Use of novel surface protein lrig specifically present on regulatory t cells

Abstract

An Lrig inhibitor or Lrig expression inhibitor according to the present invention specifically binds to an Lrig protein present on preadipocytes, adipocytes, or regulatory T cells to regulate the function of the regulatory T cells, thereby regulating the activity of effector T cells, and thus can be very efficiently used for the prevention, amelioration, or treatment of metabolic diseases.

Description

Uses of LRIG, a novel surface protein specifically present on regulatory T cells

[0001] The present invention relates to a next-generation biopharmaceutical, and more particularly, to a therapeutic pharmaceutical composition capable of preventing or treating metabolic diseases by inducing gene editing in the body by administering an inhibitor of Lrig protein or miRNA, siRNA, etc. that inhibits the expression of Lrig protein.

[0002]

[0003] Since the concept of suppressor T cells, which could potentially control and suppress the effector function of conventional T cells, was first proposed by Gershon in the early 1970s, research has been conducted in many fields of immunology to elucidate the biological characteristics and functions of regulatory T cells.

[0004] Regulatory T cells (Tregs) play a crucial role in naturally preventing excessive inflammation and immune responses. However, it has been reported that the function and number of regulatory T cells are significantly reduced in autoimmune and chronic inflammatory diseases. Therefore, for patients with immune and inflammatory diseases, it is crucial to ensure normal levels of regulatory T cells, which may be a potential treatment option.

[0005] Meanwhile, obesity refers to a state in which excessive fat is accumulated in the body. Men are considered obese when their body fat exceeds 25% of their body weight, and women when their body fat exceeds 30% of their body weight. Clinically, obesity is defined as a BMI (Body Mass Index) of 30.0 or higher. Causes of obesity include excessive calorie intake, endocrine disorders, lack of exercise, and genetic factors, but among them, excess energy accumulation is the most directly related. Obesity is divided into simple (essential) obesity, which is caused when energy expenditure is less than energy intake without any specific cause, and symptomatic obesity, which is caused by endocrine diseases (insulin-dependent diabetes mellitus, Cushing's syndrome, ovarian dysfunction, hypothyroidism) or hypothalamic disorders (which appear when the brain's satiety center is damaged). Current treatments for obesity can be broadly divided into drugs that affect appetite by acting on the central nervous system and drugs that act on the gastrointestinal tract to inhibit absorption. Drugs that act on the central nervous system include drugs such as fenfluramine and dexfenfluramine, which inhibit the serotonin (5HT) nervous system, depending on their mechanism of action; drugs such as ephedrine and caffeine, which act on the noradrenergic nervous system; and recently, drugs such as sibutramine, which acts simultaneously on the serotonin and noradrenergic nervous systems to suppress obesity. In addition, drugs that act on the gastrointestinal tract to suppress obesity include orlistat, which was recently approved as an obesity treatment, which reduces fat absorption by inhibiting the activity of lipase produced in the pancreas. However, previously used drugs such as fenfluramine have recently been banned due to side effects such as primary pulmonary hypertension and heart valve lesions. Other drugs also cause problems such as hypotension and lactic acidosis, which prevents their use in patients with heart failure or renal disease.

[0006] Accordingly, adipocyte differentiation inhibitors have emerged as a way to prevent or treat obesity with minimal side effects. Fat stored in adipocytes serves as a significant energy source for the body. However, as obesity progresses, adipocytes not only increase in number but also undergo morphological changes, including an increase in adipocyte size due to the synthesis and accumulation of large amounts of triglycerides through adipocyte differentiation. This is accompanied by changes in the expression of various genes secreted by adipocytes. This increase in adipocyte size is caused by the synthesis and storage of excess energy in the form of triglycerides. Depending on the amount of fat stored, adipocytes can increase in size by approximately 20 times their diameter, resulting in a thousands-fold increase in cell volume. This increase in adipocyte size is generally achieved through dietary control. However, dietary control cannot suppress the differentiation of new preadipocytes into adipocytes. Therefore, controlling adipocyte differentiation is crucial for the fundamental treatment or suppression of obesity. Adipocyte differentiation is promoted by stimulation by insulin, insulin-like growth factor-1 (IGF-1), and growth hormone, and during this process, an increase in transcription factors such as the CCAAT enhancer-binding protein (C / EBP) family and peroxisome proliferator-activated receptor-γ (PPAR-γ) is observed. These transcription factors, along with adipocyte regulatory factors, promote adipocyte differentiation and increase the expression of enzymes such as the fatty acid binding protein aP2 and fatty acid synthase.

[0007] Recently, active research has been conducted to explore substances that inhibit adipocyte differentiation, based on the idea that inhibiting adipocyte differentiation can control the number of adipocytes produced and release the excess energy accumulated accordingly.

[0008]

[0009] One object of the present invention is to provide a pharmaceutical composition or treatment method for preventing or treating a metabolic disease, comprising an Lrig protein inhibitor or an Lrig protein expression inhibitor as an active ingredient.

[0010] Another object of the present invention is to provide a composition for diagnosing a metabolic disease, a method for diagnosing a metabolic disease, a diagnostic device, a diagnostic kit, and a method for providing information for diagnosis, comprising an agent for measuring the expression level of a Lrig protein or a fragment thereof; or a gene encoding the same.

[0011] However, the technical problems to be solved by the present invention are not limited to the problems mentioned above, and other problems not mentioned can be clearly understood by those skilled in the art from the description below.

[0012]

[0013] Various embodiments of the present invention are described with reference to the drawings. In the following description, various specific details, such as specific configurations, compositions, and processes, are set forth to provide a thorough understanding of the present invention. However, certain embodiments may be practiced without one or more of these specific details, or in conjunction with other known methods and configurations. In other instances, well-known processes and manufacturing techniques are not described in specific detail so as not to unnecessarily obscure the present invention. Reference throughout this specification to an embodiment means that a particular feature, configuration, composition, or characteristic described in connection with the embodiment is included in one or more embodiments of the present invention. Thus, the appearance of an embodiment in various places throughout this specification does not necessarily indicate the same embodiment of the present invention. Additionally, the particular features, configurations, compositions, or characteristics may be combined in any suitable manner in one or more embodiments.

[0014] In one embodiment of the present invention, a pharmaceutical composition for preventing or treating a metabolic disease is provided, comprising an Lrig protein inhibitor or an Lrig protein expression inhibitor as an active ingredient.

[0015] In the present invention, the "Lrig-1 protein" is a transmembrane protein present on the surface of regulatory T cells, and is composed of a leucine-rich repeat (LRR) sequence on the extracellular or lumen side, three immunoglobulin-like domains, a transmembrane sequence, and a cytoplasmic tail. The LRIG gene family consists of LRIG1, LRIG2, and LRIG3, and the amino acids constituting each family are highly conserved. The LRIG1 gene is highly expressed in normal skin, and can regulate the proliferation of epithelial stem cells by being expressed in basal and hair follicle cells. Therefore, it plays an important role in maintaining epidermal homeostasis, and its absence can lead to psoriasis or skin cancer. It has been reported that when the chromosome 3p14.3 where LRIG1 is located is cut off, there is a possibility of developing into cancer cells, and in fact, it has been confirmed that the expression of LRIG1 is greatly reduced in renal cell carcinoma and cutaneous squamous cell carcinoma. However, it has recently been revealed that only about 20-30% of cancers express the Lrig-1 protein. Meanwhile, for the purpose of the present invention, the Lrig-1 protein may be a protein derived from mammals or mice, but is not limited thereto.

[0016] In one example of the present invention, the Lrig-1 protein may be an Lrig-1 protein derived from a mammal, for example, a primate such as a human or monkey, or a rodent such as a mouse or rat.

[0017] In another example of the present invention, the Lrig-1 protein may be, but is not limited to, the extracellular domain of the Lrig-1 protein.

[0018] The Lrig-1 extracellular domain of the present invention may be an extracellular domain of an Lrig-1 protein derived from a mammal, for example, a primate such as a human or monkey, or a rodent such as a mouse or rat. For the purposes of the present invention, the extracellular protein of the Lrig-1 protein may be an extracellular domain of an Lrig-1 protein derived from a human or mouse, but is not limited thereto.

[0019] In one example of the present invention, the extracellular domain of the Lrig-1 protein may correspond to, but is not limited to, the 35th to 794th amino acid sequence of the human Lrig-1 protein.

[0020] In another example of the present invention, the extracellular domain of the Lrig-1 protein may correspond to, but is not limited to, the 35th to 794th amino acid sequence of the mouse-derived Lrig-1 protein.

[0021] In the present invention, the “Lrig protein inhibitor” or “Lrig protein expression inhibitor” is not particularly limited as long as it is a substance capable of inhibiting the expression of the Lrig protein.

[0022] In another embodiment of the invention, a pharmaceutical composition is provided, wherein the Lrig protein expression inhibitor is at least one selected from the group consisting of antisense nucleotides, short interfering RNA (siRNA), microRNA (mircoRNA; miRNA), short hairpin RNA, and ribozymes, dsNRA, aptamers, peptide nucleic acids (PNA), ZFNs, TALENs, and CRISPR (cluster regularly interspaced short palindromic repeats)-related nucleic acids, single guide RNA (sgRNA), CRISPR-RNA (crRNA), and trans-activating crRNA (tracrRNA) that complementarily bind to the transcript sequence of the Lrig protein.

[0023] As used herein, the term "antisense oligonucleotide" or "antisense compound" refers to a molecule of RNA, DNA, LNA, PNA, or a mixture thereof that binds to another RNA or DNA (target RNA, DNA). For example, if it is an RNA oligonucleotide, it binds to another RNA target through RNA-RNA interaction and modulates the activity of the target RNA. Antisense oligonucleotides can upregulate or downregulate the expression and / or function of a specific polynucleotide. This definition is intended to include any foreign RNA or DNA molecule useful from a therapeutic, diagnostic, or other perspective. Such molecules include, for example, antisense RNA or DNA molecules, interfering RNA (RNAi), microRNA, decoy RNA molecules, siRNA, enzymatic RNA, therapeutic editing RNA, and antisense oligomeric compounds, antisense oligonucleotides, external guide sequence (EGS) oligonucleotides, primers, probes, and other oligomeric compounds that hybridize to at least a portion of a target nucleic acid. Therefore, these compounds can be introduced in the form of single-stranded, double-stranded, partially single-stranded, or circular oligomeric compounds.

[0024] In the context of the present invention, the term "oligonucleotide" refers to an oligomer or polymer of ribonucleic acid (RNA) or deoxyribonucleic acid (DNA), or a mimic thereof. The term "oligonucleotide" also includes linear or circular oligomers of natural and / or modified monomers or chains, including deoxyribonucleosides, ribonucleosides, their substituted and alpha-anomeric forms, peptide nucleic acids (PNAs), locked nucleic acids (LNAs), phosphorothioates, methylphosphonates, and the like. Oligonucleotides can specifically bind to a target polynucleotide by a regular pattern of monomer-monomer interactions, such as Watson-Crick type base pairing, Hoogsteen or reverse Hoogsteen type base pairing, and the like.

[0025] Oligonucleotides may be "chimeric," that is, composed of different regions. In the context of the present invention, a "chimeric" compound may be an oligonucleotide containing two or more chemical regions, such as DNA region(s), RNA region(s), PNA region(s), LNA region(s), etc.

[0026] Oligonucleotides can be composed of segments that can be linked, either sequentially, as in native DNA, or via spacers. These spacers are intended to form a covalent "bridge" between these segments and, if desired, have a length of no more than about 100 carbon atoms. These spacers can, for example, be positively or negatively charged, have specific nucleic acid binding properties (such as insertions, groove binders, toxins, fluorophores, etc.), be lipophilic, and have different functionalities that induce specific secondary structures, such as alanine-containing peptides that induce alpha-helices.

[0027] As used herein, the term "target nucleic acid" encompasses DNA, RNA transcribed from such DNA (including pre-mRNA and mRNA), and cDNA derived from such RNA encoding non-coding sequences, sense or antisense polynucleotides. The specific hybridization of an oligomeric compound with a target nucleic acid interferes with the normal function of the nucleic acid. This modulation of the function of the target nucleic acid by a compound that specifically hybridizes to the target nucleic acid is generally referred to as "antisense." Functions of the DNA interfered with include, for example, replication and transcription.

[0028] RNA interference ("RNAi") is mediated by double-stranded RNA (dsRNA) molecules with sequence-specific homology to a "target" nucleic acid sequence. In certain embodiments of the present invention, the mediator is a 5-25 nucleotide "small interfering" RNA duplex (siRNA). These siRNAs are derived from the processing of dsRNA by the RNase enzyme Dicer. The siRNA duplex product is recruited into a multi-protein siRNA complex called the RNA-Induced Silencing Complex (RISC). Without being bound by any particular theory, it is believed that the RISC is then guided to the target nucleic acid (preferably mRNA), where the siRNA duplex interacts in a sequence-specific manner to mediate catalytic cleavage. Small interfering RNAs that can be utilized in accordance with the present invention can be synthesized and utilized according to procedures well known in the art and familiar to those skilled in the art. The small interfering RNA used in the method of the present invention suitably comprises from about 1 to about 50 nucleotides (nt). In non-limiting specific examples, the siRNA may comprise from about 5 to about 40 nt, from about 5 to about 30 nt, from about 10 to about 30 nt, from about 15 to about 25 nt, or from about 20 to about 25 nucleotides.

[0029] Technology that suppresses gene expression is an important tool in the development of therapeutics and target verification for disease treatment. Among these technologies, RNA interference (hereinafter referred to as 'RNAi') has been discovered to act on sequence-specific mRNAs in various types of mammalian cells since its role was discovered. When a long RNA duplex is delivered to a cell, the delivered RNA duplex is processed into 21 to 23 base pairs (bp) duplexes by the endonuclease Dicer, resulting in short interfering RNA (hereinafter referred to as 'siRNA'). The siRNA binds to the RNA-induced silencing complex (RISC), and the guide (antisense) strand recognizes and degrades the target mRNA, thereby sequence-specifically inhibiting the expression of the target gene.

[0030] That is, in the present invention, “siRNA (small interfering RNA)” is a small RNA fragment of 18 to 23 nucleotides in size generated by cleavage of double-stranded RNA by the Dicer enzyme, and can be used to specifically bind to mRNA having a complementary sequence and suppress the expression of the protein or mRNA.

[0031] The term "nucleotide" encompasses both naturally occurring and non-naturally occurring nucleotides. It will be apparent to those skilled in the art that a variety of nucleotides previously considered "non-naturally occurring" have subsequently been discovered in nature. Therefore, the term "nucleotide" encompasses not only the known purine and pyrimidine heterocycle-containing molecules, but also their heterocyclic analogs and tautomers. Illustrative examples of other types of nucleotides are molecules containing adenine, guanine, thymine, cytosine, uracil, purine, xanthine, diaminopurine, 8-oxo-N6-methyladenine, 7-deazaxanthine, 7-deazaguanine, N4, N4-ethanocytosine, N6, N6-ethano-2,6-diaminopurine, 5-methylcytosine, 5-(C3-C6)-alkynylcytosine, 5-fluorouracil, 5-bromouracil, pseudoisocytosine, 2-hydroxy-5-methyl-4-triazolopyridine, isocytosine, isoguanine, inosine, and the "non-naturally occurring" nucleotides described in Benner et al., US Pat No. 5,432,272.

[0032] The term "nucleotide" includes all of these examples, as well as analogs and tautomers thereof. Of particular interest are nucleotides containing adenine, guanine, thymine, cytosine, and uracil, which are considered naturally occurring nucleotides for therapeutic and diagnostic applications in humans. Nucleotides include, for example, natural 2'-deoxy and 2'-hydroxyl sugars, and their analogs, as described in Kornberg and Baker, DNA Replication, 2nd Ed. (Freeman, San Francisco, 1992).

[0033] With respect to nucleotides, “analogs” include synthetic nucleotides having modified base moieties and / or modified sugar moieties (see, e.g., Scheit, Nucleotide Analogs, John Wiley, New York, 1980; Freier & Altmann, (1997) Nucl. Acid. Res., 25(22), 4429-4443, Toulme JJ, (2001) Nature Biotechnology 19:17-18; Manoharan M., (1999) Biochemica et Biophysica Acta 1489:117-139; Freier SM, (1997) Nucleic Acid Research, 25:4429-4443, Uhlman, E., (2000) Drug Discovery & Development, 3: 203-213, Herdewin P., (2000) Antisense & Nucleic Acid Drug Dev., 10:297-310); 2'-O, 3'-C-linked [3.2.0] bicycloarabinonucleosides). These analogs include synthetic nucleotides designed to enhance binding properties, such as double- or triple-helix stability, specificity, etc.

[0034] As used herein, "complementarity" refers to the ability of precise pairing between two nucleotides in one or both oligomeric strands. For example, if a nucleobase at a given position of an antisense compound can form a hydrogen bond with a nucleobase at a given position of a target nucleic acid, and the target nucleic acid is a DNA, RNA, or oligonucleotide molecule, the position of the hydrogen bond between the oligonucleotide and the target nucleic acid is considered to be a complementary position. An oligomeric compound and an additional DNA, RNA, or oligonucleotide molecule are complementary to each other when a sufficient number of complementary positions within each molecule are occupied by nucleotides capable of hydrogen bonding to each other. Therefore, the terms "specifically hybridizable" and "complementarity" are used to indicate a sufficient degree of precise pairing or complementarity over a sufficient number of nucleotides to allow stable and specific binding to occur between the oligomeric compound and the target nucleic acid.

[0035] In the art, the sequence of an oligomeric compound need not be 100% complementary to the sequence of a target nucleic acid in order to hybridize specifically. Furthermore, oligonucleotides can hybridize across more than one segment (e.g., a loop structure, a mismatch, or a hairpin structure) such that intervening or adjacent segments are not involved in the hybridization phenomenon. The oligomeric compounds of the present invention comprise at least about 70%, or at least about 75%, or at least about 80%, or at least about 85%, or at least about 90%, or at least about 95%, or at least about 99% sequence complementarity to a target region within the target nucleic acid sequence to which they are targeted. For example, if 18 of the 20 nucleotides of an antisense compound are complementary to the target region, then an antisense compound that hybridizes specifically will exhibit 90% complementarity. In these examples, the remaining non-complementary nucleotides may be clustered with the complementary nucleotides, or the complementary nucleotides may be interspersed, and need not be adjacent to each other or the complementary nucleotides. Therefore, an antisense compound having an 18-nucleotide length and four non-complementary nucleotides flanking two regions of complete complementarity with the target nucleic acid will have 77.8% overall complementarity with the target nucleic acid and thus fall within the scope of the present invention. The percent complementarity of an antisense compound with a region of the target nucleic acid can be routinely determined using BLAST (basic local alignment search tools) programs and PowerBLAST programs known in the art.The percentage of homology, sequence identity, or complementarity can be determined, for example, by the Gap program (Wisconsin Sequence Analysis Package, Version 8 for Unix, Genetics Computer Group, University Research Park, Madison Wis.) using default settings and the algorithm of Smith and Waterman, Adv. Appl. Math., (1981) 2, 482-489.

[0036] The oligonucleotides of the present invention may be modified with any chemical or natural modification. Chemical and natural modifications are well known in the art. Such modifications include, for example, modifications designed to increase binding to a target strand (i.e., to increase their melting temperature), to aid in the identification of the oligonucleotide or oligonucleotide-target complex, to increase cellular penetration, to stabilize the oligonucleotide against nucleases and other enzymes that degrade or interfere with the structure or activity of the oligonucleotide, to provide a mode of termination upon sequence-specific binding to the target, or to improve the pharmacokinetic properties of the oligonucleotide. Modifications include, but are not limited to, for example, (a) terminal modifications, such as 5' terminal modifications (phosphorylation / dephosphorylation, splicing, inverted linkage, etc.), 3' terminal modifications (splicing, DNA nucleotide, inverted linkage, etc.), (b) base modifications, such as replacement with a modified base, a stabilizing base, a destabilizing base, or a base that base pairs with an expanded repertoire of partners, or a spliced ​​base, (c) sugar modifications (e.g., at the 2' position or the 4' position) or sugar replacements, as well as (d) internucleoside linkage modifications, such as modification or replacement of a phosphodiester linkage. To the extent that such modification interferes with translation (i.e., results in a decrease in translation of, for example, 50%, 60%, 70%, 80%, or 90% or more, compared to no modification, in an in vitro translation assay), the modification may be suboptimal for the methods and compositions described herein. Non-limiting examples of modified internucleoside linkages include phosphorothioates, chiral phosphorothioates, phosphorodithioates, phosphotriesters, aminoalkylphosphotriesters, methyl and other alkyl phosphonates having normal 3'-5' linkages, such as 3'-alkylene phosphonates and chiral phosphonates, phosphinates, phosphoramidates,Examples include 3'-amino phosphoramidate and aminoalkylphosphoramidates, thionophosphoramidates, thionoalkylphosphonates, thionoalkylphosphotriesters and boranophosphates, 2'-5' linked analogs thereof, and those having inverted polarity where adjacent pairs of nucleoside units are linked 3'-5' to 5'-3' or 2'-5' to 5'-2'. Various salts, mixed salts and free acid forms are also included. In some embodiments, the modified oligonucleotide is a single-stranded modified oligonucleotide. In some embodiments, the single-stranded modified oligonucleotide consists of 10-30, 10-35, 10-40, 10-45, 10-50, 10-60, 10-70, 10-80, 10-90, 10-100, or more than 100 linked nucleosides and has a gap segment. In some embodiments, the gap segment refers to one or more linked nucleic acids consisting of deoxynucleosides located at or near the center of the modified oligonucleotide, such as the single-stranded modified oligonucleotide. In some embodiments, the gap segment consists of 2-3, 2-4, 2-5, 2-6, 2-7, 2-8, 2-9, 2-10, 2-20, 2-30, 2-40 2-50, 10-20, 10-30, 10-40, or 10-50 linked deoxynucleosides. The 5' wing segment corresponds to a nucleic acid (e.g., a nucleoside) linked from the 5'-end of the modified oligonucleotide to the nucleic acid before the first nucleic acid at the 5'-end of the gap segment. The 3' wing segment corresponds to a nucleic acid (e.g., a nucleoside) linked from after the last nucleic acid at the 3'-end of the gap segment to the last nucleic acid at the 3'-end of the modified oligonucleotide. The gap segment is located between the 5' wing segment and the 3' wing segment. In some embodiments,At least one nucleoside of the 5' wing segment and / or at least one nucleoside of the 3' wing segment comprises a modified nucleoside. In some embodiments, both the internucleoside linkages within the gap segment and the linkages connecting the gap segment to the 3' wing segment and / or the 5' wing segment are phosphorothioate linkages (*). In some embodiments, the internucleoside linkages connecting the remaining nucleosides of both the 5' and 3' wing segments are phosphodiester linkages. In some embodiments, the nucleosides in the modified oligonucleotide are modified with a 2' O-methyl group. The nucleosides in the modified oligonucleotide may also be modified with any other modification described herein. In some embodiments, the nucleobase sequence of the modified oligonucleotide consists of 10-30, 10-35, 10-40, 10-45, 10-50, 10-60, 10-70, 10-80, 10-90, 10-100, or more than 100 linked nucleosides and has a pharmaceutically acceptable salt thereof. The modified internucleoside linkages that do not contain a phosphorus atom therein include internucleoside linkages formed by short-chain alkyl or cycloalkyl internucleoside linkages, mixed heteroatoms, and alkyl or cycloalkyl internucleoside linkages, or one or more short-chain heteroatom or heterocyclic internucleoside linkages. These include those having morpholino linkages (formed in part from the sugar moiety of the nucleoside); siloxane backbones; sulfide, sulfoxide, and sulfone backbones; formacetyl and thioformacetyl backbones; Methylene formacetyl and thioformacetyl backbones; alkene-containing backbones; sulfamate backbones; methylene imino and methylenehydrazino backbones; sulfonate and sulfonamide backbones; amide backbones; and others having mixed N, O, S, and CH2 component moieties. Substituted sugar moieties include, but are not limited to, one of the following at the 2' position: H (deoxyribose); OH (ribose); F; O-, S-,Or N-alkyl; O-, S- or N-alkenyl; O-, S- or N-alkynyl; or O-alkyl-O-alkyl (wherein alkyl, alkenyl and alkynyl can be substituted or unsubstituted C1 to C10 alkyl or C2 to C10 alkenyl and alkynyl). Chemically or naturally modified oligonucleotides can comprise at least one nucleotide modified at the 2' position of the sugar, most preferably a 2'-O-alkyl, 2'-O-alkyl-O-alkyl or 2'-fluoro-modified nucleotide or terminal cap. In other embodiments, RNA modifications include 2'-fluoro, 2'-amino and 2' O-methyl modifications on the ribose of a pyrimidine, an abasic residue, or an inverted base at the 3' terminus of the RNA. Oligonucleotides useful according to the present invention can comprise a single modified nucleoside. In other embodiments, the oligonucleotide can comprise at least two modified nucleosides, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, at least ten, at least fifteen, at least twenty or more modified nucleosides up to the entire length of the oligonucleotide. The nucleosides or nucleobases include the natural purine bases adenine (A) and guanine (G), and the pyrimidine bases thymine (T), cytosine (C), and uracil (U). Modified nucleosides may be substituted with other synthetic and natural nucleobases, such as inosine, xanthine, hypoxanthine, nubularine, isoguanicine, tubercidin, 2-(halo)adenine, 2-(alkyl)adenine, 2-(propyl)adenine, 2 (amino)adenine, 2-(aminoalkyl)adenine, 2 (aminopropyl)adenine, 2 (methylthio) N6 (isopentenyl)adenine, 6 (alkyl)adenine, 6 (methyl)adenine, 7 (deaza)adenine, 8 (alkenyl)adenine, 8-(alkyl)adenine, 8 (alkynyl)adenine, 8 (amino)adenine, 8-(halo)adenine, 8-(hydroxyl)adenine, 8 (thioalkyl)adenine, 8-(thiol)adenine, N6-(isopentyl)adenine,N6 (methyl)adenine, N6, N6 (dimethyl)adenine, 2-(alkyl)guanine, 2-(propyl)guanine, 6-(alkyl)guanine, 6 (methyl)guanine, 7 (alkyl)guanine, 7 (methyl)guanine, 7 (deaza)guanine, 8 (alkyl)guanine, 8-(alkenyl)guanine, 8 (alkynyl)guanine, 8-(amino)guanine, 8 (halo)guanine, 8-(hydroxyl)guanine, 8 (thioalkyl)guanine, 8-(thiol)guanine, N (methyl)guanine, 2-(thio)cytosine, 3 (deaza) 5 (aza)cytosine, 3-(alkyl)cytosine, 3 (methyl)cytosine, 5-(alkyl)cytosine, 5-(alkynyl)cytosine, 5 (halo)cytosine, 5 (methyl)cytosine, 5 (propynyl)cytosine, 5 (propynyl)cytosine, 5 (trifluoromethyl)cytosine, 6-(azo)cytosine, N4 (acetyl)cytosine, 3 (3-amino-3-carboxypropyl)uracil, 2-(thio)uracil, 5 (methyl) 2 (thio)uracil, 5 (methylaminomethyl)-2 (thio)uracil, 4-(thio)uracil, 5 (methyl) 4 (thio)uracil, 5 (methylaminomethyl)-4 (thio)uracil, 5 (methyl) 2,4 (dithio)uracil, 5 (methylaminomethyl)-2,4 (dithio)uracil, 5 (2-aminopropyl)uracil, 5-(alkyl)uracil, 5-(alkynyl)uracil, 5-(allylamino)uracil, 5 (aminoallyl)uracil, 5 (aminoalkyl)uracil, 5 (guanidiniumalkyl)uracil, 5 (1,3-diazol-1-alkyl)uracil, 5-(cyanoalkyl)uracil, 5-(dialkylaminoalkyl)uracil, 5 (dimethylaminoalkyl)uracil, 5-(halo)uracil, 5-(methoxy)uracil, uracil-5 oxyacetic acid, 5(methoxycarbonylmethyl)-2-(thio)uracil, 5 (methoxycarbonyl-methyl)uracil, 5(propynyl)uracil, 5 (propynyl)uracil, 5 (trifluoromethyl)uracil, 6 (azo)uracil, dehydrouracil, N3 (methyl) uracil, 5-uracil (i.e., pseudouracil), 2 (thio) pseudouracil, 4 (thio) pseudouracil, 2,4-(dithio) pseudouracil, 5-(alkyl) pseudouracil, 5-(methyl) pseudouracil, 5-(alkyl)-2-(thio) pseudouracil,5-(methyl)-2-(thio)pseudouracil, 5-(alkyl)-4 (thio)pseudouracil, 5-(methyl)-4 (thio)pseudouracil, 5-(alkyl)-2,4 (dithio)pseudouracil, 5-(methyl)-2,4 (dithio)pseudouracil, 1 substituted pseudouracil, 1 substituted 2(thio)-pseudouracil, 1 substituted 4 (thio)pseudouracil, 1 substituted 2,4-(dithio)pseudouracil, 1 (aminocarbonylethylenyl)-pseudouracil, 1 (aminocarbonylethylenyl)-2(thio)-pseudouracil, 1 (aminocarbonylethylenyl)-4 (thio)pseudouracil, 1 (aminocarbonylethylenyl)-2,4-(dithio)pseudouracil, 1 (aminoalkylaminocarbonylethylenyl)-pseudouracil, 1 (aminoalkylamino-carbonylethylenyl)-2(thio)-pseudouracil, 1 (aminoalkylaminocarbonylethylenyl)-4(thio)pseudouracil, 1 (aminoalkylaminocarbonylethylenyl)-2,4-(dithio)pseudouracil, 1,3-(diaza)-2-(oxo)-phenoxazin-1-yl, 1-(aza)-2-(thio)-3-(aza)-phenoxazin-1-yl, 1,3-(diaza)-2-(oxo)-phenthiazin-1-yl, 1-(aza)-2-(thio)-3-(aza)-phenthiazin-1-yl, 7-substituted 1,3-(diaza)-2-(oxo)-phenoxazin-1-yl, 7-substituted 1-(aza)-2-(thio)-3-(aza)-phenoxazin-1-yl, 7-substituted 1,3-(diaza)-2-(oxo)-phenthiazin-1-yl, 7-substituted 1-(aza)-2-(thio)-3-(aza)-phenthiazin-1-yl, 7-(aminoalkylhydroxy)-1,3-(diaza)-2-(oxo)-phenoxazin-1-yl, 7-(aminoalkylhydroxy)-1-(aza)-2-(thio)-3-(aza)-phenoxazin-1-yl, 7-(aminoalkylhydroxy)-1,3-(diaza)-2-(oxo)-phenthiazin-1-yl, 7-(Aminoalkylhydroxy)-1-(aza)-2-(thio)-3-(aza)-phenthiazin-1-yl, 7-(guanidiniumalkylhydroxy)-1,3-(diaza)-2-(oxo)-phenoxazin-1-yl, 7-(guanidiniumalkylhydroxy)-1-(aza)-2-(thio)-3-(aza)-phenoxazin-1-yl, 7-(guanidiniumalkyl-hydroxy)-1,3-(diaza)-2-(oxo)-phenthiazin-1-yl,7-(Guanidiniumalkylhydroxy)-1-(aza)-2-(thio)-3-(aza)-pentiazin-1-yl, 1,3,5-(triaza)-2,6-(dioxa)-naphthalene, inosine, xanthine, hypoxanthine, nubularine, tubercidin, isoguanisine, inosinyl, 2-aza-inosinyl, 7-deaza-inosinyl, nitroimidazolyl, nitropyrazolyl, nitrobenzimidazolyl, nitroindazolyl, aminoindolyl, pyrrolopyrimidinyl, 3-(methyl)isocarbostyrilyl, 5-(methyl)isocarbostyrilyl, 3-(methyl)-7-(propynyl)isocarbostyrilyl, 7-(aza)indolyl, 6-(methyl)-7-(aza)indolyl, imidazopyridinyl, 9-(methyl)-imidazopyridinyl, pyrrolopyridinyl, isocarbostyrilyl, 7-(propynyl)isocarbostyrilyl, propynyl-7-(aza)indolyl, 2,4,5-(trimethyl)phenyl, 4-(methyl)indolyl, 4,6-(dimethyl)indolyl, phenyl, naphthalenyl, anthracenyl, phenanthracenyl, pyrenyl, stilbenyl, tetracenyl, pentacenyl, difluorotolyl, 4-(fluoro)-6-(methyl)benzimidazole, 4-(methyl)benzimidazole, 6-(azo)thymine, 2-pyridinone, 5 nitroindole, 3 nitropyrrole, 6-(aza)pyrimidine, 2 (Amino)purine, 2,6-(diamino)purine, 5-substituted pyrimidine, N2-substituted purine, N6-substituted purine, 06-substituted purine, substituted 1,2,4-triazole, pyrrolo-pyrimidin-2-one-3-yl, 6-phenyl-pyrrolo-pyrimidin-2-one-3-yl, para-substituted-6-phenyl-pyrrolo-pyrimidin-2-one-3-yl, ortho-substituted-6-phenyl-pyrrolo-pyrimidin-2-one-3-yl, bis-ortho-substituted-6-phenyl-pyrrolo-pyrimidin-2-one-3-yl, para-(aminoalkylhydroxy)-6-phenyl-pyrrolo-pyrimidin-2-one-3-yl, Ortho-(aminoalkylhydroxy)-6-phenyl-pyrrolo-pyrimidin-2-one-3-yl, bis-ortho-(aminoalkylhydroxy)-6-phenyl-pyrrolo-pyrimidin-2-one-3-yl, pyridopyrimidin-3-yl, 2-oxo-7-amino-pyridopyrimidin-3-yl, 2-oxo-pyridopyrimidin-3-yl,or any O-alkylated or N-alkylated derivative thereof. The antisense oligonucleotides of the present invention may be chimeric oligonucleotides. Chimeric antisense compounds of the present invention may be formed as composite structures of two or more oligonucleotides, modified oligonucleotides, oligonucleotides, and / or oligonucleotide mimetics described above. Such compounds are also referred to in the art as hybrid or mixed backbones or chimeras or gapmers. In particular, a gapmer is an oligonucleotide having at least three distinct portions, wherein two of the portions are similar, i.e., contain one or more backbone modifications, and surround a distinct (i.e., not included in the backbone modifications) region. The oligonucleotide may comprise a molecular species at one or both ends, i.e., at the 3' and / or 5' ends. As used herein, a molecular species refers to any compound that is not a naturally occurring or non-naturally occurring nucleotide. Molecular species include spacers, lipids, sterols, lipid moieties such as cholesterol moieties, cholic acid, thioethers such as hexyl-S-tritylthiol, thiocholesterol, aliphatic chains such as dodecanediol or undecyl residues, phospholipids such as di-hexadecyl-rac-glycerol or triethylammonium 1,2-di-O-hexadecyl-rac-glycero-3-H-phosphonate, polyamine or polyethylene glycol chains, or adamantane acetic acid, palmityl moieties, octadecylamine or hexylamino-carbonyl-oxycholesterol moieties, stearyl, C16 alkyl chains, bile acids, cholic acid, taurocholic acid, deoxycholate, oleyl lithocholic acid, oleoyl cholenic acid, glycolipids, phospholipids, sphingolipids, isoprenoids such as steroids, vitamins such as Vitamin E, saturated fatty acids, unsaturated fatty acids, fatty acid esters such as triglycerides, pyrene, porphyrin, texaphyrin, adamantane, acridine, biotin, coumarin, fluorescein, rhodamine, Texas-Red, digoxigenin, dimethoxytrityl,Including but not limited to t-butyldimethylsilyl, t-butyldiphenylsilyl, cyanine dyes (e.g., Cy3 or Cy576), Hoechst 33258 dye, psoralen, or ibuprofen.

[0037] The above nucleotides complementarily bind to the transcript base sequence of the Lrig protein to inhibit the expression of the Lrig protein, thereby inhibiting the differentiation of pre-adipocytes into adipocytes, thereby treating various metabolic diseases including obesity. Those skilled in the art will fully understand that the above nucleotides can inhibit the expression of the Lrig protein, thereby inhibiting the differentiation of pre-adipocytes into adipocytes, thereby treating various metabolic diseases including obesity.

[0038] In another embodiment of the present invention, a pharmaceutical composition is provided, wherein the Lrig protein is at least one selected from the group consisting of Lrig-1, Lrig-2, and Lrig-3.

[0039] The pharmaceutical composition provided in the present invention can treat various metabolic diseases including obesity by inhibiting the differentiation of pre-adipocytes into adipocytes by complementarily binding to the transcript base sequence of Lrig family proteins, i.e., Lrig-1, Lrig-2 and Lrig-3 proteins, thereby inhibiting Lrig protein expression.

[0040] In another embodiment of the present invention, a pharmaceutical composition is provided, wherein the Lrig protein is present on the surface of an adipocyte, a preadipocyte, or a regulatory T cell.

[0041] The pharmaceutical composition provided in the present invention can treat various metabolic diseases including obesity by inhibiting the differentiation of pre-adipocytes into adipocytes by inhibiting the expression of Lrig protein present on the surface of adipocytes, pre-adipocytes, or regulatory T cells.

[0042] In another embodiment of the present invention, a pharmaceutical composition is provided, wherein the metabolic disease is at least one selected from the group consisting of insulin resistance disease, obesity, diabetes, dyslipidemia, liver disease, renal damage, arteriosclerosis, and hypertension.

[0043] The pharmaceutical composition of the present invention is effective in treating and preventing metabolic diseases. Metabolic diseases are a general term for diseases caused by lifestyle factors such as obesity, lack of exercise, and overnutrition. Examples include obesity, hypertension, hyperlipidemia, and diabetes. Metabolic diseases can be accompanied by or cause inflammation and cardiovascular diseases (angina, myocardial infarction, stroke, etc.).

[0044] The term metabolic syndrome or metabolic disease began to be used in the late 1950s and has been in common use since the late 1970s, and is also called metabolic disease. In 1988, Reaven proposed insulin resistance as the cause of metabolic syndrome, and named various abnormal symptoms, namely abdominal obesity, dyslipidemia, hypertension, and fasting hyperglycemia, as Syndrome X. Metabolic syndrome is determined by genetic and environmental factors, and is also influenced by factors such as age, smoking, drinking, diet, and physical activity. Metabolic syndrome is reported to be a major risk factor for diabetes mellitus and cardiovascular disease.

[0045] The main symptoms of the metabolic syndrome include diabetes and obesity due to blood sugar metabolism disorders, increased neutral fat or dyslipidemia due to lipid metabolism disorders, high blood pressure due to increased high-density cholesterol and sodium, and gout due to increased uric acid. It is also reported that various adult diseases such as stroke, arteriosclerosis, and heart disease are caused by metabolic syndrome. According to recent research, it is estimated that about a quarter of adults in the United States have metabolic syndrome. In addition, in Korea, approximately 15-20% of those in their 30s and 30-40% of those over 40 have metabolic syndrome, and the number of patients with metabolic syndrome is reported to be increasing rapidly.

[0046] Diabetes, a representative disease of the metabolic syndrome, is a disease that occurs when there is a problem with the beta cells, which are insulin-secreting cells, or when there is a problem with the function of insulin, blood sugar control is not properly performed, resulting in high blood sugar levels.

[0047] While obesity is a social issue not only for its aesthetic appeal, its most serious problem lies in the serious health risks it can cause, such as metabolic complications like diabetes and hypertension. The symptom associated with this pathological condition is systemic chronic inflammation, which occurs in obese individuals.

[0048] Inflammation is a vital immune response that occurs within the body. When localized, it serves as a crucial defense against external pathogens and viruses. However, when this inflammatory response becomes chronically and systemically overactive due to a disruption in the body's immune balance, it can disrupt metabolic processes within the body.

[0049] In particular, the chronic inflammatory response caused by obesity has been identified as a cause of various metabolic diseases such as diabetes, cardiovascular disease, and arteriosclerosis, and is also the most important factor in defining obesity as a disease. Without the invention of secondary metabolic diseases caused by chronic inflammatory response, obesity would be merely a cosmetic issue. Recently, the World Health Organization (WHO) has classified obesity as a disease, citing the chronic inflammatory response as a reason for secondary metabolic diseases that significantly reduce quality of life, such as diabetes.

[0050] Obesity induces abnormalities in visceral adipose tissue, which secretes endocrine factors such as adiponectin, plasminogen activator inhibitor, monocyte chemotactic protein-1 (MCP-1), tumor necrosis factor-α (TNF-α), and leptin.

[0051] In particular, when monocyte chemotactic protein-1 (MCP-1) and tumor necrosis factor-α (TNF-α) are secreted excessively, immune cells such as macrophages infiltrate adipose tissue and increase the expression of inflammatory cytokines such as interleukin-6 (IL-6) in addition to monocyte chemotactic protein-1 (MCP-1) and tumor necrosis factor-α. As a result, chronic inflammation of adipose tissue occurs, and the chronic inflammatory response reduces insulin sensitivity and induces glucose tolerance, which can lead to the development of diabetes.

[0052] This is a symptom that occurs when the body loses the functions of each body due to the accumulated waste products and toxins that are generated when the balanced metabolism is not properly discharged, and it develops into metabolic syndrome, also known as insulin resistance syndrome. Metabolic syndrome is known to cause damage to the coronary arteries, which can lead to heart disease or stroke, or cause high blood pressure by reducing the kidneys' ability to remove salt, increase the proportion of neutral fat, which is a cause of cardiovascular disease, and increase the risk of blood clotting. It is also known to cause damage to the eyes, kidneys, and nerves due to reduced insulin production in type 2 diabetes.

[0053] The above diabetes refers to a disease that occurs when the amount of insulin secreted is insufficient or the action and function of insulin is not sufficient. When this disease occurs, it causes an abnormal increase in the concentration of glucose in the liver or blood due to excessive breakdown of glycogen, protein, and lipid, resulting in diabetes and ketonuria. It also causes pathological conditions such as blood concentrating state due to electrolyte loss due to abnormalities in water and electrolyte metabolism, as well as circulatory disorders and kidney disorders. Insulin is secreted from the beta cells of the islets of Langerhans in the pancreas, and is secreted when the concentration of blood glucose increases, and is suppressed when it decreases, thereby regulating the appropriate activity of energy sources. This disease is divided into insulin-dependent diabetes (type I) and non-insulin-dependent diabetes (type II). Diabetes can generally be diagnosed by measuring blood glucose concentration, but there are differences depending on the criteria. In humans, diabetes is generally diagnosed when the blood glucose is 200 mg / dL or higher on a regular basis, or 140 mg / dL or higher when fasting. Therefore, lowering the glucose concentration in the blood or liver can help treat or prevent diabetes.

[0054] The nucleic acid molecule of the present invention includes all nucleic acid molecules translated into polynucleotide sequences based on the amino acid sequence of the binding molecule provided in the present invention, as known to those skilled in the art. Therefore, various polynucleotide sequences can be produced by ORF (open reading frame), and all of these are also included in the nucleic acid molecule of the present invention.

[0055] According to another embodiment of the present invention, an expression vector into which the isolated nucleic acid molecule provided by the present invention is inserted is provided.

[0056] In the present invention, the "vector" refers to a nucleic acid molecule capable of transporting another nucleic acid to which it has been linked. One type of vector is a "plasmid," which refers to a circular double-stranded DNA into which additional DNA segments can be ligated. Another type of vector is a phage vector. Another type of vector is a viral vector, into which additional DNA segments can be ligated to the viral genome. Some vectors are capable of autonomous replication in the host cell into which they are introduced (e.g., bacterial vectors are episomal mammalian vectors with a bacterial origin of replication). Other vectors (e.g., non-episomal mammalian vectors) can integrate into the host cell genome upon introduction, thereby replicating along with the host genome. Furthermore, some vectors can direct the expression of genes to which they are linked in an operational manner. Such vectors are referred to herein as "recombinant expression vectors" or simply "expression vectors." In general, expression vectors useful in recombinant DNA techniques often exist in the form of plasmids. In this specification, “plasmid” and “vector” may be used interchangeably, as the plasmid is the most commonly used form of vector.

[0057] In the present invention, specific examples of the expression vector may be selected from the group consisting of commercially widely used pCDNA vectors, F, R1, RP1, Col, pBR322, ToL, Ti vectors; cosmids; phages such as lambda, lambdoid, M13, Mu, p1 P22, Qμμ, T-even, T2, T3, T7, etc.; and plant viruses, but are not limited thereto, and all expression vectors known to those skilled in the art as expression vectors can be used in the present invention, and when selecting an expression vector, it depends on the properties of the target host cell. When introducing a vector into a host cell, it may be performed by calcium phosphate transfection, viral infection, DEAE-dextran controlled transfection, lipofectamine transfection, or electroporation, but is not limited thereto, and those skilled in the art can select and use an introduction method appropriate for the expression vector and host cell to be used. Preferably, the vector contains one or more selectable markers, but this is not limited to this. Selection can be made based on whether a product is produced using a vector that does not contain a selectable marker. The selectable marker is selected based on the target host cell, and since this is done using methods already known to those skilled in the art, the present invention is not limited thereto.

[0058] To facilitate purification of the nucleic acid molecule of the present invention, a tag sequence can be inserted into an expression vector and fused thereto. Such tags include, but are not limited to, a hexahistidine tag, a hemagglutinin tag, a myc tag, or a flag tag, and any tag known to those skilled in the art that facilitates purification can be utilized in the present invention.

[0059] According to another embodiment of the present invention, a host cell line transformed with an expression vector provided by the present invention is provided.

[0060] In the present invention, the term "host cell" includes an individual cell or cell culture that may be or was the recipient of the vector(s) for incorporating the polypeptide insert. Host cells include progeny of a single host cell, and the progeny may not necessarily be completely identical (morphologically or in genomic DNA complement) to the original parent cell due to natural, accidental, or intentional mutation. Host cells include cells that have been transfected in vivo with the polypeptide(s) of the present disclosure.

[0061] In the present invention, the host cell may include a mammalian, plant, insect, fungal or cellular cell, and for example, bacterial cells such as Escherichia coli, Streptomyces, and Salmonella Typhimurium; fungal cells such as yeast cells and Pichia pastoris; insect cells such as Drozophylla and Spodoptera Sf9 cells; animal cells such as CHO (Chinese hamster ovary cells), SP2 / 0 (mouse myeloma), human lymphoblastoid, COS, NSO (mouse myeloma), 293T, Bow melanoma cells, HT-1080, BHK (Baby Hamster Kidney cells), HEK (Human Embryonic Kidney cells), or PERC.6 (human retinal cells); Alternatively, the host cell line may be a plant cell, but is not limited thereto, and any cell known to those skilled in the art that can be used as a host cell line may be used.

[0062] As used herein, the term "combination" or "pharmaceutical combination" refers to a product resulting from the mixing or combining of more than one active ingredient, and including both fixed and non-fixed combinations of the active ingredients. The term "fixed combination" means that the active ingredients, such as a compound of the invention and one or more additional therapeutic agents, are administered to a patient simultaneously in the form of a single entity or dosage. The term "non-fixed combination" means that the active ingredients, such as a compound of the invention and one or more additional therapeutic agents, are administered to a patient as separate entities simultaneously, concurrently, or sequentially without a specific time limit, wherein such administration provides therapeutically effective levels of the active ingredients in the patient's body. The latter also applies to cocktail therapy, for example, the administration of three or more active ingredients.

[0063] The term "composition" or "pharmaceutical composition" as used herein refers to a mixture of at least one compound of the present invention and optionally more than one other pharmaceutically acceptable chemical component, such as a carrier, stabilizer, diluent, dispersant, suspending agent, thickener and / or excipient.

[0064] The terms "treat," "treating," or "treatment," as used herein, refer to a method of alleviating, alleviating, or ameliorating the symptoms of a disease or condition, preventing additional symptoms, ameliorating or preventing the underlying metabolic cause of the symptoms, inhibiting the disease or condition, arresting the development of the disease or condition, relieving the disease or condition, causing regression of the disease or condition, alleviating a condition caused by the disease or condition, or stopping the symptoms of the disease or condition prophylactically and / or therapeutically.

[0065] In addition, siRNA and the like according to the present invention can specifically bind to Lrig protein present on preadipocytes, adipocytes or regulatory T cells to suppress the function of the regulatory T cells, thereby regulating the activity of effector T cells, thereby preventing or treating various diseases, such as metabolic diseases.

[0066] In the present invention, 'obesity' does not simply mean being overweight, but a state in which body fat is excessively accumulated. This means that even if one appears to be of normal weight on the outside, if one's body fat percentage is high, one can be considered obese. The body mass index (BMI) is usually used to determine obesity. A BMI of 23 to 24.9 is considered overweight, 25 to 29.9 is considered mild obesity, 30 to 34.9 is considered moderate obesity, and 35 or more is considered severe obesity. Obesity is caused by a combination of factors rather than a single cause, including poor eating habits, including westernized eating habits, decreased activity level, emotional factors, and genetic factors. Obesity that occurs in this way ultimately increases the risk of chronic diseases such as hyperlipidemia, diabetes, and hypertension.

[0067] In one embodiment of the present invention, the composition has the properties of inhibiting adipogenesis, inhibiting lipid accumulation in the body, or a combination thereof.

[0068] The term "adipogenesis" in this specification refers to the process by which adipocytes are formed from preadipocytes. Transcription factors such as peroxisome proliferator-activated receptor-γ (PPARγ) and C / EBP (CCAAT enhancer-binding protein) are known to be involved in the adipogenesis process. In addition, endocrine products such as insulin, IGF-1, cAMP (cyclic adenosine monophosphate), and glucocorticoids are known to induce adipogenesis. The term "adipogenesis" in this specification is used interchangeably with "adipogenesis" or "lipid differentiation."

[0069] Adipocytes are cells that primarily form adipose tissue, which stores energy as fat. Adipose tissue is a central metabolic organ that regulates energy balance and homeostasis. White adipocytes (WADs) form white adipose tissue and are primarily distributed under the skin, in the abdomen, and around internal organs. White adipose tissue has limited vascularity and primarily functions as a heat insulator, mechanical buffer, and energy source.

[0070] In one specific example of the present invention, the inhibition of adipogenesis is to inhibit the differentiation of preadipocytes into adipocytes in the early stage of the adipogenesis process.

[0071] The adipogenesis process has early, middle, and late stages, and important differentiation regulators are involved in each stage. Representative genes involved in the early stage of adipogenesis are C / EBPβ and C / EBPδ, and they are the first to act in preadipocytes by differentiation-inducing factors. Preadipocytes, whose growth was inhibited in a post-confluent state, begin the cell cycle to differentiate into adipocytes by differentiation-inducing factors. Genes expressed in the middle stage are PPARγ and C / EBPα, and during this period, lipid droplets begin to form and lipids accumulate. Genes expressed in the late stage include FABP4 and FASN, and this is the final stage of adipocyte differentiation, the stage where mature adipocytes are created.

[0072] The early, middle, and late stages of the adipogenesis process can be distinguished according to the passage of time. When the time required for the entire adipogenesis process is 7 days, the early stage can be 0 to 2 days, the middle stage can be 2 to 4 days, and the late stage can be 4 to 7 days. In one embodiment of the present invention, when YM976 was treated on days 0 to 2 of the adipogenesis process, it exhibited a superior effect in inhibiting adipogenesis compared to when YM976 was treated at other stages.

[0073] In one embodiment of the present invention, the composition inhibits the expression of a gene or protein selected from the group consisting of PPARγ (peroxisome proliferator-activated receptor-γ), C / EBPα (CCAAT enhancer-binding protein α), C / EBPβ, FASN (fatty acid synthase), FABP4 (fatty acid binding protein 4), and a combination thereof in preadipocytes.

[0074] Body fat is an essential nutrient required by all physiological organs and plays a crucial role in maintaining energy homeostasis. However, Westernized eating and lifestyle habits resulting from social and economic development have led to an imbalance in energy intake and expenditure, leading to obesity. This obesity is known to be a direct and indirect cause of various metabolic and cardiovascular diseases, such as diabetes, hypertension, hyperlipidemia, arteriosclerosis, heart disease, and stroke. The accumulation of body fat, which is the cause of obesity, occurs through the production and secretion of hormones (adipokines) secreted by body fat cells, which induce differentiation of adipocytes, leading to hypertrophy and hyperplasia, and the accumulation of fat globules generated by lipogenesis. In addition, adipocyte hyperplasia is induced by the proliferation and differentiation process of preadipocytes, and in preadipocytes, C / EBPβ and C / EBPγ are activated during the early differentiation by stimulation with dexamethasone (DEX), 3-isobutyl-1-methylxanthine (IBMX), insulin, etc., and these interact to regulate the expression of PPARγ and C / EBPα. In addition, PPARγ and C / EBPα are expressed in the early stage of differentiation and participate in adipocyte differentiation, and they induce the expression of various adipogenic genes in the later stage of differentiation. Therefore, inhibition of the above mechanism is being studied as an important target in anti-obesity research.

[0075] In the present invention, 'metabolic disease' means a condition or disease closely related to obesity or caused by obesity, and specifically may be at least one selected from the group consisting of fatty liver, type 2 diabetes, hyperlipidemia, cardiovascular disease, and arteriosclerosis.

[0076] In the present invention, fatty liver refers to a condition or disease in which fat accumulates in excessive amounts in liver cells due to a disorder of fat metabolism in the liver.

[0077] In the present invention, hyperlipidemia refers to a condition or disease in which the concentration of fat components in the blood, particularly cholesterol and triglycerides, is higher than the normal level, and is used in a broad sense to include all conditions in which it is required to lower the concentration of lipids in the blood.

[0078] In this specification, arteriosclerosis refers to a condition or disease in which the arterial wall thickens and loses elasticity, thereby reducing blood circulation to organs and tissues in the body. It also includes "atherosclerosis," which refers to a condition or disease in which fat, cholesterol, and other substances are deposited on the inner wall of the artery to form plaque, narrowing the inner lumen and reducing blood circulation. Atherosclerosis can occur anywhere in the body, and when it occurs in the blood vessels of the heart, it can cause coronary artery disease such as angina pectoris and myocardial infarction, and when it occurs in the brain, it can cause cerebral infarction, and when it occurs in the kidney, it can cause renal failure, etc.

[0079] Meanwhile, the strength, binding and expression of Foxp3, T cell receptor (TCR) of Tregs, ability to produce anti-inflammatory cytokines such as interleukin-10 (IL-10) and IL-35, cytotoxic T-lymphocyte-associated protein 4 (CTLA-4), programmed cell death protein 1 (PD-1), CD39 and CD73 are known to play important roles in immune regulation.

[0080] Recent studies have revealed that the expression levels of these proteins are related to metabolism. That is, after Treg activation, mTOR signaling is upregulated to induce lipid synthesis, mevalonate metabolism, and mitochondrial function; both CDTLA-4 and PD-1 have been described to block glycolysis, which promotes lipolysis and fatty acid oxidation by PD-1 signaling; CD39, which converts adenosine triphosphate (ATP) and adenosine diphosphate (ADP) to adenosine monophosphate (AMP), and CD73, which converts AMP to adenosine, abrogate ATP effects such as P2 receptor-mediated cytotoxicity and ATP-induced maturation of dendritic cells.

[0081] Alterations in Treg numbers and function have been widely documented in human autoimmune, infectious, and allergic diseases, as well as in cancer. Furthermore, reduced Treg levels have been reported in patients with type 2 diabetes and contribute to both hyperglycemia and high-density lipoprotein concentrations in the blood. Circulating adipose and visceral adipose Treg are decreased in obese individuals and inversely correlate with measures of adiposity, inflammation, and glucose tolerance, potentially identifying individuals at increased metabolic and cardiovascular risk. PPARγ signaling for Treg energy homeostasis maintains the inflammatory state of adipose tissue and insulin sensitivity in lean adipose tissue, whereas dysfunction impairs insulin sensitivity. Interestingly, in animal models, attenuation of acute graft-versus-host disease and multiple sclerosis was observed, but Treg frequency and suppressive capacity remained unchanged after INSR silencing.

[0082] Therefore, it can be seen that changes in the number and function of Tregs are closely related to metabolic diseases including autoimmune diseases, cancer, as well as type 2 diabetes, obesity, and cardiovascular diseases. Antisense oligonucleotides that complementarily bind to the Lrig protein family and inhibit the expression of Lrig proteins, which can change the number or function of Tregs, can prevent or treat metabolic diseases including type 2 diabetes, obesity, and cardiovascular diseases.

[0083] Meanwhile, in the present invention, “prevention” may include, without limitation, any act of blocking, suppressing or delaying the symptoms of a disease by using the pharmaceutical composition of the present invention.

[0084] Additionally, in the present invention, “treatment” may include, without limitation, any act that improves or benefits the symptoms of a disease by using the pharmaceutical composition of the present invention.

[0085] In the present invention, the pharmaceutical composition may be characterized as being in the form of a capsule, tablet, granule, injection, ointment, powder, or beverage, and the pharmaceutical composition may be characterized as being for human use.

[0086] In the present invention, the pharmaceutical composition is not limited to these, but may be formulated and used in the form of oral dosage forms such as powders, granules, capsules, tablets, aqueous suspensions, etc., external preparations, suppositories, and sterile injection solutions, respectively, according to conventional methods. The pharmaceutical composition of the present invention may include a pharmaceutically acceptable carrier. Pharmaceutically acceptable carriers may include binders, lubricants, disintegrants, excipients, solubilizers, dispersants, stabilizers, suspending agents, coloring agents, fragrances, etc. for oral administration, and buffers, preservatives, analgesics, solubilizers, isotonic agents, stabilizers, etc. for injections, and bases, excipients, lubricants, preservatives, etc. for topical administration. The formulation of the pharmaceutical composition of the present invention may be prepared in various ways by mixing with the pharmaceutically acceptable carriers described above. For example, for oral administration, it can be manufactured in the form of tablets, troches, capsules, elixirs, suspensions, syrups, wafers, etc., and for injections, it can be manufactured in the form of unit dose ampoules or multiple doses. In addition, it can be formulated in the form of solutions, suspensions, tablets, capsules, sustained-release preparations, etc.

[0087] Meanwhile, examples of carriers, excipients, and diluents suitable for formulation include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, microcrystalline cellulose, polyvinylpyrrolidone, water, methyl hydroxybenzoate, propyl hydroxybenzoate, talc, magnesium stearate, or mineral oil. In addition, fillers, anti-coagulants, lubricants, wetting agents, fragrances, emulsifiers, preservatives, and the like may be additionally included.

[0088] The routes of administration of the pharmaceutical composition of the present invention include, but are not limited to, oral, intravenous, intramuscular, intraarterial, intramedullary, intrathecal, intracardiac, transdermal, subcutaneous, intraperitoneal, intranasal, enteral, topical, sublingual, or rectal. Oral or parenteral administration is preferred.

[0089] The term "parenteral" in the present invention includes subcutaneous, intradermal, intravenous, intramuscular, intraarticular, intrasynovial, intrasternal, intrathecal, intralesional, and intracranial injection or infusion techniques. The pharmaceutical composition of the present invention may also be administered in the form of a suppository for rectal administration.

[0090] The pharmaceutical composition of the present invention may vary depending on various factors including the activity of the specific compound used, age, body weight, general health, sex, dosage form, administration time, administration route, excretion rate, drug combination, and severity of the specific disease to be prevented or treated, and the dosage of the pharmaceutical composition may vary depending on the patient's condition, body weight, degree of disease, form of medicine, administration route, and period, but may be appropriately selected by those skilled in the art, and may be administered at 0.0001 to 50 mg / kg or 0.001 to 50 mg / kg per day. Administration may be administered once a day or divided into several times. The dosage does not limit the scope of the present invention in any way. The pharmaceutical composition according to the present invention may be formulated as a pill, a dragee, a capsule, a liquid, a gel, a syrup, a slurry, or a suspension.

[0091] In the present invention, the "subject" refers to a subject suspected of having a metabolic disease, and the subject suspected of having a metabolic disease refers to a mammal including a human, a rat, a livestock, etc. that has developed or may develop the disease. However, a subject that can be treated with an antisense oligonucleotide that complementarily binds to the Lrig protein family of the present invention and inhibits the expression of the Lrig protein is included without limitation.

[0092] The method of the present invention may include administering a pharmaceutically effective amount of an antisense oligonucleotide or the like that complementarily binds to the Lrig protein family and inhibits the expression of the Lrig protein. The appropriate total daily dosage may be determined by the treating physician within the scope of sound medical judgment, and may be administered once or in several divided doses. However, for the purposes of the present invention, it is preferable that a specific therapeutically effective amount for a specific patient be applied differently depending on various factors, including the type and degree of response to be achieved, the specific composition including whether other agents are used in some cases, the patient's age, weight, general health, sex, and diet, the time of administration, the route of administration, and the excretion rate of the composition, the treatment period, drugs used together or concurrently with the specific composition, and similar factors well known in the medical field.

[0093] Meanwhile, but not limited thereto, the method for preventing or treating the disease may be a combination therapy further comprising administering a compound or substance having therapeutic activity against one or more diseases.

[0094] In the present invention, the term "combination" should be understood to refer to simultaneous, separate, or sequential administration. If the administration is sequential or separate, the interval between administrations of the secondary components should be such that the beneficial effects of the combination are not lost.

[0095] In the present invention, the administration dose of the antibody or antibody-drug conjugate may be about 0.0001 μg to 500 mg per 1 kg of patient body weight, but is not limited thereto.

[0096] In one embodiment of the present invention, a composition for diagnosing a metabolic disease is provided, comprising an agent for measuring the expression level of a Lrig protein or a fragment thereof; or a gene encoding the same.

[0097] The diagnostic composition of the present invention may include, but is not limited to, a formulation for measuring the expression level of a gene encoding the Lrig-1 protein or its extracellular domain present on the surface of an activated regulatory T cell, i.e., an activated regulatory T cell having an inhibitory ability of an effector T cell; or a gene encoding the same.

[0098] The diagnostic composition of the present invention may further include an agent for measuring the expression level of one or more proteins selected from the group consisting of CD25, TIGIT, LAG3, CTLA-4, GITR, OX40, ICOS, PD-1, TIM-3, CCR4, FR4, CD15s, PI-16 and Lrig-1 proteins present on the surface of T cells; or a gene encoding the same. In this way, when the expression levels of various proteins present on the surface of T cells; or the genes encoding the same are further measured, a remarkable synergistic effect can be exhibited in diagnosing the desired disease compared to when the Lrig-1 protein or its extracellular domain; or the gene encoding the same is measured alone.

[0099] The agent for measuring the expression level of the Lrig-1 protein or its extracellular domain of the present invention and the expression level of one or more proteins selected from the group consisting of CD25, TIGIT, LAG3, CTLA-4, GITR, OX40, ICOS, PD-1, TIM-3, CCR4, FR4, CD15s, PI-16 and Lrig-1 protein is not particularly limited, but may include, for example, one or more selected from the group consisting of antibodies, oligopeptides, ligands, PNA (peptide nucleic acid) and aptamers that specifically bind to the proteins.

[0100] The "PNA (Peptide Nucleic Acid)" in the present invention refers to an artificially synthesized polymer similar to DNA or RNA, and was first introduced in 1991 by Professors Nielsen, Egholm, Berg, and Buchardt of the University of Copenhagen, Denmark. While DNA has a phosphate-ribose sugar backbone, PNA has a repeated N-(2-aminoethyl)-glycine backbone linked by peptide bonds, which greatly increases its binding affinity and stability to DNA or RNA, and is used in molecular biology, diagnostic analysis, and antisense therapy. PNA is described in detail in the literature [Nielsen PE, Egholm M, Berg RH, Buchardt O (December 1991). "Sequence-selective recognition of DNA by strand displacement with a thymine-substituted polyamide". Science 254 (5037): 1497-1500].

[0101] In the present invention, the "aptamer" is an oligonucleotide or peptide molecule, and the general contents of the aptamer are disclosed in detail in the literature [Bock LC et al., Nature 355(6360):5646(1992); Hoppe-Seyler F, Butz K "Peptide aptamers: powerful new tools for molecular medicine". J Mol Med. 78(8):42630(2000); Cohen BA, Colas P, Brent R. "An artificial cell-cycle inhibitor isolated from a combinatorial library". Proc Natl Acad Sci USA. 95(24): 142727(1998)].

[0102] A preparation for measuring the expression level of a gene encoding the Lrig-1 protein of the present invention (i.e., LAIR1) or a gene encoding the extracellular domain of the Lrig-1 protein, and a gene encoding at least one protein selected from the group consisting of CCD25, TIGIT, LAG3, CTLA-4, GITR, OX40, ICOS, PD-1, TIM-3, CCR4, FR4, CD15s, PI-16 and Lrig-1 protein may include at least one selected from the group consisting of primers, probes, LNAs and antisense nucleotides that specifically bind to the gene.

[0103] The "primer" of the present invention is a fragment that recognizes a target gene sequence, and includes a pair of forward and reverse primers, but is preferably a pair of primers that provide analysis results with specificity and sensitivity. When the nucleic acid sequence of the primer is a sequence that does not match the non-target sequence present in the sample, and thus the primer only amplifies the target gene sequence containing the complementary primer binding site and does not cause non-specific amplification, high specificity can be imparted.

[0104] The "probe" of the present invention refers to a substance that can specifically bind to a target substance to be detected in a sample, and refers to a substance that can specifically confirm the presence of the target substance in the sample through the binding. The type of the probe is not limited to a substance commonly used in the art, but is preferably PNA (peptide nucleic acid), LNA (locked nucleic acid), peptide, polypeptide, protein, RNA, or DNA, and most preferably PNA. More specifically, the probe includes a biomaterial derived from or similar to a living organism or manufactured in vitro, and may be, for example, an enzyme, a protein, an antibody, a microorganism, an animal or plant cell and organ, a nerve cell, DNA, and RNA. DNA includes cDNA, genomic DNA, and oligonucleotides, RNA includes genomic RNA, mRNA, and oligonucleotides, and examples of proteins include antibodies, antigens, enzymes, peptides, etc.

[0105] In the present invention, the "LNA (Locked nucleic acids)" refers to a nucleic acid analog containing a 2'-O, 4'-C methylene bridge [J Weiler, J Hunziker and J Hall Gene Therapy (2006) 13, 496.502]. LNA nucleosides contain common nucleic acid bases of DNA and RNA and can form base pairs according to the Watson-Crick base pairing rule. However, due to the 'locking' of the molecule caused by the methylene bridge, LNA cannot form an ideal shape in Watson-Crick binding. When LNA is included in a DNA or RNA oligonucleotide, LNA can pair with a complementary nucleotide chain more quickly and increase the stability of the double helix.

[0106] In the present invention, the term "antisense" refers to an oligomer having a sequence of nucleotide bases and an intersubunit backbone that allows the antisense oligomer to hybridize with a target sequence within RNA by Watson-Crick base pairing, typically allowing the formation of an mRNA and RNA:oligomer heteroduplex within the target sequence. The oligomer may have exact sequence complementarity or approximate sequence complementarity to the target sequence.

[0107] Since the information on the Lrig1 protein according to the present invention or the gene encoding it (LRIG1) is known, a person skilled in the art will be able to easily design a primer, probe or antisense nucleotide that specifically binds to the gene encoding the protein based on this.

[0108] According to another embodiment of the present invention, there is provided a diagnostic kit for metabolic diseases comprising the diagnostic composition provided by the present invention.

[0109] The kit of the present invention may be, but is not limited to, an RT-PCR kit, a DNA chip kit, an ELISA kit, a protein chip kit, a rapid kit, or an MRM (Multiple reaction monitoring) kit.

[0110] The kit of the present invention may further comprise one or more other component compositions, solutions or devices suitable for the analysis method.

[0111] The kit of the present invention may further comprise, for example, essential elements necessary for performing a reverse transcription polymerase reaction. The reverse transcription polymerase reaction kit comprises a primer pair specific for a gene encoding a marker protein. The primer pair is a nucleotide sequence having a sequence specific to the nucleic acid sequence of the gene, and may have a length of, for example, about 7 bp to 50 bp, or about 10 bp to 30 bp. It may also comprise a primer pair specific for the nucleic acid sequence of a control gene. In addition, the reverse transcription polymerase reaction kit may further comprise a test tube or other appropriate container, a reaction buffer (with various pH and magnesium concentrations), deoxynucleotides (dNTPs), an enzyme such as Taq polymerase and reverse transcriptase, DNase, RNase inhibitor DEPC-water, sterile water, etc.

[0112] The kit of the present invention may include essential elements necessary for performing a DNA chip. The DNA chip kit may include a substrate to which cDNA or oligonucleotides corresponding to a gene or fragment thereof are attached, and reagents, preparations, enzymes, etc. for producing a fluorescently labeled probe. The substrate may also include cDNA or oligonucleotides corresponding to a control gene or fragment thereof.

[0113] The kit of the present invention may include the essential elements necessary for performing an ELISA. The ELISA kit includes an antibody specific for the protein. The antibody has high specificity and affinity for the marker protein and little cross-reactivity with other proteins, and may be a monoclonal antibody, a polyclonal antibody, or a recombinant antibody. The ELISA kit may also include an antibody specific for a control protein. In addition, the ELISA kit may include reagents capable of detecting bound antibodies, such as labeled secondary antibodies, chromophores, enzymes (e.g., conjugated to antibodies), and their substrates or other substances capable of binding to antibodies.

[0114] According to another embodiment of the present invention, a method for providing information for diagnosing a metabolic disease is provided, comprising: a step of determining that a metabolic disease has developed or is likely to develop if the expression level of the measured Lrig protein or a gene encoding the same is higher than that of a control group in a biological sample isolated from a target individual;

[0115] The step of measuring the expression level of the present invention may further measure the expression level of one or more proteins selected from the group consisting of CD25, TIGIT, LAG3, CTLA-4, GITR, OX40, ICOS, PD-1, TIM-3, CCR4, FR4, CD15s, PI-16 and Lrig-1 proteins present on the surface of T cells; or a gene encoding the same. In this way, when the expression levels of various proteins present on the surface of T cells; or the genes encoding the same are further measured, a remarkable synergistic effect can be exhibited in diagnosing the desired disease compared to when the Lrig-1 protein; or the gene encoding the same is measured alone.

[0116] The “target subject” of the present invention refers to a subject whose onset of the disease is uncertain, and has a high possibility of developing the disease.

[0117] The "biological sample" of the present invention means any material, biological fluid, tissue or cell obtained from or derived from an individual, for example, whole blood, leukocytes, peripheral blood mononuclear cells, buffy coat, plasma, serum, sputum, tears, mucus, nasal washes, nasal aspirate, breath, urine, semen, saliva, peritoneal washings, pelvic fluids, cystic fluid, meningeal fluid, amniotic fluid, glandular fluid, pancreatic fluid, lymph fluid, pleural fluid, nipple aspirate, It may include, but is not limited to, bronchial aspirate, synovial fluid, joint aspirate, organ secretions, cells, cell extracts, or cerebrospinal fluid.

[0118] The Lrig-1 protein of the present invention; or its extracellular domain may be expressed on the cell surface of T cells, particularly regulatory T cells, for example, activated regulatory T cells, i.e., regulatory T cells with an activated suppressive capacity of effector T cells.

[0119] The method for measuring or comparing the expression level of the Lrig-1 protein or its extracellular domain and one or more proteins selected from the group consisting of CD25, TIGIT, LAG3, CTLA-4, GITR, OX40, ICOS, PD-1, TIM-3, CCR4, FR4, CD15s, PI-16 and Lrig-1 protein of the present invention includes protein chip analysis, immunoassay, ligand binding assay, MALDI-TOF (Matrix Assisted Laser Desorption / Ionization Time of Flight Mass Spectrometry) analysis, SELDI-TOF (Surface Enhanced Laser Desorption / Ionization Time of Flight Mass Spectrometry) analysis, radioimmunoassay, radioimmunodiffusion, Okteroni immunodiffusion, rocket immunoelectrophoresis, tissue immunostaining, complement fixation assay, two-dimensional electrophoresis, liquid chromatography-mass spectrometry, These include, but are not limited to, LC-MS (Liquid Chromatography-Mass Spectrometry / Mass Spectrometry), LC-MS / MS (Liquid Chromatography-Mass Spectrometry), Western blotting, or ELISA (Enzyme linked immunosorbent assay).

[0120] In order to confirm the presence and expression level of a gene encoding the Lrig-1 protein or its extracellular domain of the present invention and a gene encoding at least one protein selected from the group consisting of CD25, TIGIT, LAG3, CTLA-4, GITR, OX40, ICOS, PD-1, TIM-3, CCR4, FR4, CD15s, PI-16 and Lrig-1 protein, analytical methods for measuring the expression level of the gene include, but are not limited to, reverse transcription polymerase chain reaction (RT-PCR), competitive RT-PCR, real-time RT-PCR, RNase protection assay (RPA), Northern blotting or DNA chip.

[0121] The present invention may include a step of predicting that the likelihood of developing a metabolic disease is high when the expression level of the Lrig-1 protein or its extracellular domain, or a gene encoding it, measured for a biological sample of the subject of interest is changed compared to a normal control group.

[0122] In the present invention, the "control group" may be a normal control group, and more specifically, may be obtained from a serum sample of a patient confirmed to not have a bone-related disease, and may be an average or median value of the expression level of TregL1 of a patient who underwent a bone density test, ultrasound, and CT and was finally confirmed to not have a bone-related disease. The expression level of a marker protein or a gene encoding the same in the control group can be compared with the expression level of a marker protein or a gene encoding the same in a biological sample derived from a patient with a bone-related disease to be analyzed, and a bone-related disease or a metabolic disease can be diagnosed by determining whether there is a significant change in the expression level.

[0123] In the present invention, "high level" means a significantly higher expression level than that of a normal control group to a measurable degree, and specifically, for example, it means a case where it is expressed by 20% or more than that of a normal control group, more specifically, it may mean a case where it is expressed by 30% or more, even more specifically, it may mean a case where it is expressed by 40% or more, and most specifically, it may mean a case where it is expressed by 50% or more, but is not limited thereto.

[0124] The method may further include a step of predicting that there is a high possibility of developing a metabolic disease when the expression level of one or more proteins selected from the group consisting of CD25, TIGIT, LAG3, CTLA-4, GITR, OX40, ICOS, PD-1, TIM-3, CCR4, FR4, CD15s, PI-16 and Lrig-1 proteins present on the surface of the T cells of the present invention; or a gene encoding the same, is changed (increased or decreased) compared to a normal control group.

[0125] In another specific example of the present invention, the method may include a step of predicting that the likelihood of developing a metabolic disease is high when the expression level of the Lrig-1 protein or a gene encoding the same is reduced compared to a normal control group.

[0126] Furthermore, in the present invention, when the expression level of the Lrig-1 protein or its extracellular domain; or the gene encoding it is measured for a biological sample of the desired individual as described above, and thus the possibility of developing a metabolic disease is predicted or diagnosed, the method may further include a step of administering a drug for the disease to the desired individual.

[0127] In the present invention, the term “diagnosis” means confirming the presence or characteristics of a pathological condition, and more specifically, determining a metabolic disease.

[0128] According to another embodiment of the present invention, the present invention relates to a diagnostic device for diagnosing metabolic diseases.

[0129] The measuring unit of the diagnostic device of the present invention can measure the expression level of TregL1 using a preparation for measuring the expression level of TregL1 for a biological sample obtained from a target individual.

[0130] In the present invention, the object of the present invention may be selected from the group consisting of humans, rats, mice, guinea pigs, hamsters, rabbits, monkeys, dogs, cats, cows, horses, pigs, sheep, and goats, and may specifically be humans, but is not limited thereto.

[0131] In the present invention, the biological sample is whole blood, leukocytes, peripheral blood mononuclear cells, buffy coat, plasma, serum, sputum, tears, mucus, nasal washes, nasal aspirate, breath, urine, semen, saliva, peritoneal washings, ascites, cystic fluid, meningeal fluid, amniotic fluid, glandular fluid, pancreatic fluid, lymph fluid, pleural fluid, nipple aspirate, bronchial aspirate, synovial fluid, joint aspirate. The present invention may be, but is not limited to, one or more selected from the group consisting of aspirate, organ secretions, cells, cell extracts, and tissues.

[0132] The agent used in the measurement unit of the diagnostic device of the present invention may be an agent for measuring the expression level of TregL1. More specifically, it may include one or more selected from the group consisting of primers, probes, and antisense nucleotides that specifically bind to the gene.

[0133] By checking the expression level of the gene using the preparation in the measuring unit of the diagnostic device of the present invention, the presence or absence of a metabolic disease can be predicted.

[0134] The diagnostic device for metabolic disease of the present invention may further include a detection unit that predicts and outputs the presence or absence of a metabolic disease in the target individual from the expression level of the gene obtained from the measurement unit.

[0135] In the present invention, the detection unit can diagnose a metabolic disease by generating information and classifying it as having developed or having a high possibility of developing a metabolic disease when the expression level of the gene obtained from the measurement unit is higher than the expression level of TregL1 measured in the control group.

[0136] In addition, in the present invention, the detection unit can diagnose a metabolic disease by generating information and classifying it as having developed or having a high possibility of developing a metabolic disease when the expression level of the gene obtained from the measurement unit is higher than the expression level of TregL1 measured in the control group.

[0137] In another embodiment of the present invention, a diagnostic composition is provided, wherein the agent for measuring the expression level of the protein comprises at least one selected from the group consisting of an antibody, an oligopeptide, a ligand, a peptide nucleic acid (PNA), and an aptamer that specifically binds to the protein.

[0138] In another embodiment of the present invention, a diagnostic composition is provided, wherein the agent for measuring the expression level of the gene comprises at least one selected from the group consisting of a primer, a probe, and an antisense nucleotide that specifically bind to the gene.

[0139] In another embodiment of the present invention, a diagnostic composition is provided, wherein the composition is for application to a biological sample isolated from a desired individual.

[0140] In another embodiment of the present invention, the biological sample is whole blood, leukocytes, peripheral blood mononuclear cells, buffy coat, plasma, serum, sputum, tears, mucus, nasal washes, nasal aspirate, breath, urine, semen, saliva, peritoneal washings, ascites, cystic fluid, meningeal fluid, amniotic fluid, glandular fluid, pancreatic fluid, lymph fluid, pleural fluid, nipple aspirate, bronchial aspirate, synovial fluid, joint A diagnostic composition is provided, comprising at least one selected from the group consisting of joint aspirate, organ secretions, cells, cell extracts, and cerebrospinal fluid.

[0141] In one embodiment of the present invention, a diagnostic kit comprising the diagnostic composition is provided.

[0142] In another embodiment of the present invention, a diagnostic kit is provided, wherein the kit is an RT-PCR kit, a DNA chip kit, an ELISA kit, a protein chip kit, a rapid kit, or an MRM (Multiple reaction monitoring) kit.

[0143] In another embodiment of the present invention, the method for measuring the expression level of the protein is provided by protein chip analysis, immunoassay, ligand binding assay, MALDI-TOF (Matrix Assisted Laser Desorption / Ionization Time of Flight Mass Spectrometry) analysis, SELDI-TOF (Sulface Enhanced Laser Desorption / Ionization Time of Flight Mass Spectrometry) analysis, radioimmunoassay, radioimmunodiffusion, aukteroni immunodiffusion, rocket immunoelectrophoresis, tissue immunostaining, complement fixation assay, two-dimensional electrophoresis analysis, liquid chromatography-mass spectrometry (LC-MS), liquid chromatography-mass spectrometry / mass spectrometry (LC-MS), Western blotting, enzyme linked immunosorbent assay (ELISA), or multiple reaction monitoring (MRM) method.

[0144] In another embodiment of the present invention, the expression level of the gene is measured by a method including reverse transcription polymerase chain reaction (RT-PCR), competitive reverse transcription polymerase reaction (Competitive RT-PCR), real-time RT-PCR, RNase protection assay (RPA), Northern blotting, or DNA chip.

[0145] In one embodiment of the present invention, a diagnostic device for a metabolic disease is provided, comprising: (a) a measuring unit for measuring the expression level of an Lrig protein or a gene encoding the protein for a biological sample obtained from a target individual; and (b) a detection unit for outputting the presence or absence of a metabolic disease from the expression level of the gene or the protein encoded by the gene measured by the measuring unit.

[0146]

[0147] The Lrig inhibitor, Lrig expression inhibitor, or antisense oligonucleotide complementarily binding to the Lrig protein according to the present invention specifically binds to the Lrig protein present on preadipocytes, adipocytes, or regulatory T cells, thereby regulating the function of the regulatory T cells, and as a result, regulating the activity of effector T cells, so that it can be used very effectively for the prevention, improvement, or treatment of various diseases.

[0148]

[0149] Figure 1 is the experimental protocol.

[0150] Figure 2 shows the FACS results confirming the level of Lrig-1 protein expression after Lrig-1 knockdown.

[0151] Figure 3 shows the FACS results confirming the level of Lrig-1 protein expression and whether preadipocytes were differentiated into adipocytes after Lrig-1 knockdown.

[0152] Figure 4 shows the kinetics of Lrig1 expression.

[0153] Figure 5 shows the results of confirming the differentiation of 3T3-L1 cells.

[0154] Figure 6a shows the results of confirming the differentiation of 3T3-L1 cells using a 100x microscope.

[0155] Figure 6b shows the results of confirming the differentiation of 3T3-L1 cells using a 400x microscope.

[0156] Figure 7 shows the results of flow cytometry analysis confirming Lrig-1 knockdown.

[0157] Figure 8 shows the results of confirming the expression of Lrig-1 protein when Lrig-1 is knocked down.

[0158] Figure 9 shows the results of analyzing lipid droplets when Lrig-1 was knocked down.

[0159] Figure 10 shows the results of confirming FABP4 expression through Western blot.

[0160] Figure 11 shows the results of confirming the relative FABP4 protein expression over time.

[0161] Figure 12 shows the results of examining the body weight changes of Lrig-1 knockout mice for males and females.

[0162] Figure 13 shows the results of examining the change in body weight of the first confirmed Lrig-1 knockout male mouse by dividing it into cases where the genotype was hetero or homo.

[0163] Figure 14 shows the results of examining changes in adipocytes in Lrig-1 knockout male mice, first identified, by dividing them into visceral fat and subcutaneous fat. In the figure, PGF represents eWAT, i.e., visceral fat, and SCF represents iWAT, i.e., subcutaneous fat.

[0164] Figure 15 shows the results of examining the change in body weight of the first confirmed Lrig-1 knockout female mouse by dividing it into cases where the genotype was hetero or homo.

[0165] Figure 16 shows the first confirmed changes in adipocytes in Lrig-1 knockout female mice, divided into visceral fat and subcutaneous fat. In the figure, PGF represents eWAT, i.e., visceral fat, and SCF represents iWAT, i.e., subcutaneous fat.

[0166] Figure 17 shows the results of examining the change in body weight of Lrig-1 knockout male mice confirmed for the second time, divided into cases where the genotype was hetero or homo.

[0167] Figure 18 shows the results of examining changes in adipocytes in the second confirmed Lrig-1 knockout male mouse, divided into visceral fat and subcutaneous fat. In the figure, PGF represents eWAT, i.e., visceral fat, SCF represents iWAT, i.e., subcutaneous fat, WAT represents white adipose tissue, i.e., white adipocytes, and BAT represents brown adipose tissue, i.e., brown adipocytes.

[0168] Figure 19 shows the results of examining the change in body weight of Lrig-1 knockout female mice confirmed for the second time, divided into cases where the genotype was hetero or homo.

[0169] Figure 20 shows the results of examining changes in adipocytes in Lrig-1 knockout female mice, divided into visceral fat and subcutaneous fat, for the second time. In the figure, PGF represents eWAT, i.e., visceral fat, and SCF represents iWAT, i.e., subcutaneous fat.

[0170] Figure 21 illustrates the experimental process for differentiating 3T3 cell lines to express Lrig-1.

[0171] Figure 22 shows the expression level of Lrig-1 confirmed through flow cytometry analysis.

[0172] Figure 23 shows the expression level of Lrig-1 over time in differentiated adipocytes.

[0173] Figure 24 shows the expression level of Lrig-1 over time in differentiated adipocytes.

[0174]

[0175] Hereinafter, the present invention will be described in more detail through examples. These examples are intended solely to illustrate the present invention more specifically, and it will be apparent to those skilled in the art that the scope of the present invention is not limited by these examples, in accordance with the gist of the present invention.

[0176] [Example]

[0177] [Example 1] Lrig-1 knockdown using siRNA in 3T3-L1 cells (Lrig-1 knockdown in 3T3-L1)

[0178] [Example 1-1] Induction for 3T3-L1 differentiation

[0179] First, 3T3-L1 cells (pre-adipocytes) were seeded at 1 x 10 in a 24-well plate. 5 / well and plated in DMI medium, i.e., Dulbecco's modified Eagle's medium (DMEM) from Cytiva® Cat. No. SH30243.01 supplemented with 10% fetal bovine serum from Cytiva® Cat. No. SV30207.02, 1% penicillin-streptomycin from Thermo Fisher® Cat. No. 15140122, 1 uM dexamethasone from Sigma® Cat. No. D4902, 520 μM 3-isobutyl-1-methylxanthine (IBMX) from Sigma® Cat. No. 228420010 and 100 μM 3-isobutyl-1-methylxanthine (IBMX) from Sigma® Cat. No. After culturing for 3 days in a medium containing 1 μM of insulin from I5500, the cells were cultured for 3 days in an insulin medium (i.e., Dulbecco's modified Eagle's medium (DMEM) from Cytiva® Cat. No. SH30243.01 supplemented with 10% fetal bovine serum from Cytiva® Cat. No. SV30207.02, 1% penicillin-streptomycin from Thermo Fisher® Cat. No. 15140122, and 167 nM of insulin from Sigma® Cat. No. I5500). Thereafter, the cells were cultured for 3 days in a complete DMEM medium (i.e., Dulbecco's modified Eagle's medium (DMEM) from Cytiva® Cat. No. SH30243.01 supplemented with 10% fetal bovine serum from Cytiva® Cat. No. SV30207.02, 1% penicillin-streptomycin from Thermo Fisher® Cat. No. 15140122, and 167 nM of insulin from Sigma® Cat. No. I5500). The cells were cultured for 6 days in a medium containing 10% fetal bovine serum (SV30207.02) and 1% penicillin-streptomycin (Thermo Fisher® Cat. No. 15140122). The detailed experimental protocol is shown in Fig. 1, and the medium conditions are shown in Table 1 below.

[0180] ReagentsConcentrationCatalog No.Completed DMEM (DMEM)Dulbecco's modified Eagle's medium (DMEM) Cytiva, SH30243.01Fetal bovine serum10%Cytiva, SV30207.02Penicillin-Streptomycin1%Thermo Fisher, 15140122Maintain media (MM)Dulbecco's modified Eagle's medium (DMEM) Cytiva, SH30243.01Bovine Calf serum10%Cytiva, SH30073.03HIPenicillin-Streptomycin1%Thermo Fisher, 15140122DMIDulbecco's modified Eagle's medium (DMEM) Cytiva, SH30243.01Fetal bovine serum 10%Cytiva, SV30207.02Penicillin-Streptomycin 1%Thermo Fisher, 15140122Dexamethasone (Dex) 1 μMsigma, D49023-isobutyl-1-methylxanthine (IBMX) 520 μMsigma, 228420010Insulin (I) 1 μMsigma, I5500Insulin mediaDulbecco's modified Eagle's medium (DMEM) Cytiva, SH30243.01Fetal bovine serum 10%Cytiva, SV30207.02 Penicillin-Streptomycin 1% Thermo Fisher, 15140122 Insulin (I) 167 nMSigma, I5500 FACS analysis Mouse LRIG1 Alexa488-conjugated Antibody R&D systems, FAB3688 GBoDIPY™ 493 / 503 (4,4-Difluoro-1,3,5,7,8-Pentamethyl-4-Bora-3a,4a-Diaza-s-Indacene) 1 μM Invitrogen, D3922.

[0181] [Example 1-2] siRNA transfection

[0182] 3T3-L1 cells derived from Example 1-1 were cultured at 1 x 10 5 After 3 days of seeding cells / well, siRNA transfection was performed at 100% density and cell cycle arrest. Control and Lrig-1 siRNAs were prepared according to Table 2 below. In addition, specific Lrig-1 siRNA information is shown in Table 3.

[0183] siRNAStock Conc.Working Conc.Media siRNA-Control (AM1611, Thermo Fisher)5μM240nMOpti-MEMsiRNA-Lrig1 (s68320, Thermo Fisher)10μM240nMOpti-MEM

[0184]

[0185] [Example 1-3] Confirmation of Lrig-1 expression after knockdown

[0186] Five days after knockdown and four days after induction, the expression level of Lrig-1 protein was analyzed by FACS. Specifically, the expression level of Lrig-1 protein was analyzed by FACS using Mouse LRIG-1 Alexa488-conjugated Antibody (R&D systems® Cat. No. FAB3688G).

[0187] As a result, it was confirmed that Lrig-1 protein was knocked down in the siRNA-treated group, as shown in Fig. 2.

[0188] [Example 1-4] Confirmation of inhibition of adipocyte differentiation upon Lrig-1 knockdown

[0189] The expression level of Lrig-1 protein was analyzed by FACS on the 13th day after knockdown and the 12th day after induction. Specifically, the expression of Lrig-1 protein was confirmed using Mouse LRIG-1 Alexa488-conjugated Antibody (R&D systems® Cat. No. FAB3688G), and the differentiation of pre-adipocytes into adipocytes was confirmed by FACS using 1 μM BODIPY™ 493 / 503 (4,4-Difluoro-1,3,5,7,8-Pentamethyl-4-Bora-3a,4a-Diaza-s-Indacene) (Invitrogen® Cat. No. D3922).

[0190] As a result, as shown in Fig. 3, it was confirmed that Lrig-1 protein was knocked down in the group treated with Lrig-1 siRNA, and differentiation of pre-adipocytes into adipocytes was inhibited compared to the control group.

[0191] [Example 1-5] Confirmation of inhibition of adipocyte differentiation upon Lrig-1 siRNA treatment

[0192] Previously, the inhibition of differentiation into adipocytes confirmed in Example 1-4 was specifically confirmed in terms of the kinetics of Lrig-1 expression and lipid droplet 3T3-L1 differentiation.

[0193] As a result, as shown in Fig. 4, when Lrig-1 expression was knocked down in 3T3-L1 cells by treating them with Lrig-1 siRNA, it was confirmed that Lrig-1 expression was suppressed in adipocytes and preadipocytes, and as shown in Fig. 5, it was confirmed that lipid droplet formation was suppressed in 3T3-L1 cells treated with Lrig-1 siRNA.

[0194] Additionally, as shown in Figures 6a and 6b, it was confirmed through microscopic photographs that the formation of lipid droplets in 3T3-L1 cells was inhibited in Lrig-1 knockdown cells.

[0195] [Example 2] MOA (Mode of Action) Analysis: Lrig-1 Knockdown

[0196] [Example 2-1] Induction for 3T3-L1 differentiation

[0197] Similar to Example 1, 3T3-L1 cells (pre-adipocytes) were first seeded at 1 x 10 in a 24-well plate. 5 / well and plated in DMI medium, i.e., Dulbecco's modified Eagle's medium (DMEM) from Cytiva® Cat. No. SH30243.01 supplemented with 10% fetal bovine serum from Cytiva® Cat. No. SV30207.02, 1% penicillin-streptomycin from Thermo Fisher® Cat. No. 15140122, 1 uM dexamethasone from Sigma® Cat. No. D4902, 520 μM 3-isobutyl-1-methylxanthine (IBMX) from Sigma® Cat. No. 228420010 and 100 μM 3-isobutyl-1-methylxanthine (IBMX) from Sigma® Cat. No. After culturing for 3 days in a medium containing 1 μM of insulin from I5500, the cells were cultured for 3 days in an insulin medium (i.e., Dulbecco's modified Eagle's medium (DMEM) from Cytiva® Cat. No. SH30243.01 supplemented with 10% fetal bovine serum from Cytiva® Cat. No. SV30207.02, 1% penicillin-streptomycin from Thermo Fisher® Cat. No. 15140122, and 167 nM of insulin from Sigma® Cat. No. I5500). Thereafter, the cells were cultured for 3 days in a complete DMEM medium (i.e., Dulbecco's modified Eagle's medium (DMEM) from Cytiva® Cat. No. SH30243.01 supplemented with 10% fetal bovine serum from Cytiva® Cat. No. SV30207.02, 1% penicillin-streptomycin from Thermo Fisher® Cat. No. 15140122, and 167 nM of insulin from Sigma® Cat. No. I5500). The strain was cultured for 8 days in a medium containing 10% fetal bovine serum (SV30207.02) and 1% penicillin-streptomycin (Thermo Fisher® Cat. No. 15140122).

[0198] [Example 2-2] siRNA transfection

[0199] 3T3-L1 cells derived from Example 2-1 were cultured at 1 x 10 5 After 3 days of seeding cells / well, siRNA transfection was performed at 100% density and cell cycle arrest. Control and Lrig-1 siRNAs were prepared according to Table 4 below, and Lrig knockdown was performed through siRNA transfection using siRNAs other than the siRNAs indicated in Tables 2 and 3 of Example 1-2.

[0200] StockWorkingMediasiRNA-Control (Thermo Fisher 4390843)10μM240nMOpti-MEM + S / F DMEMsiRNA-Lrig1 (Thermo Fisher 4390815)

[0201] Next, RANiMAX (Stock 1 μL / Opti-MEM 50 μL) was prepared, and 50 μL of the siRNA prepared in Table 4 was mixed with 50 μL of RANiMAX to prepare 100 μL of an siRNA mixture. The 3T3-L1 cells prepared in Example 2-1 were washed twice with S / F DMEM, and then treated with 500 μL of S / F DMEM. Subsequently, 100 μL of each siRNA mixture was treated to the negative control, control, and siRNA treatment groups (total 600 μL, Final conc. 20 nM, 24 h o / n). Afterwards, differentiation into adipocytes was induced (Adipocyte induction).

[0202] [Example 2-3] MoA (Mode of Action) Analysis

[0203] To determine whether Lrig-1 knockdown affects lipid metabolism, we performed a MoA analysis. Specifically, cells were harvested with PBS, washed twice, fixed at 4°C for 30 minutes, and then washed. Subsequently, FxCycle-Bv421 staining was performed at 4°C for 30 minutes, followed by flow cytometry analysis (time points: 0, 24, 48, and 96 hours; cell cycle: G1, S, G2, and M).

[0204] As a result, as shown in Fig. 7, it was confirmed through flow cytometry that Lrig-1 knockdown was properly performed, and as shown in Fig. 8, it was confirmed that the expression of Lrig-1 protein was reduced in the case of Lrig-1 knockdown, and as shown in Fig. 9, it was confirmed that the formation of lipid droplets was reduced in the case of Lrig-1 knockdown.

[0205] In addition, as shown in FIGS. 10 and 11, it was confirmed that the expression of FABP4 (Fatty Acid Binding Protein 4), a protein that transports fatty acids and is particularly expressed in large amounts in adipocytes and involved in regulating carbohydrate and lipid metabolism, was reduced in the case of Lrig-1 knockdown.

[0206] [Example 3] Analysis of experimental results from Treg Lrig-1 knockout mice

[0207] [Example 3-1] Preparation of L1 wKO (Lrig-1 CreERT2) mice

[0208] The L1 wKO (Lrig-1 CreERT2) mouse is a Lrig1 whole-body knockout mouse in which the creERT2 sequence is inserted into the translational initiation site of the endogenous Lrig1 locus to knock out Lrig1 and report the expression of Lrig1 as creERT2.

[0209] [Example 3-2] Observation of weight changes in L1 wKO (Lrig-1 CreERT2) mice

[0210] The L1 wKO (Lrig-1 CreERT2) mice prepared in Example 3-1 were divided into homo, hetero knockout mice, and male and female mice, and body weight changes were observed for 6 to 8 weeks.

[0211] The changes in genotype and body weight of L1 wKO (Lrig-1 CreERT2) mice are shown in Table 5 and Figure 12.

[0212]

[0213] [Example 3-3] Confirmation of decreased fat tissue and body weight in L1 wKO (Lrig-1 CreERT2) mice.

[0214] The weight of adipose tissue and changes in body weight were observed in L1 wKO (Lrig-1 CreERT2) mice twice. Specifically, changes in adipose tissue and body weight in male and female mice were observed over 5 to 12 weeks, and are shown in Figures 13 to 16.

[0215] As a result, in the case of male homo knockout mice, body weight decreased by 12.51% compared to the control group at 12 weeks, and visceral fat and subcutaneous fat also decreased. In the case of female mice, body weight decreased by 14.82% compared to the control group at 12 weeks, and visceral fat and subcutaneous fat also decreased.

[0216] The changes in adipose tissue and body weight of male and female mice were confirmed again for 6 to 12 weeks and are shown in Figures 17 to 20.

[0217] As a result, in the case of male homo knockout mice, body weight decreased by 8.98% compared to the control group at 12 weeks, and visceral fat and subcutaneous fat also decreased, and in particular, it was additionally confirmed that brown adipocytes also decreased. In the case of female mice, body weight decreased by 14.84% compared to the control group at 12 weeks, and visceral fat and subcutaneous fat also decreased.

[0218] Accordingly, it was confirmed that suppressing the function of Treg L1 reduces fat cells and body weight, and thus has a therapeutic effect on metabolic diseases including obesity.

[0219] [Example 4] Diagnosis of metabolic disease by confirming increased Lrig-1 expression in adipocytes.

[0220] [Example 4-1] Differentiation of 3T3-L1 cells into adipocytes

[0221] 3T3-L1 cell line, which is a cell line before differentiation into adipocytes and expresses Lrig-1, was seeded at 1x10 in a 24-well plate. 5 / well and cultured in DMI medium for two days, then in insulin medium for two days, and then in DMEM medium for two days.

[0222] The specific experimental protocol is shown in Table 6 and Figure 21 below.

[0223]

[0224] [Example 4-2] Analysis of Lrig-1 expression level in differentiated adipocytes

[0225] Here, we analyzed the level of Lrig-1 expression in differentiated adipocytes by flow cytometry using mouse Lrig-1 Alexa488-conjugated antibody. Specifically, cells were harvested and washed twice with PBS, and then stained with 1 μM BODIPY at 37 °C for 30 minutes. After washing, the cells were stained with anti-Lrig1-Alexa488 antibody, and then flow cytometry was performed.

[0226] The results are shown in Figures 22 to 24, and it was found that the expression of Lrig-1 increased in differentiated adipocytes from the 4th day, which can be used to diagnose metabolic diseases including obesity.

[0227] While specific aspects of the present invention have been described in detail above, it should be apparent to those skilled in the art that these specific descriptions are merely preferred embodiments and do not limit the scope of the present invention. Therefore, the substantial scope of the present invention is defined by the appended claims and their equivalents.

Claims

1. A pharmaceutical composition for preventing or treating metabolic diseases, comprising an Lrig protein inhibitor or an Lrig protein expression inhibitor as an active ingredient.

2. In paragraph 1, A pharmaceutical composition wherein the above Lrig protein expression inhibitor is an antisense nucleotide that complementarily binds to the transcript sequence of the Lrig protein.

3. In paragraph 2, A pharmaceutical composition, wherein the antisense nucleotide is at least one selected from the group consisting of small interfering RNA (siRNA), microRNA (mircoRNA; miRNA), short hairpin RNA, ribozyme, dsNRA, aptamer, PNA (peptide nucleic acid), ZFN, TALEN, and CRISPR (cluster regularly interspaced short palindromic repeats)-related nucleic acid, single guide RNA (sgRNA), CRISPR-RNA (crRNA), and trans-activating crRNA (tracrRNA).

4. In paragraph 3, A pharmaceutical composition wherein the Lrig protein is at least one selected from the group consisting of Lrig-1, Lrig-2 and Lrig-3.

5. In paragraph 4, A pharmaceutical composition wherein the Lrig protein is present on the surface of adipocytes, preadipocytes or regulatory T cells.

6. In paragraph 5, A pharmaceutical composition, wherein the metabolic disease is at least one selected from the group consisting of insulin resistance disease, obesity, diabetes, dyslipidemia, liver disease, renal damage, arteriosclerosis, and hypertension.

7. A composition for diagnosing a metabolic disease, comprising a preparation for measuring the expression level of a Lrig protein or a fragment thereof; or a gene encoding the same.

8. In paragraph 7, A diagnostic composition comprising at least one selected from the group consisting of antibodies, oligopeptides, ligands, peptide nucleic acids (PNAs), and aptamers that specifically bind to the protein, wherein the agent for measuring the expression level of the protein comprises:

9. In paragraph 7, A diagnostic composition comprising at least one selected from the group consisting of a primer, a probe, and an antisense nucleotide that specifically binds to the gene, wherein the agent for measuring the expression level of the gene is a composition.

10. In paragraph 7, A diagnostic composition, wherein the composition is intended for application to a biological sample isolated from a target object.

11. In paragraph 10, The biological samples include whole blood, leukocytes, peripheral blood mononuclear cells, buffy coat, plasma, serum, sputum, tears, mucus, nasal washes, nasal aspirate, breath, urine, semen, saliva, peritoneal washings, ascites, cystic fluid, meningeal fluid, amniotic fluid, glandular fluid, pancreatic fluid, lymph fluid, pleural fluid, nipple aspirate, bronchial aspirate, synovial fluid, joint aspirate, trachea. A diagnostic composition comprising at least one selected from the group consisting of organ secretions, cells, cell extracts, and cerebrospinal fluid.

12. A diagnostic kit comprising a diagnostic composition according to any one of claims 7 to 11.

13. In paragraph 12, The above kit is a diagnostic kit, which is an RT-PCR kit, a DNA chip kit, an ELISA kit, a protein chip kit, a rapid kit, or an MRM (Multiple reaction monitoring) kit.

14. In a biological sample isolated from the target organism, A method for providing information for diagnosing a metabolic disease, comprising: a step of determining that a metabolic disease has developed or is likely to develop if the expression level of the measured Lrig protein or the gene encoding it is higher than that of a control group; 15. In paragraph 14, The measurement of the expression level of the above protein is a method by protein chip analysis, immunoassay, ligand binding assay, MALDI-TOF (Matrix Assisted Laser Desorption / Ionization Time of Flight Mass Spectrometry) analysis, SELDI-TOF (Sulface Enhanced Laser Desorption / Ionization Time of Flight Mass Spectrometry) analysis, radioimmunoassay, radioimmunodiffusion, aukteroni immunodiffusion, rocket immunoelectrophoresis, tissue immunostaining, complement fixation assay, two-dimensional electrophoresis, liquid chromatography-mass spectrometry (LC-MS), liquid chromatography-mass spectrometry / mass spectrometry (LC-MS), Western blotting, enzyme linked immunosorbent assay (ELISA), or multiple reaction monitoring (MRM) method.

16. In paragraph 14, The expression level of the above gene can be measured by a method such as reverse transcription polymerase chain reaction (RT-PCR), competitive reverse transcription polymerase reaction (Competitive RT-PCR), real-time RT-PCR, RNase protection assay (RPA), Northern blotting, or DNA chip. 17.(a) A measuring unit for measuring the expression level of Lrig protein or a gene encoding the protein in a biological sample obtained from a target individual; and (b) A diagnostic device for metabolic diseases, comprising a detection unit that outputs the presence or absence of a metabolic disease from the expression level of a gene or a protein encoded by the gene measured in the above measurement unit.

18. A method for preventing or treating a metabolic disease comprising an Lrig protein inhibitor or an Lrig protein expression inhibitor as an active ingredient.

19. A method for diagnosing a metabolic disease, comprising a preparation for measuring the expression level of a Lrig protein or a fragment thereof; or a gene encoding the same.

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