was conjugated to TN13, thereby preparing TAT-TN13. TN13 has a high level of safety compared to existing chemical p38 inhibitors. While existing inhibitors require dissolution in organic solvents that are toxic to cells, TN13 is highly soluble in water, so it has no solvent-induced toxicity during processing, and exhibits high absorption. Furthermore, unlike existing inhibitors that simultaneously inhibit both p38α and p38β isoforms, TN13 selectively inhibits only p38α with a short amino acid sequence, and is therefore expected to have fewer side effects. While chemical drugs have a high possibility of causing side effects due to the uncertain persistence period in the body, peptide drugs have a relatively low possibility of causing side effects due to their short persistence period in the body. The thioredoxin-binding protein-derived peptide according to the present invention has anti-inflammatory effects, an effect of inhibiting lipogenesis in liver cancer cell lines, an anti-obesity effect, an effect of inhibiting metabolic fatty liver formation, and other minor agonistic effects. These agonistic effects enable the prevention and / or treatment of liver diseases such as obesity or fatty liver disease, especially metabolic fatty liver disease.

[0020] A peptide derived from a thioredoxin binding protein comprising the amino acid sequence of sequence number 1 of the present invention may be composed of a sequence in which one or several amino acids of the protein are added, deleted, or substituted, as long as it has the same activity as the protein or has the same gene location encoding the thioredoxin binding protein on the chromosome.

[0021] The above thioredoxin binding protein-derived peptide is composed of a sequence having at least 80% homology, more specifically at least 90% homology, and most specifically at least 95%, 96%, 97%, 98%, 99% or 99.5% homology to the amino acid sequence of SEQ ID NO: 1, but is not limited thereto.

[0022] For example, a peptide derived from a thioredoxin binding protein comprising an amino acid sequence of SEQ ID NO: 1 according to the present invention may have an amino acid sequence of 30 or less and may include an amino acid sequence of SEQ ID NO: 1. This means that the number of consecutive amino acid sequences is at most 50. Preferably, it means having an amino acid sequence of 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30. More preferably, it may include or consist of a sequence of 7 to 30 consecutive amino acids, or 10 to 15 consecutive amino acids.

[0023] For example, the amino acid sequence of sequence number 1 may further include at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, or 17 amino acids at the N-terminus and / or C-terminus of the Core sequence.

[0024] Additionally, the thioredoxin binding protein-derived peptide can be fused with a cell-penetrating peptide as needed.

[0025] A cell-penetrating peptide is a type of signal peptide, a peptide sequence intended to transport substances into cells. It typically consists of a peptide sequence of 7-30 amino acids. Any sequence that has a signal peptide capable of being delivered into cells may be included in the present invention.

[0026] For example, cell-penetrating peptides mainly contain basic amino acid residues such as lysine / arginine, and thus play a role in allowing proteins fused with them to penetrate the cell membrane and enter the cell. The cell-penetrating peptides include, but are not limited to, HIV-1 Tat protein, the homeodomain (penetratin) of Drosophila Antennapedia, HSV VP22 transcriptional regulatory protein, MTS peptide derived from vFGF, PTD-5, transportan, or sequences derived from Pep-1 peptide, etc. In this way, various CPPs (e.g., TAT48-60, penetratin (pAntp)43-58, polyarginine, Pep-1, transportan, etc.) identified / synthesized from viruses or cationic peptides have the activity of internalizing cells to mediate the movement of biologically active substances and drug carriers.

[0027] These exemplary sequences are shown in Table 1 below:

[0028] HIV-1 Tat (SEQ ID NO: 2)YGRKKRRQRRRPenetratin (SEQ ID NO: 3)RQIKIWFQNRRMKWKKHSV VP22 (SEQ ID NO: 4)DAATATRGRSAASRPTERPRAPARSASRPRRPVEMTS (SEQ ID NO: 5)AAVALLPAVLLAAPTransportan (SEQ ID NO: 6)GWTLNSAGYLLGKINLKALAALAKKILPEP-1 (SEQ ID NO: 7)KETWWETWWTEWSQPKKKRKVPoly arginine (SEQ ID NO: 8)RRRRRRRPTD-5 (SEQ ID NO: 9)RRQRRTSKLMKR

[0029] That is, the peptide derived from the thioredoxin binding protein may be linked to any one cell-penetrating peptide selected from the group consisting of SEQ ID NOs: 2 to 9. More specifically, the cell-penetrating peptide may be an amino acid sequence of SEQ ID NO: 2, and the TN13 peptide according to the present invention may be fused with the cell-penetrating peptide to form a fusion peptide.

[0030] In a preferred embodiment, the cell penetrating peptide may be a TAT peptide, and is preferably bound to the terminus of a peptide comprising the amino acid sequence of SEQ ID NO: 1, wherein the terminus may be either the amino terminus (5' terminus, N-terminus) or the carboxy terminus (3' terminus, C-terminus), but the amino terminus (5' terminus, N-terminus) is most preferred.

[0031] The above peptide may be further modified by phosphorylation, acetylation, methylation, glycosylation, etc., and may bind to other proteins. However, as long as the protein is not altered to the extent that its function is lost, it can be considered to be the same as the protein before modification.

[0032] The peptide derived from the thioredoxin binding protein of the present invention may have a cell-penetrating peptide linked to its N-terminus. More specifically, a TAT peptide described in SEQ ID NO: 2 is preferably linked, but is not limited thereto.

[0033] The connection between the above cell-penetrating peptide and the TN13 peptide may be direct or may be connected via a linker or spacer peptide.

[0034] The term "linker" or "spacer" refers to a short amino acid sequence used to separate two functionally different peptides in the construction of a fusion protein. The absence of a linker between two or more individual domains in a protein can result in reduced or inappropriate function of the protein domains due to steric hindrance, such as reduced catalytic activity or binding affinity for receptors / ligands. Using an artificial linker to connect protein domains in a chimeric protein can increase the space between the domains. Preferably, the linker or spacer is not particularly limited to the following, as long as it exhibits the effect of enhancing the activity of the conjugate of a cell-penetrating peptide and a vesicle-targeting peptide. While it may not have a specific biological activity other than to join the domains or to preserve some minimal distance or other spatial relationship between them, the constituent amino acids may be selected to influence some property of the molecule, such as folding, net charge, or hydrophobicity.

[0035] For example, it is preferred that the linker include a sequence recognized by a signal peptidase enzyme and be cleaved through this. That is, a sequence recognizable by a signal peptidase enzyme can be added between the cell-penetrating peptide and the TN13 peptide sequence.

[0036] In the present invention, the term “signal peptidase cleavage peptide” refers to a linker sequence that can be recognized by a signal peptidase and provide cleavage between a TN13 peptide and a cell-penetrating peptide sequence. The “signal peptidase cleavage peptide” can recognize the C-terminus of the cell-penetrating peptide sequence and the above cleavage peptide sequence and provide peptide cleavage between them. Since the cell-penetrating peptide sequence is derived from the signal sequence, it can have “AXA” or “VXA” (wherein X is any amino acid sequence) at the C-terminus. When an “EA” sequence connected thereto is added as a linker sequence, the signal peptidase enzyme can recognize the cleavage site between the above “AXA” or “VXA” and “EA” sequences and cleave them. Through this, the target TN13 can be delivered well into cells, thereby enhancing drug action.

[0037] The above liver disease may be any one selected from the group consisting of hepatic fibrosis, cholesterolosis, cirrhosis, viral and alcoholic hepatitis, Wilson's disease, hemochromatosis, steatosis, metabolic fatty liver disease (MASLD) and metabolic associated steatohepatitis (MASH).

[0038] In the present invention, “metabolic dysfunction-associated steatotic liver disease (MASLD)” is a disease in which fat accumulates in liver tissue regardless of significant alcohol consumption. Metabolic dysfunction-associated steatotic liver disease is divided into primary and secondary depending on the cause. Primary is caused by hyperlipidemia, diabetes, or obesity, which are characteristics of metabolic syndrome, and secondary is caused by nutritional causes (rapid weight loss, starvation, intestinal bypass surgery, etc.), various drugs, toxic substances (poisonous mushrooms, bacterial toxins, etc.), metabolic causes, and other factors. In the present invention, metabolic dysfunction-associated steatotic liver disease includes both primary and secondary metabolic dysfunction fatty liver diseases.

[0039] In the present invention, "simple fatty liver" is also called simple hepatic steatosis, and refers to a state of steatosis in which fat accumulates more than 5% of the normal liver fat. Simple fatty liver is not accompanied by inflammation and thus does not cause liver cell damage, and is therefore distinct from steatohepatitis, which is accompanied by inflammation.

[0040] In the present invention, “metabolic dysfunction-associated steatohepatitis (MASH)” is an advanced type of metabolic fatty liver disease, which means a state in which inflammation of hepatocytes is induced by fatty liver, and in people who do not consume significant alcohol, it shows tissue changes such as inflammation and necrosis, which are histologically similar to alcoholic liver disease. Metabolic dysfunction-associated steatohepatitis is accompanied by abnormal fat accumulation or deposition (steatosis) in the liver, hepatocyte inflammation and necrosis, and fibrosis due to tissue damage, and because it is a progressive disease, it can develop from fatty liver to steatohepatitis, liver fibrosis, cirrhosis, and even liver cancer.

[0041] In the present invention, "liver fibrosis," also known as hepatic fibrosis, refers to a condition in which extracellular matrix proteins, including collagen, accumulate excessively in liver tissue. Liver fibrosis is a fibrotic phenomenon that occurs due to the persistent destruction of hepatocytes and damage to liver tissue caused by various causes. It is caused by an imbalance between increased production of extracellular matrix and relatively decreased degradation. If liver fibrosis persists, it can develop into cirrhosis.

[0042] In the present invention, "liver cirrhosis" also refers to liver cirrhosis, which occurs when normal liver tissue is replaced by fibrous tissue, such as regenerative nodules, due to chronic inflammation, preventing the liver from properly performing its original function, resulting in a decline in liver function. While symptoms are not apparent in the early stages of cirrhosis, if liver damage progresses significantly, complications such as jaundice, ascites, hepatic encephalopathy, and variceal bleeding may occur.

[0043] In the present invention, the liver disease may be any one selected from the group consisting of, for example, hepatic fibrosis, cirrhosis, metabolic fatty liver disease (MASLD), and metabolically associated steatohepatitis (MASH).

[0044] The term “prevention” as used in the present invention means any act of suppressing or delaying the onset of obesity or liver disease by administering a composition.

[0045] In the present invention, “treatment” means any act in which the symptoms of obesity or liver disease are improved or beneficially changed by administration of the composition.

[0046] The pharmaceutical composition of the present invention may further comprise one or more pharmaceutically acceptable carriers in addition to the peptide derived from thioredoxin binding protein having the amino acid sequence of SEQ ID NO: 1 as an active ingredient for administration. Pharmaceutically acceptable carriers may include saline solution, sterile water, Ringer's solution, buffered saline, dextrose solution, maltodextrin solution, glycerol, ethanol, and mixtures of one or more of these components. If necessary, other conventional additives such as antioxidants, buffers, and bacteriostatic agents may be added. In addition, diluents, dispersants, surfactants, binders, and lubricants may be additionally added to formulate the composition into an injectable formulation such as an aqueous solution, suspension, or emulsion, or into a pill, capsule, granule, or tablet. Accordingly, the pharmaceutical composition of the present invention may be a patch, a liquid, a pill, a capsule, a granule, a tablet, a suppository, or the like. These preparations can be prepared by conventional methods used in formulation in the art or by methods disclosed in Remington's Pharmaceutical Science (latest edition), Mack Publishing Company, Easton PA, and can be formulated into various preparations depending on each disease or ingredient.

[0047] The composition of the present invention is administered in a pharmaceutically effective amount. The term "pharmaceutically effective amount" refers to an amount sufficient to treat a disease at a reasonable benefit / risk ratio applicable to medical treatment and not causing side effects. The effective dosage level may be determined based on factors including the patient's health condition, the type and severity of the disease, the activity and sensitivity of the drug, the method of administration, the time of administration, the route and excretion rate, the duration of treatment, drugs used in combination or simultaneously, and other factors well known in the medical field. The composition of the present invention may be administered as an individual therapeutic agent or in combination with other therapeutic agents, may be administered sequentially or simultaneously with conventional therapeutic agents, and may be administered singly or in multiple doses. It is important to administer an amount that achieves the maximum effect with the minimum amount without side effects by taking all of the above factors into consideration, and this can be easily determined by those skilled in the art. The daily dosage of the composition according to the present invention is 0.0001 to 10 mg / ml, preferably 0.0001 to 5 mg / ml, and it is more preferable to administer it once or several times a day.

[0048] The term "administration" in the present invention means introducing a predetermined substance into a patient by an appropriate method, and the administration route of the composition may be administered through any common route as long as it can reach the target tissue. In addition, the pharmaceutical composition of the present invention may be administered by any device that allows the active substance to move to the target tissue. For example, it may be administered by transdermal administration, oral administration, intrathecal administration, intraperitoneal administration, intravenous administration, intramuscular administration, subcutaneous administration, intradermal administration, topical administration, intranasal administration, intrapulmonary administration, rectal administration, inner ear administration, intrauterine epidural administration, sublingual administration, and intracerebrovascular injection, but is not limited thereto.

[0049] In the present invention, the pharmaceutical composition may appropriately include a suspending agent, a solubilizing agent, a stabilizer, an isotonic agent, a preservative, an adsorption inhibitor, a surfactant, a diluent, an excipient, a pH adjuster, a soothing agent, a buffer, a reducing agent, an antioxidant, etc., if necessary depending on the administration method or formulation. Pharmaceutically acceptable carriers and formulations suitable for the present invention, including those exemplified above, are described in detail in the literature [Remington's Pharmaceutical Sciences, 19th ed., 1995]. The pharmaceutical composition may be manufactured in a unit dose form or may be manufactured by inserting it into a multi-dose container by formulating it using a pharmaceutically acceptable carrier and / or excipient according to a method that can be easily performed by a person skilled in the art to which the present invention pertains. At this time, the formulation may be in the form of a solution, suspension, or emulsion in an oil or aqueous medium, or in the form of a powder, granules, tablets, or capsules.

[0050] Solid preparations for oral administration include tablets, pills, powders, granules, capsules, etc., and these solid preparations are formulated by mixing at least one excipient, such as starch, calcium carbonate, sucrose, lactose, gelatin, etc., with the above composition. In addition to simple excipients, lubricants such as magnesium stearate and talc may be used.

[0051] Oral liquid preparations include suspensions, solutions, emulsions, and syrups, and may include various excipients, such as wetting agents, sweeteners, flavoring agents, and preservatives, in addition to commonly used simple diluents such as water and liquid paraffin.

[0052] Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solutions, suspensions, emulsions, lyophilized preparations, and suppositories. Non-aqueous solutions and suspensions can include propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate. Suppository bases include withepsol, macrogol, Tween 61, cacao butter, laurin, and glycerogelatin. Injections can also include conventional additives such as solubilizers, isotonic agents, suspending agents, emulsifiers, stabilizers, and preservatives.

[0053] The route of administration of the pharmaceutical composition of the present invention may be through any general route as long as it can reach the target tissue, but may be through subcutaneous injection using an osmotic pump, intradermal injection, intravein injection, intraperitoneal injection, or intravitreal injection.

[0054] The composition of the present invention may additionally contain one or more active ingredients exhibiting the same or similar function. The composition of the present invention contains the protein in an amount of 0.0001 to 10 wt%, preferably 0.001 to 1 wt%, based on the total weight of the composition.

[0055] In another aspect, the present invention also provides a method for preventing or treating obesity or liver disease, comprising administering to a subject in need thereof a composition comprising a peptide derived from a thioredoxin binding protein comprising the amino acid sequence of SEQ ID NO: 1.

[0056] In the above treatment method, “thioredoxin binding protein-derived peptide”, “obesity”, “liver disease”, “prevention”, “treatment” and “composition” are as described above.

[0057] The term "subject" of the present invention refers to any animal that has developed or may develop obesity or liver disease, and may typically be an animal that can show a beneficial effect by treatment with a thioredoxin binding protein-derived peptide comprising the amino acid sequence of SEQ ID NO: 1 of the present invention, but includes without limitation any subject that has symptoms of obesity or liver disease or is likely to have such symptoms. As described above, by administering the pharmaceutical composition of the present invention to a subject, the obesity or liver disease can be effectively prevented or treated. The pharmaceutical composition of the present invention can be administered as an individual therapeutic agent, or can be administered in combination with existing obesity or liver disease therapeutic agents, and can be administered sequentially or simultaneously with the existing therapeutic agents.

[0058] The term "administration" in the present invention means introducing a predetermined substance into a patient by an appropriate method, and the route of administration of the composition may be administered through any common route as long as it can reach the target tissue. In addition, the pharmaceutical composition of the present invention may be administered by any device that allows the active substance to move to the target tissue. For example, it may be administered by oral administration, intrathecal administration, intraperitoneal administration, intravenous administration, intramuscular administration, subcutaneous administration, intradermal administration, topical administration, intranasal administration, intrapulmonary administration, rectal administration, inner ear administration, intrauterine epidural administration, sublingual administration, and intracerebrovascular injection, but is not limited thereto. Solid preparations for oral administration include tablets, pills, powders, granules, capsules, etc., and preparations for parenteral administration include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilized preparations, and suppositories.

[0059] The pharmaceutical composition of the present invention may be administered as an individual therapeutic agent or in combination with another therapeutic agent, and may be administered sequentially or simultaneously with conventional therapeutic agents. Taking all of the above factors into account, the pharmaceutical composition may be administered in an amount that achieves maximum efficacy with minimal side effects, as can be readily determined by those skilled in the art.

[0060] The present invention also provides a use of a peptide derived from a thioredoxin binding protein comprising the amino acid sequence of SEQ ID NO: 1 for the manufacture of a medicament for the prevention or treatment of obesity or liver disease.

[0061] The peptide derived from a thioredoxin binding protein containing the amino acid sequence of the above-mentioned sequence number 1 for the manufacture of a pharmaceutical agent can be mixed with acceptable adjuvants, diluents, carriers, etc., and can be manufactured into a complex preparation together with other active agents to have a synergistic effect of the active ingredients.

[0062] The matters mentioned in the uses, compositions and methods of the present invention apply equally unless they are contradictory to each other.

[0063] Food composition for preventing or improving obesity or liver disease comprising a peptide derived from a thioredoxin binding protein according to the present invention

[0064] In another aspect, a food composition for preventing or improving obesity or liver disease is provided, comprising a peptide derived from a thioredoxin binding protein having an amino acid sequence of sequence number 1.

[0065] In the above food composition, “thioredoxin binding protein-derived peptide”, “obesity”, “fatty liver disease”, “prevention”, “treatment” and “composition” are as described above.

[0066] In addition to the active ingredient, the above composition may contain a food additive acceptable from a food science perspective.

[0067] The term "food supplement additive" used in the present invention refers to a component that can be added to food as an auxiliary, and can be appropriately selected and used by those skilled in the art as added in the manufacture of health functional foods of each formulation. Examples of food supplement additives include various nutrients, vitamins, minerals (electrolytes), flavoring agents such as synthetic flavoring agents and natural flavoring agents, coloring agents and fillers, pectic acid and its salts, alginic acid and its salts, organic acids, protective colloid thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohol, carbonating agents used in carbonated beverages, etc., but the types of food supplement additives of the present invention are not limited by the above examples.

[0068] The food composition of the present invention may include a health functional food composition. The term "health functional food" as used herein refers to a food manufactured and processed in the form of tablets, capsules, powders, granules, liquids, pills, etc. using raw materials or ingredients that have functionality useful to the human body. Here, "functionality" means obtaining a beneficial effect for health purposes, such as regulating nutrients for the structure and function of the human body or physiological functions. The health functional food of the present invention can be manufactured by a method commonly used in the art, and during the manufacturing process, raw materials and ingredients commonly added in the art can be added. In addition, the formulation of the health functional food can be manufactured without limitation as long as it is a formulation recognized as a health functional food. The food composition of the present invention can be manufactured in various forms, and unlike general drugs, it has the advantage of not having side effects that may occur with long-term administration of drugs using food as a raw material, and is highly portable, so the health functional food of the present invention can be consumed as a supplement to enhance the anti-obesity effect or the effect of preventing or improving fatty liver disease.

[0069] In addition, the health functional food composition is preferably manufactured in any one dosage form selected from powder, granules, pills, tablets, capsules, candy, syrup, and beverage, but is not limited thereto. The health functional food composition of the present invention can be manufactured by adding the active ingredient as it is or mixing it with other foods or food ingredients, and can be manufactured appropriately according to a conventional method. Examples of foods to which the mixed extract of the root and tangerine peel can be added include dairy products including caramel, meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gum, ice cream, various soups, beverages, tea, drinks, alcoholic beverages, and vitamin complexes, and include all health functional foods in the conventional sense. That is, there is no particular limitation on the type of the food. The above health functional food composition may contain various nutrients, vitamins, minerals (electrolytes), synthetic and natural flavorings, colorings and enhancers (cheese, chocolate, etc.), pectic acid and its salts, alginic acid and its salts, organic acids, protective colloid thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohol, carbonating agents used in carbonated beverages, etc. In addition, it may contain fruit pulp for producing natural fruit juice and vegetable beverages. The above ingredients may be used independently or in combination. In addition, the health functional food composition of the present invention may contain various flavorings or natural carbohydrates as additional ingredients, and the natural carbohydrates are monosaccharides such as glucose and fructose, disaccharides such as maltose and sucrose, polysaccharides such as dextrin and cyclodextrin, and sugar alcohols such as xylitol, sorbitol and erythritol. Sweeteners that can be used include natural sweeteners such as thaumatin and stevia extract, and synthetic sweeteners such as saccharin and aspartame.

[0070] The peptide derived from thioredoxin-interacting protein (TXNIP) according to the present invention inhibits fat production, exhibits an anti-obesity effect, and has the effect of inhibiting fatty liver production and liver fibrosis related to metabolic disorders, and thus can be used for the prevention or treatment of obesity or liver disease.

[0071] Figure 1 shows the results of a CCK-8 assay on the viability of RAW264.7 cells treated with a peptide (TN13) derived from thioredoxin binding protein (TNXIP) according to the present invention (A), a Western blotting analysis on the inhibitory effect on p38 activity (B), and an optical microscopic analysis of morphological changes induced by LPS (C) (statistical analysis was performed using an unpaired two-tailed t-test. *p < 0.05, **p < 0.01, and ***p < 0.001).

[0072] Figure 2 shows the results of Western blotting analysis on the inhibitory effect of TN13 of the present invention on the p38-NFκB signal pathway according to palmitic acid treatment in RAW264.7 cells (statistical analysis was performed using an unpaired two-tailed t-test. *p < 0.05, **p < 0.01, and ***p < 0.001).

[0073] Figure 3 shows the results of CCK-8 analysis on the viability of HepG2 cells treated with TN13 according to the present invention (A), flow cytometry analysis (B), and optical microscopy analysis of cell permeability of FITC-TAT-TN13 (C).

[0074] Figure 4 shows the results of analyzing the time-dependent activation of p38 induced by palmitic acid (PA) using Western blotting (A), the results of analyzing the inhibitory effect of TN13 of the present invention on p38 MAPK using Western blotting (B), and the results of evaluating adipogenesis induced by palmitic acid in HepG2 cells using Oil Red O staining (C).

[0075] Figure 5 is a schematic diagram showing the timeline of high-fat diet (HFD) and TN13 treatment in a mouse model experiment.

[0076] Figure 6 shows the results of body weight measurements (A, B) and photographs of the mice (C) for 15 weeks in an in vivo experiment using a mouse model, respectively, of the normal diet group (ND), high-fat diet group (HFD), and TN13-administered high-fat diet group (HFD+TN13).

[0077] Figure 7 shows the results of blood glucose level analysis of the normal diet group (ND), high-fat diet group (HFD), and TN13-administered high-fat diet group (HFD+TN13) in an in vivo experiment using a mouse model.

[0078] Figure 8 shows the results of measuring the mass of eWAT (epididymal white adipose tissue) in the normal diet group (ND), high-fat diet group (HFD), and TN13-administered high-fat diet group (HFD+TN13) in an in vivo experiment using a mouse model (A), the results of photographing the appearance of eWAT (B), and the results of analyzing the size of fat cells in eWAT using H&E staining (C).

[0079] Figure 9 shows the results of an analysis of the inflammatory response in the body due to a high-fat diet in a normal diet group (ND), a high-fat diet group (HFD), and a high-fat diet group administered TN13 (HFD+TN13) in an in vivo experiment using a mouse model, including the results of a CBC analysis on inflammatory immune cells (A, B, C) and the results of measuring the level of pro-inflammatory cytokines in serum by ELISA analysis (D, E, F).

[0080] Figure 10 shows the results of imaging the liver of each group of the normal diet group (ND), high-fat diet group (HFD), and high-fat diet group administered TN13 (HFD+TN13) in an in vivo experiment using a mouse model (A), the results of measuring liver mass (B), the results of measuring TG levels in the liver to analyze fatty liver (C), the results of liver tissue examination through H&E staining and Oil Red O staining (D), and the results of evaluating protein levels related to lipogenesis and p38 by Western blotting (E) (Statistical analysis was performed using an unpaired two-tailed t-test. *p < 0.05, **p < 0.01, and ***p < 0.001).

[0081] Figure 11 shows the results of measuring the levels of proinflammatory cytokines in the livers of each group of the normal diet group (ND), high-fat diet group (HFD), and TN13-administered high-fat diet group (HFD+TN13) using an ELISA assay in an in vivo experiment using a mouse model (statistical analysis was performed using an unpaired two-tailed t-test. *p < 0.05, **p < 0.01, and ***p < 0.001).

[0082] Figure 12 shows the results of IHC staining analysis (D) and cell counting (E, F) of macrophages and neutrophils in the liver in each group of the normal diet group (ND), high-fat diet group (HFD), and TN13-administered high-fat diet group (HFD+TN13) in an in vivo experiment using a mouse model (statistical analysis was performed using an unpaired two-tailed t-test. *p < 0.05, **p < 0.01, and ***p < 0.001).

[0083] Figure 13 shows an image confirming liver fibrosis using Sirius Red staining (A), and the results of measuring the levels of fibrosis-activating cytokines in each group of the normal diet group (ND), high-fat diet group (HFD), and TN13-administered high-fat diet group (HFD+TN13) in an in vivo experiment using a mouse model using ELISA analysis (B), and the results of evaluating the levels of proteins related to fibrosis using Western blotting (C) (statistical analysis was performed using an unpaired two-tailed t-test. *p < 0.05, **p < 0.01, and ***p < 0.001).

[0084] Figure 14 shows the results of confirming liver damage by measuring ALT and AST levels in each group of the normal diet group (ND), high-fat diet group (HFD), and TN13-administered high-fat diet group (HFD+TN13) in an in vivo experiment using a mouse model (statistical analysis was performed using an unpaired two-tailed t-test. *p < 0.05, **p < 0.01, and ***p < 0.001).

[0085] Hereinafter, it will be described in detail by examples.

[0086] However, the following examples are only illustrative of the present invention, and the content of the present invention is not limited to the following examples.

[0087] Example 1. Preparation of peptide derived from thioredoxin binding protein (TNXIP)

[0088] Example 1-1. Preparation of TN13 peptide

[0089] TN13, a domain fragment of TXNIP containing the amino acid sequence of sequence number 1, was produced. The sequence number 1 is as disclosed in Table 1 below.

[0090] Example 1-2. Preparation of TAT-TN13 peptide (TN13)

[0091] TN13, a domain fragment of TXNIP containing the amino acid sequence of SEQ ID NO: 1 according to Example 1-1, was produced, and the HIV TAT transduction domain sequence (SEQ ID NO: 2) was conjugated to the N-terminus of TN13 to produce a TAT-TN13 peptide. SEQ ID NOs: 1 and 2 are as disclosed in Table 2 below.

[0092] Name Sequence Sequence Number TN13GSKKVILDLPLVI Sequence Number 1 Gly Ser Lys Lys Val Ile Leu Asp Leu Pro Leu Val Ile HIV TAT domain (TAT) YGRKKRRQRRR Sequence Number 2 Tyr Gly Arg Lys Lys Arg Arg Gln Arg Arg Arg

[0093] Example 1-3. Preparation of Fitc(Fluorescein isothiocyanate)-TAT-TN13 peptide

[0094] A peptide composed of Fitc (Fluorescein isothiocyanate)-TAT-TN13, which is a combination of the TAT-TN13 peptide (TN13) manufactured according to Example 1-2 and the fluorescent dye FITC, was synthesized by requesting synthesis from Peptron Co., Ltd. (Daejeon, Korea).

[0095] Example 1-4. Test Preparation

[0096] (1) Cell culture

[0097] The mouse macrophage cell line RAW264.7 was cultured in RPMI (Welgene Inc., Gyeongsan, Korea) containing 10% FBS (R&D Systems, Minneapolis, MN, USA). The human hepatoma cell line HepG2 was cultured in DMEM (Welgene Inc., Gyeongsan, Korea) containing 10% FBS, and all cells were cultured at 37°C in an atmosphere of 5% CO2.

[0098] (2) Cell viability analysis

[0099] Cell viability was measured using the Cell Counting Kit (CCK)-8 (Dojindo Laboratories, Kumamoto, Japan). 2x10 RAW264.7 cells or HepG2 cells were seeded in a 96-well plate. 4 Cells were dispensed per well and stabilized in a 5% CO2 incubator at 37°C, after which TN13 was treated at various concentrations. After 24 hours, CCK-8 solution (10 μl per well) was treated and additionally incubated for 30 minutes to 1 hour at 37°C and 5% CO2, and the absorbance at 450 nm was measured using a microplate reader.

[0100] (3) Western blot analysis

[0101] Western blot analysis was performed on RAW264.7 cells, HepG2 cells, and mouse hepatocytes. In vitro cells were harvested after being washed twice with phosphate-buffered saline (PBS) under appropriate conditions. Cells were lysed using radioimmunoprecipitation assay (RIPA) cell lysis buffer containing protease and phosphatase inhibitors, and protein amounts were quantified using BCA protein assay (Thermo, USA). Proteins were separated by size by loading onto a 10% SDS-PAGE gel so that equal amounts of protein per sample could be loaded. After transfer to PVDF membrane (Millipore, USA), membranes were blocked with 5% BSA, and primary antibodies were reacted overnight at 4°C. The primary antibodies used, Phospho-p38 MAPK (#9211), p38 MAPK (#8690), Phospho-NF-κB p65 (#3033), NF-κB p65 (#8242), Fatty Acid Synthase (#3180), PPARγ (#2435), C / EBPα (#8178), Phospho-Acetyl-CoA Carboxylase (#3661), Acetyl-CoA Carboxylase (#3676), α-Smooth Muscle Actin (#14968), Phospho-SMAD2 (#18338), Smad2 (#5339), were purchased from Cell Signaling Technology (Cell Signaling Technology, Danvers, MA, USA), and SREBP-1 (sc-365513) and β-Actin (sc-47778) were purchased from Santa Cruz Biotechnology. (Santa Cruz Biotechnology, CA, USA) was purchased.Afterwards, the membrane was incubated with a secondary antibody of peroxidase (HRP)-conjugated goat anti-rabbit (Thermo Fisher, #31460) or anti-mouse IgG (Thermo Fisher, #31430) at room temperature for 1 hour. After washing with PBST, protein expression was measured on a PVDF membrane reacted with SuperSignal West Pico Chemiluminescent Substrate (Thermo Fisher, #34078), and the results were analyzed using the CSAnalyzer 4 (ATTO Technology, NY, USA) program.

[0102] (4) ELISA analysis

[0103] Enzyme-linked immunosorbent assays (ELISAs) were performed to measure the amount of cytokines secreted from cells or tissues. The types of cytokines analyzed were IL-1 beta (#88-7013-22), IL-6 (#88-7064-88), TNF alpha (#88-7324-88), and TGF beta-1 (#88-8350-88). All analyses were performed using ELISA kits (Invitrogen, Carlsbad, CA, USA) and the experiments were performed according to the provided protocol. To measure the amount of cytokines secreted in in vivo samples, mouse serum and liver tissue were used. Serum was isolated from blood. To obtain liver tissue samples, liver tissue isolated from mice was lysed using radioimmunoprecipitation assay (RIPA) cell lysis buffer containing protease and phosphatase inhibitors and 1 mM PMSF. ELISA analysis was performed using liver tissue samples obtained through this method.

[0104] (5) Preparation of experimental animals and administration of TN13 (in vivo experiment)

[0105] Five-week-old male C57BL / 6 mice were purchased from Dooyeolbiotech (Seoul, Korea). All mice were housed in a Specific Pathogen Free (SPF) environment with a 12-hour light / dark cycle and constant temperature and humidity. After a one-week acclimation period, they were randomly assigned to negative control, positive control, and experimental groups. The negative control group was fed a normal diet (Inotiv, #2018s), while the positive control and experimental groups were fed an adjusted calorie 60% HFD (Inotiv, #TD.06414). The experimental group was fed a high-fat diet and simultaneously received 25 mg / kg of TN13 by intraperitoneal injection twice a week. At the 15th week, the mice were sacrificed, and blood, epididymal fat (eWAT), and liver were isolated for analysis.

[0106] (6) Blood analysis measurement

[0107] Blood glucose and complete blood count (CBC) were measured in whole blood from mice using a biochemical analyzer. Serum was used for ELISA analysis and aspartate transaminase (AST) and alanine transferase (ALT) analysis. To obtain serum, blood was collected, left at room temperature for 30 minutes, and then centrifuged at 3,000 rpm for 10 minutes at 4°C. ELISA analysis was performed as described above, and AST and ALT analyses were performed using a biochemical analyzer.

[0108] (7) Histological staining

[0109] Adipose tissue and liver tissue were each prepared as paraffin blocks, and then sectioned to prepare paraffin sections. Adipose tissue sections were stained with hematoxylin and eosin (H&E). Liver tissue sections were stained with H&E, Sirius Red, and immunohistochemistry (IHC) using F4 / 80 (CST, #70076) and Neutrophil (Abcam, #ab2557) antibodies. Additionally, liver tissue was prepared as frozen sections and stained with Oil Red O.

[0110] (8) TG measurement

[0111] Liver tissue was pulverized with 5% TritonX-100 solution, and triglyceride (TG) was dissolved in a constant-temperature water bath at 80-100°C for 3-5 minutes. The supernatant was centrifuged at 13,000 rpm for 5 minutes, and the TG value was measured. The TG assay kit (Bioassay system, #ETGA-200) was used, and the measurement method was performed according to the instructions provided by the manufacturer.

[0112] Example 2. Analysis of the anti-inflammatory effect of TNXIP-derived peptides in mouse macrophages.

[0113] In order to confirm the cytotoxicity of the TN13 peptide, a TNXIP-derived peptide, on cell lines, TAT-TN13 (TN13) prepared according to Example 1-1 was treated to cells, and cell viability was confirmed 24 hours later through CCK8 analysis.

[0114] The analysis results showed that there was no significant effect on cell survival up to a concentration of 20 μM (Fig. 1A).

[0115] In addition, to confirm the inhibitory effect of TN13 on p38 phosphorylation (phospho p38, p-p38) in RAW246.7 cells, cells were pretreated with various concentrations of TN13 for 1 hour, and then treated with 100 ng / ml LPS for 30 minutes, and protein expression was confirmed through Western blotting.

[0116] As a result, compared to the positive control group treated with only LPS, the experimental group treated with TN13 showed a decrease in p-p38 expression in a TN13 concentration-dependent manner (Fig. 1B).

[0117] In addition, when cells were pretreated with 20 μM TN13 for 1 hour and then treated with 100 ng / ml LPS for 16 hours, morphological changes induced by LPS-induced macrophage activation were observed under a microscope.

[0118] The analysis results confirmed that morphological changes induced by LPS-induced macrophage activation were suppressed when TN13 was treated (Fig. 1C).

[0119] Through these analysis results, it was confirmed that TN13 according to the present invention has an anti-inflammatory effect.

[0120] Example 3. Analysis of the inhibitory effect of TNXIP-derived peptides on macrophage activation by palmitic acid (PA).

[0121] Palmitic acid (PA), a saturated fatty acid, is a common saturated fatty acid found in fats and waxes, including olive oil, palm oil, and body fat, and is known to induce liver damage, fatty liver disease, insulin resistance, and metabolic-related fatty liver disease, depending on the dose.

[0122] In addition, the inhibitory effect of p38-NFκB signaling pathway in RAW246.7 cells upon treatment with Palmitic Acid (PA) was confirmed through Western blotting of protein expression, and the results are shown in Figure 2.

[0123] As can be seen in Fig. 2, treatment with PA increased the phosphorylation of p38 and p65, and TN13 of the present invention was found to significantly inhibit this.

[0124] Therefore, we confirmed that TN13 effectively acts in the inflammatory response induced by saturated fatty acids by inhibiting the phosphorylation of p38, which is activated by the MAPK pathway that activates the inflammatory response, and p65, which is activated by the NF-κB pathway.

[0125] Example 4. Analysis of cell viability and cell permeability

[0126] Cells were treated with various concentrations of TNXIP-derived peptide (TN13) for 24 hours, and cell viability was confirmed through CCK-8 assay.

[0127] As a result, even when treated with a concentration of 100 μM TN13, no significant difference in cell viability was observed, indicating excellent cell viability (Fig. 3A).

[0128] In addition, to confirm cell permeability, an experiment was conducted using FITC-TAT-TN13 of Example 1-2, which was conjugated with the fluorescent dye FITC. FITC-TAT-TN13 was treated to cells for 2 hours, and the results were confirmed using a flow cytometer and fluorescence microscope.

[0129] As a result, cell permeability was found to increase in a concentration-dependent manner (Figures 3B and 3C).

[0130] As a result of the cell viability and cell penetration analysis as above, it was confirmed that the concentration of 50 μM TN13 did not affect cell viability and was a concentration that penetrated well into cells.

[0131] Example 5. Analysis of the lipogenesis inhibitory effect of TNXIP-derived peptides in liver cancer cell lines.

[0132] We investigated whether a TNXIP-derived peptide (TN13) reduced adipogenesis through p-p38 inhibition in PA-induced HepG2 cells. To confirm the increase in p-p38 levels upon PA treatment, cells were treated with 100 μM PA, harvested at 15 min, 30 min, 1 h, 2 h, 4 h, 8 h, 12 h, and 24 h, and western blotting was performed.

[0133] As a result, the level of p-p38 was highest when PA was treated for 15 minutes, and its expression gradually decreased over time (Fig. 4A).

[0134] Based on these results, HepG2 cells were pretreated with TN13 for 1 hour and then treated with PA for 15 minutes to confirm changes in p-p38 levels.

[0135] As a result, it was shown that the expression of p-p38 increased by PA treatment decreased in a TN13 concentration-dependent manner (Fig. 4B).

[0136] Additionally, to induce adipogenesis in HepG2 cells, we treated them with PA and performed Oil Red O staining, a fat-soluble dye that can detect fat accumulation in cells, to confirm whether TN13 effectively inhibits it.

[0137] As a result, it was shown that fat accumulation was reduced by TN13 (Fig. 4C).

[0138] Example 6. Analysis of the anti-obesity effect of TNXIP-derived peptide (TN13) in vivo using a mouse model.

[0139] To confirm the effects of TNXIP-derived peptide (TN13) on obesity suppression and improvement of chronic inflammation and metabolic fatty liver disease, mice were divided into three experimental groups and an experiment was conducted. Six-week-old C57BL / 6J mice were randomly divided into a normal diet group (negative control group), a high-fat diet group (positive control group), and a TN13-administered high-fat diet group (experimental group). They were fed regular feed or a high-fat diet (60% fat) for 15 weeks, and their body weights were measured every week.

[0140] The experimental group was administered TN13 at a concentration of 25 mg / kg twice a week via intraperitoneal injection during the same period (Fig. 5).

[0141] The analysis results showed that changes in body weight between the normal diet group and the high-fat diet group began to appear one week after starting the diet, and from the 10th week, a large difference was observed with a weight increase of more than 50% compared to the normal diet group.

[0142] A significant difference in body weight was observed between the high-fat diet group and the experimental group from the third week, and ultimately, the body weight of the experimental group was found to have decreased by more than 30% (Fig. 6A, B, C).

[0143] At week 15, whole blood was collected via orbital puncture after anesthesia with Evertin. After complete sacrifice via CO2 inhalation, epididymal fat and liver were isolated. Obesity is often accompanied by hyperglycemia. To confirm this, blood glucose levels were measured, and the experimental group showed lower values ​​than the high-fat diet group (Fig. 7).

[0144] In addition, when measuring the weight of fat, the most direct factor for judging obesity, the epididymal fat of the experimental group was found to be reduced in weight compared to the high-fat diet group (Figs. 8A, 8B). To visually observe changes in the size of fat cells, epididymal white adipose tissue (eWAT), a white fat, was prepared as a paraffin section and H&E staining was performed. As a result, it was found that the size of fat cells was significantly reduced in the experimental group (Fig. 8C).

[0145] Example 7. In vivo analysis of the anti-inflammatory effect of TNXIP-derived peptide (TN13) using a mouse model.

[0146] The anti-inflammatory effect was confirmed using the animal model of Example 6. As a result of examining the distribution of white blood cells through complete blood cell count (CBC) analysis, it was found that the number of neutrophils, monocytes, and basophils, which are known to significantly contribute to the inflammatory response, decreased when treated with a TNXIP-derived peptide (TN13) (Fig. 9A, B, C).

[0147] In this way, cells involved in the inflammatory response promote the inflammatory response by secreting pro-inflammatory cytokines such as TNF-α, IL-1β, and IL-6. When the level of secretion of these cytokines was examined in serum, it was found that all of them were reduced when treated with a TNXIP-derived peptide (TN13) (Fig. 9D, E, F).

[0148] These results confirmed that the TNXIP-derived peptide (TN13) of the present invention is effective not only in suppressing obesity but also in improving chronic inflammation occurring throughout the body.

[0149] Example 8. Analysis of the inhibitory effect of TNXIP-derived peptide (TN13) on fatty liver formation in vivo using a mouse model.

[0150] The inhibitory effect on fatty liver formation due to metabolic abnormalities was confirmed using the animal model of Example 6. Compared to the normal diet group (ND), the high-fat diet group (HFD) showed an increase in liver size and weight and a pale liver color, but the experimental group administered TN13 (HFD+TN13) showed liver weight and morphological characteristics similar to those of the normal diet group (Fig. 10A, B).

[0151] Fatty liver disease occurs when lipid metabolites such as triglycerides accumulate in hepatocytes. In the experimental group, triglyceride levels decreased to the level of the normal diet group (Figure 10C).

[0152] In addition, H&E-stained liver tissues showed pathological features of fatty liver, such as steatosis, intralobular inflammation, and hepatocellular ballooning. Oil Red O staining confirmed changes in the size and accumulation of lipid droplets within hepatocytes. This confirmed a significant reduction in steatosis in the experimental group (Fig. 10D). In addition to these visual inspections and histopathological evaluations, differences in protein expression between the positive control group and the experimental group were also confirmed.

[0153] In addition, a decrease in the protein expression of FAS, SREBP-1, PPAR-g, C / EBPa, p-ACC, and ACC, which are factors related to fat production, was confirmed in the experimental group, and a decrease in the protein expression of p-p38 by TN13 treatment was confirmed (Fig. 10E).

[0154] These results confirmed that the TNXIP-derived peptide (TN13) effectively inhibits the phosphorylation of p38, thereby exhibiting pharmacological changes in the inhibitory effect on the development of fatty liver disease.

[0155]

[0156] Example 9. Analysis of the inhibitory effect of TNXIP-derived peptide (TN13) on metabolic abnormalities such as steatohepatitis and liver fibrosis in vivo using a mouse model.

[0157] Using the animal model of Example 6, the inhibitory effects of TN13 on metabolic abnormalities, steatohepatitis, and liver fibrosis were confirmed. Exacerbation of fatty liver disease can progress to steatohepatitis or liver fibrosis. Experiments were conducted to determine the inhibitory effects of TN13 on these conditions.

[0158] As a result, in the experimental group administered TN13 (HFD+TN13), the levels of pro-inflammatory cytokines TNF-α, IL-1β, and IL-6 in liver tissue were significantly reduced, showing an excellent effect on the inflammatory response (Fig. 11A, B, C).

[0159] Furthermore, the number of macrophages and neutrophils actually distributed in liver tissue was quantified using IHC staining, which can detect the presence or absence of specific proteins in tissue sections. As a result, the number of these immune cells was significantly reduced in the experimental group administered TN13 (HFD+TN13) (Fig. 12A, B, C). These results suggest that TN13 may be effective not only in simple hepatic steatosis but also in steatohepatitis.

[0160] In addition, the results of Sirius red staining analysis, a histological examination method that can confirm liver fibrosis, showed a significant effect of inhibiting liver fibrosis in the experimental group administered TN13 (HFD+TN13) (Fig. 13A). In addition, the experimental group administered TN13 (HFD+TN13) showed a significant decrease in the secretion of TGF-β, a cytokine that promotes fibrosis (Fig. 13B), and there were accompanying changes in the protein expression of p-SMAD2 activated by TGF-β and α-SMA, a fibrogenic factor (Fig. 13C). These results suggest that TN13 can inhibit the progression of liver fibrosis.

[0161] Additionally, we observed increases in aspartate aminotransferase (AST) and alanine aminotransferase (ALT), indicators that can comprehensively assess liver damage. Serum was separated from the collected blood and measured. The ALT and AST levels in the experimental group were significantly reduced compared to the high-fat diet group (Fig. 14A, B).

[0162] The present invention has been described by way of example, and those skilled in the art will appreciate that various modifications may be made without departing from the essential characteristics of the present invention. Therefore, the embodiments disclosed herein are intended to illustrate, rather than limit, the present invention, and the spirit and scope of the present invention are not limited by these embodiments. The scope of protection of the present invention should be construed according to the claims below, and all techniques within the scope equivalent thereto should be construed as being included within the scope of the present invention.

Claims

1. A pharmaceutical composition for preventing or treating obesity or liver disease, comprising a peptide derived from a thioredoxin binding protein having an amino acid sequence of sequence number 1.

2. A pharmaceutical composition according to claim 1, wherein the peptide derived from the thioredoxin binding protein has an amino acid sequence of 30 or fewer amino acids and includes an amino acid sequence of sequence number 1.

3. A pharmaceutical composition according to claim 1, wherein the peptide derived from the thioredoxin binding protein consists of an amino acid sequence of sequence number 1.

4. A pharmaceutical composition according to claim 1, wherein the thioredoxin-binding protein-derived peptide is linked to any one cell-penetrating peptide selected from the group consisting of amino acid sequences of SEQ ID NOs: 2 to 9.

5. A pharmaceutical composition according to claim 4, wherein the cell-penetrating peptide comprises an amino acid sequence of sequence number 2.

6. A pharmaceutical composition according to claim 1, wherein the liver disease is any one selected from the group consisting of hepatic fibrosis, cholesterolosis, cirrhosis, viral and alcoholic hepatitis, Wilson's disease, hemochromatosis, steatosis, metabolic fatty liver disease (MASLD), and metabolic associated steatohepatitis (MASH).

7. A pharmaceutical composition according to claim 1, wherein the liver disease is any one selected from the group consisting of liver fibrosis, cirrhosis, metabolic fatty liver disease (MASLD), and metabolic-associated steatohepatitis (MASH).

8. A food composition for preventing or improving obesity or liver disease, comprising a peptide derived from a thioredoxin binding protein having an amino acid sequence of sequence number 1.

9. A food composition according to claim 8, wherein the peptide derived from the thioredoxin binding protein has an amino acid sequence of 30 or fewer amino acids and includes an amino acid sequence of sequence number 1.

10. A food composition according to claim 8, wherein the peptide derived from the thioredoxin binding protein consists of an amino acid sequence of sequence number 1.

11. A food composition according to claim 8, wherein the thioredoxin-binding protein-derived peptide is linked to any one cell-penetrating peptide selected from the group consisting of amino acid sequences of SEQ ID NOs: 2 to 9.

12. A food composition according to claim 8, wherein the cell-penetrating peptide comprises an amino acid sequence of sequence number 2.

13. A food composition according to claim 8, wherein the liver disease is any one selected from the group consisting of hepatic fibrosis, cholesterolosis, cirrhosis, viral and alcoholic hepatitis, Wilson's disease, hemochromatosis, steatosis, metabolic fatty liver disease (MASLD), and metabolic associated steatohepatitis (MASH).

14. A food composition in claim 8, wherein the liver disease is any one selected from the group consisting of liver fibrosis, cirrhosis, metabolic fatty liver disease (MASLD), and metabolic-associated steatohepatitis (MASH).

15. A method for preventing or treating obesity or liver disease, comprising administering to a subject in need thereof a composition comprising a peptide derived from a thioredoxin binding protein comprising an amino acid sequence of sequence number 1.

16. Use of a peptide derived from a thioredoxin binding protein comprising an amino acid sequence of sequence number 1 for the manufacture of a medicament for preventing or treating obesity or liver disease.

17. A pharmaceutical composition comprising a peptide derived from a thioredoxin binding protein comprising an amino acid sequence of sequence number 1 for use in the prevention or treatment of obesity or liver disease.

18. A method according to claim 15, wherein the liver disease is any one selected from the group consisting of hepatic fibrosis, cholesterolosis, cirrhosis, viral and alcoholic hepatitis, Wilson's disease, hemochromatosis, steatosis, metabolic fatty liver disease (MASLD), and metabolic associated steatohepatitis (MASH).

19. The use according to claim 16, wherein the liver disease is any one selected from the group consisting of hepatic fibrosis, cholesterolosis, cirrhosis, viral and alcoholic hepatitis, Wilson's disease, hemochromatosis, steatosis, metabolic fatty liver disease (MASLD) and metabolic associated steatohepatitis (MASH).

20. A pharmaceutical composition according to claim 17, wherein the liver disease is any one selected from the group consisting of hepatic fibrosis, cholesterolosis, cirrhosis, viral and alcoholic hepatitis, Wilson's disease, hemochromatosis, steatosis, metabolic fatty liver disease (MASLD), and metabolic associated steatohepatitis (MASH).

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