WO2020214012A1 - Preventive or therapeutic pharmaceutical composition for hyperlipidemia comprising triple agonist acting on all of glucagon, glp-1 and gip receptors, or conjugate thereof, and preventive or therapeutic method - Google Patents

Preventive or therapeutic pharmaceutical composition for hyperlipidemia comprising triple agonist acting on all of glucagon, glp-1 and gip receptors, or conjugate thereof, and preventive or therapeutic method Download PDF

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WO2020214012A1
WO2020214012A1 PCT/KR2020/005233 KR2020005233W WO2020214012A1 WO 2020214012 A1 WO2020214012 A1 WO 2020214012A1 KR 2020005233 W KR2020005233 W KR 2020005233W WO 2020214012 A1 WO2020214012 A1 WO 2020214012A1
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cysteine
glutamine
lysine
alanine
glutamic acid
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PCT/KR2020/005233
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French (fr)
Korean (ko)
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조효상
김정국
이아람
김상윤
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한미약품 주식회사
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Publication of WO2020214012A1 publication Critical patent/WO2020214012A1/en

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/22—Hormones
    • A61K38/26—Glucagons
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/56—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/62—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being a protein, peptide or polyamino acid
    • A61K47/64—Drug-peptide, drug-protein or drug-polyamino acid conjugates, i.e. the modifying agent being a peptide, protein or polyamino acid which is covalently bonded or complexed to a therapeutically active agent
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/68—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an antibody, an immunoglobulin or a fragment thereof, e.g. an Fc-fragment
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P3/00—Drugs for disorders of the metabolism
    • A61P3/06—Antihyperlipidemics
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/46—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates
    • C07K14/47—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates from mammals
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/575—Hormones
    • C07K14/605—Glucagons

Definitions

  • the present invention relates to the therapeutic use of a triple activator or a conjugate thereof having activity on both glucagon, GLP-1 and GIP receptors for hyperlipidemia.
  • Lipids typically include cholesterol and triglycerides, and cholesterol is divided into low-density lipoprotein (LDL) cholesterol and high-density lipoprotein (HDL) cholesterol.
  • LDL low-density lipoprotein
  • HDL high-density lipoprotein
  • a condition in which cholesterol and triglyceride levels in the blood are out of the normal range (including both increase and decrease) and hypercholesterolemia a condition in which total cholesterol is high in the blood, hyperlipidemia is It is a disease that is a generic term for a case in which one or both of low-density lipoprotein cholesterol or triglycerides is high, and is a disease that is distinguished from dyslipidemia and hypercholesterolosis.
  • the pathogenesis and the patient's condition are different from each other, and different treatment and diet control methods are required.
  • it is a disease caused by abnormal regulation of lipid levels in the body, and abnormally low cholesterol levels can be a problem in dyslipidemia.
  • cholesterol levels essential for biological activities through dietary regimens such as dietary supplements It is important to increase.
  • cardiovascular disease and cerebrovascular disease are the three major causes of death in Korea along with cancer.
  • Hyperlipidemia is increasing as the obese population increases due to Western-style eating habits, and the number of hyperlipidemia patients is increasing by about 10% each year in Korea. It is known that obesity and alcohol are the causes of such hyperlipidemia. Obesity is the biggest etiology among them.
  • statin which acts as an HMGCR (HMG-CoA reductase) inhibitor and has an effect of inhibiting the synthesis of cholesterol
  • NPC1L1 Niemann-Pick C1-Like 1
  • Ezetimibe, and evolocumab which inhibits PCSK9 (Proprotein convertase subtilisin/kexin type 9), which interferes with LDL absorption, has been developed and is being used as a therapeutic agent.
  • Ezetimibe has a disadvantage in that its efficacy in improving blood lipids in clinical practice is significantly lower than that of conventional drugs.
  • Evolocumab has been reported to be ineffective in patients with genetic mutations in LDLR (LDL receptor).
  • LDLR LDL receptor
  • GLP-1 Glucagon-like peptide-1
  • GIP Glucose-dependent insuliontropic polypeptide
  • GLP-1 is a hormone secreted from the small intestine when stimulated by food intake. It promotes the secretion of insulin in the pancreas in a blood sugar concentration-dependent manner and helps to lower blood sugar levels. In addition, it acts as a satiety factor, slowing the digestion of the stomach and delaying the passage time of food digests, thereby reducing food intake. Moreover, when administered to rats, it was reported that there was an effect of inhibiting food intake and reducing weight, and these effects were found to be the same in both normal and obese states, showing the potential as a treatment for obesity.
  • GIP GIP
  • ApoB48 Apolipoprotein B
  • Glucagon is produced by the pancreas when blood sugar starts to drop due to drug treatment, disease, or hormone or enzyme deficiency. Glucagon signals the liver to break down glycogen to induce glucose to be released, and to raise blood sugar levels to normal levels. In addition, it has been reported that glucagon can suppress appetite, brown fat cells, and promote energy expenditure in animals and humans in addition to synergistic effects on blood sugar.
  • One object of the present invention is for the prevention or treatment of hyperlipidemia comprising a peptide or conjugate having activity against a glucagon receptor, a GLP-1 (Glucagon-like peptide-1) receptor, and a GIP (Glucose-dependent insuliontropic polypeptide) receptor. It is to provide a pharmaceutical composition.
  • Another object of the present invention is to provide a method for preventing or treating hyperlipidemia, comprising administering the peptide, conjugate, or composition containing the same to an individual in need thereof.
  • Another object of the present invention is to provide the use of the peptide, conjugate or a composition comprising the same in the manufacture of a medicament for the prevention or treatment of hyperlipidemia.
  • One aspect embodying the present invention is for the prevention or treatment of hyperlipidemia comprising a peptide having activity against a glucagon receptor, a GLP-1 (Glucagon-like peptide-1) receptor, and a GIP (Glucose-dependent insuliontropic polypeptide) receptor. It is a pharmaceutical composition.
  • the amino acid sequence added from the peptide is characterized in that it is derived from a native GLP-1, a native GIP, or a native exendin-4 amino acid sequence.
  • the peptide is characterized in that it is a peptide comprising an amino acid sequence represented by the following general formula 1:
  • Xaa1 is histidine (His, H), 4-imidazoacetyl (CA), or tyrosine (Tyr, Y)
  • Xaa2 is glycine (Gly, G), alpha-methyl-glutamic acid, or Aib (aminoisobutyric acid)
  • Xaa3 is glutamic acid (Glu, E) or glutamine (Gln, Q)
  • Xaa7 is threonine (Thr, T) or isoleucine (Ile, I)
  • Xaa10 is leucine (Leu, L), tyrosine (Tyr, Y), lysine (Lys, K), cysteine (Cys, C), or valine (Val, V)
  • Xaa12 is lysine (Lys, K), serine (Ser, S), or isoleucine (Ile, I)
  • Xaa13 is glutamine (Gln, Q),
  • R1 is cysteine (Cys, C), GKKNDWKHNIT (SEQ ID NO: 106), m-SSGAPPPS-n (SEQ ID NO: 107), or m-SSGQPPPS-n (SEQ ID NO: 108), or absent, wherein m is -Cys- , -Pro-, or -Gly-Pro-, and n is -Cys-, -Gly-, -Ser-, or -His-Gly-, or it is characterized in that it does not exist.
  • the peptide is characterized in that in the general formula 1, Xaa14 is leucine or methionine, and Xaa15 is cysteine, aspartic acid, or leucine.
  • Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib
  • Xaa7 is threonine
  • Xaa10 is tyrosine, cysteine, or valine
  • Xaa12 Is lysine or isoleucine
  • Xaa13 is tyrosine, alanine, glutamine, or cysteine
  • Xaa14 is leucine, cysteine, or methionine
  • Xaa15 is cysteine, leucine, glutamic acid, or aspartic acid
  • Xaa17 is glutamine, arginine, isoleucine, cysteine.
  • Xaa18 is alanine, glutamine, arginine, or histidine
  • Xaa19 is alanine, glutamine, valine, or cysteine
  • Xaa20 is lysine, arginine, or glutamine
  • Xaa21 is glutamic acid, glutamine, leucine, cysteine , Or aspartic acid
  • Xaa23 is isoleucine or valine
  • Xaa24 is cysteine, alanine, glutamine, asparagine, glutamic acid, or aspartic acid
  • Xaa27 is leucine or lysine.
  • the peptide is characterized in that it is a peptide comprising an amino acid sequence represented by the following general formula 2.
  • Xaa1 is 4-imidazoacetyl, histidine, or tyrosine
  • Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib
  • Xaa10 is tyrosine, or cysteine
  • Xaa13 is alanine, glutamine, tyrosine, Or cysteine
  • Xaa14 is leucine, methionine, or tyrosine
  • Xaa15 is aspartic acid, glutamic acid, or leucine
  • Xaa16 is glycine, glutamic acid, or serine
  • Xaa17 is glutamine, arginine, isoleucine, glutamic acid, cysteine, or lysine.
  • Xaa18 is alanine, glutamine, arginine, or histidine
  • Xaa19 is alanine, glutamine, cysteine, or valine
  • Xaa20 is lysine, glutamine, or arginine
  • Xaa21 is cysteine, glutamic acid, glutamine, leucine, or aspartic acid
  • Xaa23 is isoleucine or valine
  • Xaa24 is cysteine, alanine, glutamine, asparagine, or glutamic acid
  • Xaa28 is lysine, cysteine, asparagine, or aspartic acid
  • Xaa29 is glycine, glutamine, cysteine, or histidine
  • Xaa30 is cysteine, Glycine, lysine, or histidine
  • Xaa31 is proline or cysteine
  • Xaa40 is cysteine or absent.
  • Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib
  • Xaa7 is threonine
  • Xaa10 is tyrosine, cysteine, or valine
  • Xaa12 Is lysine or isoleucine
  • Xaa13 is tyrosine, alanine, or cysteine
  • Xaa14 is leucine or methionine
  • Xaa15 is cysteine or aspartic acid
  • Xaa17 is glutamine
  • Xaa18 is alanine, arginine.
  • Xaa19 is alanine, glutamine, or cysteine
  • Xaa20 is lysine or glutamine
  • Xaa21 is glutamic acid, cysteine, or aspartic acid
  • Xaa23 is valine
  • Xaa24 is alanine, glutamine, cysteine, asparagine, or aspartic acid. It is an acid
  • Xaa27 is characterized in that it is leucine or lysine.
  • Xaa13 is alanine, tyrosine, or cysteine
  • Xaa15 is aspartic acid or glutamic acid
  • Xaa17 is glutamine
  • Xaa18 is alanine, arginine, or histidine
  • Xaa21 is cysteine, glutamic acid, glutamine, or aspartic acid
  • Xaa23 is isoleucine or valine
  • Xaa24 is cysteine, glutamine, or asparagine
  • Xaa28 is cysteine, asparagine, or aspartic acid
  • Xaa29 is glutamine, cysteine, or histidine
  • Xaa30 is cysteine, lysine, or histidine.
  • Xaa2 is alpha-methyl-glutamic acid or Aib
  • Xaa7 is threonine
  • Xaa10 is tyrosine or cysteine
  • Xaa12 is lysine or isoleucine
  • Xaa13 is tyrosine, alanine, or cysteine
  • Xaa14 is leucine or methionine
  • Xaa15 is cysteine or aspartic acid
  • Xaa16 is glutamic acid
  • Xaa17 is arginine
  • Xaa18 is alanine, arginine, or histidine.
  • Xaa19 is alanine, glutamine, or cysteine
  • Xaa20 is lysine or glutamine
  • Xaa21 is glutamic acid or aspartic acid
  • Xaa23 is valine
  • Xaa24 is glutamine, asparagine, or aspartic acid
  • Xaa27 is leucine
  • Xaa28 is It is characterized in that it is cysteine, alanine, asparagine, or aspartic acid.
  • Xaa10 is tyrosine
  • Xaa12 is isoleucine
  • Xaa13 is alanine or cysteine
  • Xaa14 is methionine
  • Xaa15 is aspartic acid
  • Xaa16 is glutamic acid
  • Xaa17 is isoleucine or lysine
  • Xaa18 is alanine or histidine
  • Xaa20 is lysine
  • Xaa21 is aspartic acid
  • Xaa23 is valine
  • Xaa24 is asparagine
  • Xaa27 is leucine
  • Xaa28 is alanine or asparagine
  • Xaa29 is glutamine or threonine
  • Xaa30 is cysteine or It is characterized by being lysine or absent.
  • the peptide is characterized in that it is a peptide comprising the amino acid sequence of the following general formula 3.
  • Xaa1 is histidine or tyrosine
  • Xaa2 is alpha-methyl-glutamic acid or Aib
  • Xaa13 is alanine, tyrosine or cysteine
  • Xaa17 is arginine, cysteine, or lysine
  • Xaa18 is alanine or arginine
  • Xaa19 is alanine or cysteine
  • Xaa21 is glutamic acid or aspartic acid
  • Xaa24 is glutamine or asparagine
  • Xaa28 is cysteine or aspartic acid
  • Xaa29 is cysteine, histidine, or glutamine
  • Xaa30 is cysteine or histidine
  • Xaa31 is proline Or cysteine
  • Xaa40 is cysteine or absent.
  • composition wherein the peptide is a cysteine, GKKNDWKHNIT (SEQ ID NO: 106), CSSGQPPPS (SEQ ID NO: 109), GPSSGAPPPS (SEQ ID NO: 110), GPSSGAPPPSC (SEQ ID NO: 111), PSSGAPPPS (SEQ ID NO: Number 112), PSSGAPPPSG (SEQ ID NO: 113), PSSGAPPPSHG (SEQ ID NO: 114), PSSGAPPPSS (SEQ ID NO: 115), PSSGQPPPS (SEQ ID NO: 116), or PSSGQPPPSC (SEQ ID NO: 117), or absent.
  • GKKNDWKHNIT SEQ ID NO: 106
  • CSSGQPPPS SEQ ID NO: 109
  • GPSSGAPPPS SEQ ID NO: 110
  • GPSSGAPPPSC SEQ ID NO: 111
  • PSSGAPPPS SEQ ID NO: Number 112
  • PSSGAPPPSG SEQ ID NO: 113
  • PSSGAPPPSHG SEQ
  • composition according to any one of the preceding embodiments, wherein the peptide comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 1 to 102.
  • composition according to any one of the preceding embodiments, wherein the peptide is amidated at its C-terminus.
  • composition according to any one of the preceding embodiments, wherein the pharmaceutical composition has any one or more of an effect of increasing LDL absorption, inhibiting 3-Hydroxy-3-methylglutaryl-CoA reductase (HMGCR) activity, and promoting fatty acid degradation. To do.
  • HMGCR 3-Hydroxy-3-methylglutaryl-CoA reductase
  • Another aspect embodying the present invention is a pharmaceutical composition for the prevention or treatment of hyperlipidemia comprising a conjugate comprising the peptide.
  • the conjugate may be characterized in that it is represented by the following Formula 1:
  • X is a peptide having activity against the glucagon receptor, GLP-1 receptor, and GIP receptor,
  • L is polyethylene glycol
  • a is 0 or a natural number, provided that when a is 2 or more, each L is independent of each other,
  • F is an immunoglobulin Fc region.
  • the peptide is characterized in that it is a peptide comprising an amino acid sequence represented by the following general formula 1:
  • Xaa1 is histidine (His, H), 4-imidazoacetyl (CA), or tyrosine (Tyr, Y),
  • Xaa2 is glycine (Gly, G), alpha-methyl-glutamic acid, or Aib (aminoisobutyric acid),
  • Xaa3 is glutamic acid (Glu, E) or glutamine (Gln, Q),
  • Xaa7 is threonine (Thr, T) or isoleucine (Ile, I),
  • Xaa10 is leucine (Leu, L), tyrosine (Tyr, Y), lysine (Lys, K), cysteine (Cys, C), or valine (Val, V),
  • Xaa12 is lysine (Lys, K), serine (Ser, S), or isoleucine (Ile, I),
  • Xaa13 is glutamine (Gln, Q), tyrosine (Tyr, Y), alanine (Ala, A), or cysteine (Cys, C),
  • Xaa14 is leucine (Leu, L), methionine (Met, M), or tyrosine (Tyr, Y),
  • Xaa15 is cysteine (Cys, C), aspartic acid (Asp, D), glutamic acid (Glu, E), or leucine (Leu, L),
  • Xaa16 is glycine (Gly, G), glutamic acid (Glu, E), or serine (Ser, S),
  • Xaa17 is glutamine (Gln, Q), arginine (Arg, R), isoleucine (Ile, I), glutamic acid (Glu, E), cysteine (Cys, C), or lysine (Lys, K),
  • Xaa18 is alanine (Ala, A), glutamine (Gln, Q), arginine (Arg, R), or histidine (His, H),
  • Xaa19 is alanine (Ala, A), glutamine (Gln, Q), cysteine (Cys, C), or valine (Val, V),
  • Xaa20 is lysine (Lys, K), glutamine (Gln, Q), or arginine (Arg, R),
  • Xaa21 is glutamic acid (Glu, E), glutamine (Gln, Q), leucine (Leu, L), cysteine (Cys, C), or aspartic acid (Asp, D),
  • Xaa23 is isoleucine (Ile, I) or valine (Val, V),
  • Xaa24 is alanine (Ala, A), glutamine (Gln, Q), cysteine (Cys, C), asparagine (Asn, N), aspartic acid (Asp, D), or glutamic acid (Glu, E),
  • Xaa27 is valine (Val, V), leucine (Leu, L), lysine (Lys, K), or methionine (Met, M),
  • Xaa28 is cysteine (Cys, C), lysine (Lys, K), alanine (Ala, A), asparagine (Asn, N), or aspartic acid (Asp, D),
  • Xaa29 is cysteine (Cys, C), glycine (Gly, G), glutamine (Gln, Q), threonine (Thr, T), glutamic acid (Glu, E), or histidine (His, H),
  • Xaa30 is cysteine (Cys, C), glycine (Gly, G), lysine (Lys, K), or histidine (His, H) or absent,
  • R1 is cysteine (Cys, C), GKKNDWKHNIT (SEQ ID NO: 106), m-SSGAPPPS-n (SEQ ID NO: 107), or m-SSGQPPPS-n (SEQ ID NO: 108), or absent,
  • m is -Cys-, -Pro-, or -Gly-Pro-,
  • n is -Cys-, -Gly-, -Ser-, or -His-Gly- or absent.
  • the pharmaceutical composition is characterized in that it comprises a conjugate in which the peptide represented by the general formula 1 is bound to an immunoglobulin Fc region through polyethylene glycol.
  • composition according to any one of the preceding embodiments, wherein the pharmaceutical composition has any one or more of an effect of increasing LDL absorption, inhibiting 3-Hydroxy-3-methylglutaryl-CoA reductase (HMGCR) activity, and promoting fatty acid degradation. To do.
  • HMGCR 3-Hydroxy-3-methylglutaryl-CoA reductase
  • Another embodiment of the present invention is a method for preventing or treating hyperlipidemia, comprising administering the peptide, conjugate, or a composition containing the same to an individual in need thereof.
  • Another aspect embodying the present invention is the use of the peptide, conjugate, or a composition comprising the same in the manufacture of a medicament for the prevention or treatment of hyperlipidemia.
  • the triple activator long-acting conjugate according to the present invention has activity against the glucagon receptor, the GLP-1 (Glucagon-like peptide-1) receptor, and the GIP (Glucose-dependent insuliontropic polypeptide) receptor, and can be applied to the treatment of hyperlipidemia.
  • 1 is a diagram confirming in vivo the effect of treating hyperlipidemia of the triple activator long-acting conjugate of the present invention.
  • Figure 2 is a diagram confirming that LDL absorption is increased by the triple-active agent long-acting conjugate of the present invention.
  • HMGCR 3-Hydroxy-3-methylglutaryl-CoA reductase
  • Figure 4 is a diagram confirming the effect of promoting fatty acid decomposition of the triple active substance long-acting conjugate of the present invention.
  • One aspect for embodying the present invention is the prevention of hyperlipidemia, including a peptide having activity against glucagon receptor, GLP-1 (Glucagon-like peptide-1) receptor, and GIP (Glucose-dependent insuliontropic polypeptide) receptor Or a therapeutic pharmaceutical composition.
  • hyperlipidemia including a peptide having activity against glucagon receptor, GLP-1 (Glucagon-like peptide-1) receptor, and GIP (Glucose-dependent insuliontropic polypeptide) receptor Or a therapeutic pharmaceutical composition.
  • the triple activator of the present invention is characterized by having a therapeutic or preventive effect on hyperlipidemia.
  • hyperlipidemia unlike dyslipidemia, in which cholesterol and triglyceride levels in the blood are out of the normal range (including both increase and decrease) and hypercholesterolemia, in which total cholesterol in the blood is high, is low-density lipoprotein in the blood. It is a disease that is a generic term for a case in which one or both of cholesterol or triglycerides is high, and is a disease distinguished from dyslipidemia and hypercholesterolosis.
  • glucagon receptor GLP-1 receptor
  • peptide having activity against the GIP receptor may be used interchangeably as a triple activator in the present invention.
  • Such peptides include various substances, such as various peptides, with significant levels of activity against glucagon, GLP-1, and GIP receptors.
  • the triple activator having a significant level of activity against glucagon, GLP-1, and GIP receptors is one or more of glucagon, GLP-1, and GIP receptors, specifically two or more. More receptors, more specifically, in vitro activity against all three receptors is 0.1% or more, 1% or more, 2% compared to the natural ligands of the receptor (natural glucagon, natural GLP-1, and natural GIP).
  • a method of measuring the in vitro activity of such a triple activator may refer to Experimental Example 1 of the present specification, but is not particularly limited thereto.
  • the peptide is characterized by having one or more, two or more, specifically three activities, specifically significant activities of the following i) to iii):
  • activating the receptor means that the in vitro activity against the receptor is 0.1% or more, 1% or more, 2% or more, 3% or more, 4% or more, 5% or more, 6% or more, 7% or more, 8% or more, 9% or more, 10% or more, 20% or more, 30% or more, 40% or more, 50% or more, 60% or more, 70% or more, 80% or more, 90% or more, or 100% or more An example is the case. However, it is not limited thereto.
  • the composition according to the present invention contains an isolated peptide having activity against a glucagon receptor, a GLP-1 (Glucagon-like peptide-1) receptor, and a GIP (Glucose-dependent insuliontropic polypeptide) receptor, thereby increasing LDL absorption and synthesizing cholesterol. It may exhibit an effect of any one or more of the inhibitory and fatty acid decomposition promoting effects, thereby exhibiting an excellent therapeutic effect for hyperlipidemia. For example, the activity of 3-Hydroxy-3-methylglutaryl-CoA reductase (HMGCR) 24 to 48 hours after administration of the composition may be less than 50%.
  • HMGCR 3-Hydroxy-3-methylglutaryl-CoA reductase
  • the peptide may have an increased half-life in the body compared to any one of natural GLP-1, natural glucagon, and natural GIP, but is not particularly limited thereto.
  • such a peptide may be non-naturally occurring.
  • the isolated peptide may be an analog of natural glucagon, but is not particularly limited thereto.
  • the natural-type glucagon analog according to the present invention includes a peptide having one or more differences in amino acid sequence compared to the natural-type glucagon, a peptide modified through modification of the natural-type glucagon sequence, and a mimic of natural-type glucagon.
  • the native glucagon may have the following amino acid sequence:
  • the peptide is selected from the group consisting of substitution, addition, deletion, modification, and combinations thereof in at least one amino acid in the natural glucagon sequence. It may be an analog of glucagon, but is not particularly limited thereto.
  • substitution of the amino acid includes both substitution with amino acids and substitution with non-natural compounds.
  • addition may be made at the N-terminus and/or C-terminus of the peptide.
  • the length of the added amino acid is not particularly limited thereto, and 1 or more, 2 or more, 3 or more, 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, 9 or more, 10 or more, or 11 or more amino acids are added It may be, and broadly includes the addition of a polypeptide, but is not particularly limited thereto.
  • the glucagon analog is 1 times, 2 times, 3 times, 7 times, 10 times, 12 times, 13 times, 14 times, 15 times, 16 times, 17 times, 18 times in the natural glucagon amino acid sequence, 1 or more, 2 or more, 3 or more, 4 or more, 5 or more, 6 or more, 7 or more, selected from the group consisting of 19 times, 20 times, 21 times, 23 times, 24 times, 27 times, 28 times and 29 times, 8 or more, 9 or more, 10 or more, 11 or more, 12 or more, 13 or more, 14 or more, 15 or more, 16 or more, 17 or more, 18 or more, 19 or more, or 20 amino acids may be substituted with other amino acids,
  • 1 or more, 2 or more, 3 or more, 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, 9 or more, 10 or more, or 11 or more amino acids may be added to the C-terminus thereof. However, it is not particularly limited thereto.
  • the glucagon analog is 1, 2, 3, 10, 12, 13, 14, 15, 16, 17, 18, 19, in the natural glucagon amino acid sequence, 1 or more, 2 or more, 3 or more, 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, selected from the group consisting of 20, 21, 23, 24, 27, 28 and 29, 9 or more, 10 or more, 11 or more, 12 or more, 13 or more, 14 or more, 15 or more, 16 or more, 17 or more, 18 or more, or 19 amino acids may be substituted with other amino acids, and independently or additionally 1 or more, 2 or more, 3 or more, 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, 9 or more, 10 or more, or 11 or more amino acids may be added to the C-terminus, but is particularly limited thereto It is not.
  • the glucagon analog is 1, 2, 3, 10, 13, 14, 15, 16, 17, 18, 19, 20, in the natural glucagon amino acid sequence, 1 or more, 2 or more, 3 or more, 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, 9 or more, 10 or more, selected from the group consisting of 21 times, 23 times, 24 times, 28 times and 29 times, 11 or more, 12 or more, 13 or more, 14 or more, 15 or more, 16 or more, or 17 amino acids may be substituted with other amino acids, and also independently or additionally at the C-terminus of 1 or more, 2 or more, 3 or more , 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, 9 or more, 10 or more, or 11 or more amino acids may be added, but is not particularly limited thereto.
  • the glucagon analog is 1, 2, 13, 16, 17, 18, 19, 20, 21, 23, 24, 27, in the natural glucagon amino acid sequence, 1 or more, 2 or more, 3 or more, 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, 9 or more, 10 or more, 11 or more, 12 or more, 13 or more, selected from the group consisting of 28 and 29, Or 14 amino acids may be substituted with other amino acids, and independently or additionally at the C-terminus of 1 or more, 2 or more, 3 or more, 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, 9 or more , 10 or more, or 11 or more amino acids may be added, but is not particularly limited thereto.
  • amino acids introduced from the natural glucagon are tyrosine, alpha-methyl-glutamic acid, Aib, methionine, glutamic acid, histidine, lysine, leucine, isoleucine, glutamine, valine, glycine, alanine, cysteine, serine, alanine, aspartic acid, and arginine. It may be selected from the group consisting of, but is not particularly limited thereto.
  • the added amino acid sequence may be one or more amino acid sequences derived from native GLP-1, native GIP, or native exendin-4 amino acid sequence.
  • glucagon analogs or triple activators may include intramolecular bridges (e.g., covalent bridges or non-covalent bridges), and specifically may be in a form including a ring, such as glucagon analogs or triples.
  • a ring may be formed between amino acids 16 and 20 of the active substance, but is not particularly limited thereto.
  • Non-limiting examples of the ring may include a lactam bridge (or a lactam ring).
  • the glucagon analogue or triple activator includes all those modified to include an amino acid capable of forming a ring at a desired position to include a ring.
  • the pair of amino acids 16 and 20 of the glucagon analog or triple activator may be substituted with glutamic acid or lysine capable of forming a ring, but is not limited thereto.
  • Such a ring may be formed between the side chains of amino acids in the glucagon analog or triple activator, for example, a lactam ring may be formed between the side chain of lysine and the side chain of glutamic acid, but is not particularly limited thereto.
  • an analogue of glucagon prepared by a combination of these methods, in which one or more amino acid sequences are different from natural glucagon, and the alpha-carbon of the N-terminal amino acid residue has been removed, glucagon receptor, GLP-1 receptor, and GIP receptor.
  • glucagon receptor GLP-1 receptor
  • GIP receptor GIP receptor
  • amino acids may be substituted with other amino acids or non-natural compounds in order to avoid the recognition of activator decomposing enzymes to increase half-life in the body.
  • it may be a peptide having an increased half-life in the body by avoiding the recognition of a degrading enzyme through substitution of the second amino acid sequence of the amino acid sequence of the triple active agent, but amino acid substitution or Changes are included without limitation.
  • modifications for the production of analogs of natural glucagon include modifications using L-type or D-type amino acids, and/or non-natural amino acids; And/or by modifying the native sequence, for example, modification of side chain functional groups, intramolecular covalent bonds, such as inter-side chain ring formation, methylation, acylation, ubiquitination, phosphorylation, aminohexanation, biotinylation, etc. It includes everything you do.
  • substituted or added amino acids not only the 20 amino acids commonly observed in human proteins, but also atypical or non-naturally occurring amino acids may be used.
  • Commercial sources of atypical amino acids include Sigma-Aldrich, ChemPep and Genzyme pharmaceuticals. Peptides containing such amino acids and typical peptide sequences can be synthesized and purchased through commercial peptide synthesis companies, such as American peptide company or Bachem in the US, or Anygen in Korea.
  • Amino acid derivatives can also be obtained in the same way, and to name only some examples, 4-imidazoacetic acid or the like can be used.
  • the peptide according to the present invention has its N-terminus and/or C-terminus chemically modified or protected with an organic terminal in order to protect from protein cleavage enzymes in vivo and increase stability, or amino acid is added to the peptide terminal. It may be added and modified.
  • the N-terminus is acetylated and/or the C-terminus is amidated to remove such charge. It can be, but is not particularly limited thereto.
  • the peptide according to the present invention includes the peptide itself, a salt thereof (eg, a pharmaceutically acceptable salt of the peptide), or a solvate thereof.
  • the peptide may be in any pharmaceutically acceptable form.
  • the kind of the salt is not particularly limited. However, it is preferable that it is a safe and effective form for individuals, such as mammals, but is not particularly limited thereto.
  • pharmaceutically acceptable refers to a substance that can be effectively used for a desired purpose without causing excessive toxicity, irritation, or allergic reaction within the scope of medical judgment.
  • the term "pharmaceutically acceptable salt” includes salts derived from pharmaceutically acceptable inorganic acids, organic acids, or bases.
  • suitable acids include hydrochloric acid, bromic acid, sulfuric acid, nitric acid, perchloric acid, fumaric acid, maleic acid, phosphoric acid, glycolic acid, lactic acid, salicylic acid, succinic acid, toluene-p-sulfonic acid, tartaric acid, acetic acid, citric acid, methanesulfonic acid, formic acid.
  • Benzoic acid, malonic acid, naphthalene-2-sulfonic acid, and benzenesulfonic acid may include alkali metals such as sodium and potassium, alkaline earth metals such as magnesium, and ammonium.
  • solvate refers to the formation of a complex with a solvent molecule of the peptide or salt thereof according to the present invention.
  • the peptide may include an amino acid sequence represented by the following general formula 1.
  • Xaa1 is histidine, 4-imidazoacetyl, or tyrosine
  • Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib
  • Xaa3 is glutamic acid or glutamine
  • Xaa7 is threonine or isoleucine
  • Xaa10 is Leucine, tyrosine, lysine, cysteine, or valine
  • Xaa12 is lysine, serine, or isoleucine
  • Xaa13 is glutamine, tyrosine, alanine, or cysteine
  • Xaa14 is leucine, methionine, or tyrosine
  • Xaa15 is cysteine, aspartic acid.
  • Xaa16 is glycine, glutamic acid, or serine
  • Xaa17 is glutamine, arginine, isoleucine, glutamic acid, cysteine, or lysine
  • Xaa18 is alanine, glutamine, arginine, or histidine
  • Xaa19 is alanine, glutamine , Cysteine, or valine
  • Xaa20 is lysine, glutamine, or arginine
  • Xaa21 is glutamic acid, glutamine, leucine, cysteine, or aspartic acid
  • Xaa23 is isoleucine or valine
  • Xaa24 is alanine, glutamine, cysteine, asparagine, aspartic acid.
  • Xaa27 is valine, leucine, lysine, or methionine
  • Xaa28 is cysteine, lysine, alanine, asparagine, or aspartic acid
  • Xaa29 is cysteine, glycine, glutamine, threonine, glutamic acid, or histidine
  • Xaa30 Is cysteine, glycine, lysine, or histidine, or is absent
  • R1 is cysteine, GKKNDWKHNIT (SEQ ID NO: 106), m-SSGAPPPS-n (SEQ ID NO: 107), or m-SSGQPPPS-n (SEQ ID NO: 108), or absent,
  • n is -Cys-, -Gly-, -Ser-, or -His-Gly-, or may be absent.
  • triple activator examples include those comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 1 to 102, and those consisting of (essentially) an amino acid sequence selected from the group consisting of SEQ ID NOs: 1 to 102, but are limited thereto. It does not become.
  • Xaa14 may be leucine or methionine
  • Xaa15 may be cysteine, aspartic acid, or leucine.
  • a peptide examples include, but are not limited to, a peptide comprising or (essentially) an amino acid sequence selected from the group consisting of SEQ ID NOs: 1 to 12, 14 to 17, and 21 to 102.
  • These peptides may significantly activate one or more of the glucagon receptor, the GLP-1 receptor, and the GIP receptor, but are not particularly limited thereto. Specifically, it may significantly activate GLP-1, or significantly activate the glucagon receptor and/or the GIP receptor, but is not particularly limited thereto.
  • Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib
  • Xaa7 is threonine
  • Xaa10 is tyrosine, cysteine, or valine
  • Xaa12 is lysine or isoleucine
  • Xaa13 is tyrosine.
  • Xaa14 is leucine, cysteine, or methionine
  • Xaa15 is cysteine, leucine, glutamic acid, or aspartic acid
  • Xaa17 is glutamine, arginine, isoleucine, cysteine, glutamic acid, or lysine
  • Xaa18 is Alanine, glutamine, arginine, or histidine
  • Xaa19 is alanine, glutamine, valine, or cysteine
  • Xaa20 is lysine, arginine, or glutamine
  • Xaa21 is glutamic acid, glutamine, leucine, cysteine, or aspartic acid
  • Xaa23 is isoleucine.
  • Xaa24 is cysteine, alanine, glutamine, asparagine, glutamic acid, or aspartic acid
  • Xaa27 may be leucine or lysine, or a peptide, but is not particularly limited thereto.
  • Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib
  • Xaa7 is threonine
  • Xaa10 is tyrosine, cysteine, or valine
  • Xaa12 is lysine or isoleucine
  • Xaa13 is tyrosine.
  • Xaa14 is leucine or methionine
  • Xaa15 is cysteine or aspartic acid
  • Xaa17 is glutamine
  • Xaa18 is alanine, arginine, or histidine
  • Xaa19 is alanine
  • Xaa20 is lysine or glutamine
  • Xaa21 is glutamic acid, cysteine, or aspartic acid
  • Xaa23 is valine
  • Xaa24 is alanine, glutamine, cysteine, asparagine, or aspartic acid
  • Xaa27 is leucine or lysine.
  • Xaa2 is alpha-methyl-glutamic acid or Aib
  • Xaa7 is threonine
  • Xaa10 is tyrosine or cysteine
  • Xaa12 is lysine or isoleucine
  • Xaa13 is tyrosine, alanine, or cysteine.
  • Xaa14 is leucine or methionine
  • Xaa15 is cysteine or aspartic acid
  • Xaa16 is glutamic acid
  • Xaa17 is arginine, isoleucine, cysteine, or lysine
  • Xaa18 is alanine, arginine, or histidine
  • Xaa19 is alanine, glutamine, or Cysteine
  • Xaa20 is lysine or glutamine
  • Xaa21 is glutamic acid or aspartic acid
  • Xaa23 is valine
  • Xaa24 is glutamine, asparagine, or aspartic acid
  • Xaa27 is leucine
  • Xaa28 is cysteine, alanine, asparagine, or aspartic acid.
  • I can.
  • Xaa1 is histidine or 4-imidazoacetyl
  • Xaa2 is alpha-methyl-glutamic acid or Aib
  • Xaa3 is glutamine
  • Xaa7 is threonine
  • Xaa10 is tyrosine
  • Xaa12 is isoleucine.
  • Xaa13 is alanine or cysteine
  • Xaa14 is methionine
  • Xaa15 is aspartic acid
  • Xaa16 is glutamic acid
  • Xaa17 is isoleucine or lysine
  • Xaa18 is alanine or histidine
  • Xaa19 is glutamine or cysteine
  • Xaa20 is lysine
  • Xaa21 is aspartic acid
  • Xaa23 is valine
  • Xaa24 is asparagine
  • Xaa27 is leucine
  • Xaa28 is alanine or asparagine
  • Xaa29 is glutamine or threonine
  • Xaa30 is cysteine or lysine, or may be absent.
  • Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib
  • Xaa3 is glutamine
  • Xaa7 is threonine
  • Xaa10 is tyrosine, cysteine, or valine
  • Xaa12 is lysine
  • Xaa13 Is tyrosine
  • Xaa14 is leucine
  • Xaa15 is aspartic acid
  • Xaa16 is glycine, glutamic acid, or serine
  • Xaa17 is glutamine
  • Xaa18 is alanine, arginine, or histidine
  • Xaa19 is alanine.
  • Xaa20 is lysine or glutamine
  • Xaa21 is glutamic acid, cysteine, or aspartic acid
  • Xaa23 is valine
  • Xaa24 is alanine, glutamine, or cysteine
  • Xaa27 is leucine or lysine
  • Xaa29 is glycine, glutamine, It may be threonine or histidine, but is not particularly limited thereto.
  • These peptides have a significant degree of activation of the GLP-1 receptor and the glucagon receptor, and are higher than that of the GIP receptor;
  • the degree of activation of the GLP-1 receptor, the glucagon receptor, and the GIP receptor are all significant;
  • the degree of activation of the GLP-1 receptor and the GIP receptor is significant, and may correspond to a case that is higher than that of the glucagon receptor, but is not particularly limited thereto.
  • Examples of such peptides SEQ ID NO: 8, 9, 21 to 37, 39, 42, 43, 49 to 61, 64 to 83, 85, 86, 88, 89, 91 to 93, selected from the group consisting of 95 to 102 Peptides including or (essentially) composed of an amino acid sequence may be mentioned, but are not particularly limited thereto.
  • the peptide may include an amino acid sequence represented by the following general formula 2.
  • Xaa1 is 4-imidazoacetyl, histidine, or tyrosine
  • Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib
  • Xaa10 is tyrosine or cysteine
  • Xaa13 is alanine, glutamine, tyrosine, or cysteine
  • Xaa14 is leucine, methionine, or tyrosine
  • Xaa15 is aspartic acid, glutamic acid, or leucine
  • Xaa16 is glycine, glutamic acid, or serine
  • Xaa17 is glutamine, arginine, isoleucine, glutamic acid, cysteine, or lysine
  • Xaa18 is alanine, glutamine, arginine, or histidine
  • Xaa19 is alanine, glutamine, cysteine, or valine
  • Xaa20 is
  • Xaa13 is alanine, tyrosine, or cysteine
  • Xaa15 is aspartic acid or glutamic acid
  • Xaa17 is glutamine, arginine, cysteine, or lysine
  • Xaa18 is alanine, Arginine, or histidine
  • Xaa21 is cysteine, glutamic acid, glutamine, or aspartic acid
  • Xaa23 is isoleucine or valine
  • Xaa24 is cysteine, glutamine, or asparagine
  • Xaa28 is cysteine, asparagine, or aspartic acid
  • Xaa29 is glutamine, cysteine, or histidine
  • Xaa30 can be cysteine, lysine, or histidine.
  • the peptide may include the amino acid sequence of the following general formula 3.
  • Xaa1 is histidine or tyrosine
  • Xaa2 is alpha-methyl-glutamic acid or Aib
  • Xaa13 is alanine, tyrosine or cysteine
  • Xaa17 is arginine, cysteine, or lysine
  • Xaa18 is alanine or arginine
  • Xaa19 is alanine or cysteine
  • Xaa21 is glutamic acid or aspartic acid
  • Xaa24 is glutamine or asparagine
  • Xaa28 is cysteine or aspartic acid
  • Xaa29 is cysteine, histidine, or glutamine
  • Xaa30 is cysteine or histidine
  • Xaa31 is proline or cysteine
  • Xaa40 may be cysteine or absent.
  • Such peptides include, or (essentially) a peptide consisting of an amino acid sequence selected from the group consisting of SEQ ID NOs: 21, 22, 42, 43, 50, 64 to 71, 75 to 77, and 96 to 102. However, it is not particularly limited thereto.
  • R1 is cysteine, GKKNDWKHNIT (SEQ ID NO: 106), CSSGQPPPS (SEQ ID NO: 109), GPSSGAPPPS (SEQ ID NO: 110), GPSSGAPPPSC (SEQ ID NO: 111), PSSGAPPPS (SEQ ID NO: 112), PSSGAPPPSG (SEQ ID NO: 113), PSSGAPPPSHG (SEQ ID NO: 114), PSSGAPPPSS (SEQ ID NO: 115), PSSGQPPPS (SEQ ID NO: 116), or PSSGQPPPSC (SEQ ID NO: 117), or may be absent, but is not particularly limited thereto.
  • the peptide of the present invention can be synthesized by a method well known in the art, for example, an automatic peptide synthesizer, depending on its length, or can be produced by genetic engineering techniques.
  • the peptides of the present invention can be prepared by standard synthetic methods, recombinant expression systems, or any other art method.
  • the peptides according to the invention can be synthesized by a number of methods, including, for example, methods including:
  • the glucagon receptor, the GLP-1 receptor, and the peptide having an activity against the GIP receptor in vivo for increasing its half-life in vivo. It may be in the form of a combination, in which a suitable substance is combined.
  • the biocompatible material may be mixed with a carrier.
  • the conjugate of the peptide may exhibit increased persistence of potency compared to the peptide to which no carrier is bound, and in the present invention, the conjugate is referred to as a "persistent conjugate".
  • such a conjugate may be non-naturally occurring.
  • Another aspect for implementing the present invention comprises a conjugate comprising a peptide having activity against a glucagon receptor, a GLP-1 (Glucagon-like peptide-1) receptor, and a GIP (Glucose-dependent insuliontropic polypeptide) receptor.
  • a conjugate comprising a peptide having activity against a glucagon receptor, a GLP-1 (Glucagon-like peptide-1) receptor, and a GIP (Glucose-dependent insuliontropic polypeptide) receptor.
  • GLP-1 Glucagon-like peptide-1 receptor
  • GIP Glucose-dependent insuliontropic polypeptide
  • the conjugate is a conjugate represented by the following formula (1):
  • X is a peptide having activity against the glucagon receptor, GLP-1 receptor, and GIP receptor;
  • L is polyethylene glycol;
  • a is 0 or a natural number, provided that when a is 2 or more, each L is independent of each other;
  • F is an immunoglobulin Fc region.
  • Peptides having activity against X in the conjugate that is, glucagon receptor, GLP-1 receptor, and GIP receptor are as described above.
  • F is a substance capable of increasing the half-life of the peptide, having activity against X, that is, the glucagon receptor, the GLP-1 receptor, and the GIP receptor, and is a component of the moiety constituting the conjugate of the present invention. It corresponds.
  • the F may be bonded to each other by a covalent chemical bond or a non-covalent chemical bond with X, and F and X may be bonded to each other through L through a covalent chemical bond, a non-covalent chemical bond, or a combination thereof.
  • the immunoglobulin Fc region may specifically be an IgG Fc region, but is not particularly limited thereto.
  • One or more amino acid side chains in the peptides of the invention may be conjugated to such biocompatible materials to increase solubility and/or half-life and/or increase bioavailability in vivo. Such modifications can also reduce the clearance of therapeutic proteins and peptides.
  • the biocompatible material described above may be water-soluble (amphiphilic or hydrophilic) and/or non-toxic and/or pharmaceutically acceptable.
  • the F may be directly linked to X (ie, a is 0 in Formula 1), or linked through a linker (L).
  • L may be a non-peptidyl linker polyethylene glycol.
  • a may be 1 or more, and when a is 2 or more, each L may be independent.
  • the conjugate is shared with each other through a non-peptidyl linker including F at both ends, specifically an immunoglobulin Fc region and X, specifically a reactive group capable of binding to a peptide drug. It may be connected in combination.
  • the molecular weight of polyethylene glycol which is a non-peptidyl linker that can be used in the present invention, is in the range of 1 to 100 kDa, specifically 1 to 20 kDa, but is not limited thereto.
  • the non-peptidyl linker of the present invention bonded to the polypeptide corresponding to F may be a combination of different types of polymers as well as one type of polymer.
  • both ends of the non-peptidyl linker may be bonded to F, such as an amine group or thiol group of an immunoglobulin Fc region, and an amine group or thiol group of X.
  • the non-peptidyl polymer is a reactive group capable of binding to F (eg, immunoglobulin Fc region) and X at both ends, specifically X, or the N-terminus of F (eg, immunoglobulin Fc region) Or it may include a reactive group that can be bonded to the amine group located in the lysine or the thiol group of cysteine, but is not limited thereto.
  • the reactive group of the non-peptide polymer which may be bound to F, such as an immunoglobulin Fc region and X, may be selected from the group consisting of an aldehyde group, a maleimide group, and a succinimide derivative, but is not limited thereto.
  • the aldehyde group may be a propion aldehyde group or a butyl aldehyde group, but is not limited thereto.
  • the succinimide derivatives include succinimidyl valereate, succinimidyl methylbutanoate, succinimidyl methylpropionate, succinimidyl butanoate, succinimidyl propionate, N-hydroxysuccini Mid, hydroxy succinimidyl, succinimidyl carboxymethyl or succinimidyl carbonate may be used, but is not limited thereto.
  • the non-peptidyl linker may be connected to X and F through such a reactive group, but is not particularly limited thereto.
  • the final product produced by reductive amination by aldehyde bonds is much more stable than those linked by amide bonds.
  • the aldehyde reactor reacts selectively to the N-terminus at a low pH, and can form a covalent bond with a lysine moiety at a high pH, for example, pH 9.0.
  • the reactive groups at both ends of the non-peptidyl linker may be the same or different from each other, for example, a maleimide group at one end and an aldehyde group, a propion aldehyde group, or a butyl aldehyde group at the other end.
  • F, specifically immunoglobulin Fc region and X can be bonded to each end of the non-peptide linker, it is not particularly limited thereto.
  • one end of the non-peptide linker may include a maleimide group as a reactive group, and the other end may include an aldehyde group, a propion aldehyde group, or a butyl aldehyde group.
  • the hydroxy group may be activated with the various reactive groups by a known chemical reaction, or a polyethylene glycol having a commercially available modified reactive group may be used.
  • the long-acting protein conjugate of the invention can be prepared.
  • the non-peptide polymer may be linked to a cysteine residue of X, more specifically to a -SH group of cysteine, but is not limited thereto.
  • cysteine residue 10 cysteine 13
  • cysteine 15 cysteine 17, cysteine 19
  • cysteine 21 cysteine 24, cysteine 28, 29
  • the non-peptidyl polymer may be linked to a cysteine residue, a cysteine residue 30, a cysteine 31, a cysteine 40, or a cysteine 41, but is not particularly limited thereto.
  • a reactive group of a non-peptidyl polymer may be connected to the -SH group of the cysteine residue, and all of the above descriptions apply to the reactive group.
  • maleimide-PEG-aldehyde is used, the maleimide group is linked by a -SH group of X and a thioether bond, and the aldehyde group is F, specifically, a reductive amination reaction with the -NH 2 group of immunoglobulin Fc. It can be connected through, but is not limited thereto, and this corresponds to one example.
  • the reactive group of the non-peptide polymer may be linked to -NH 2 located at the N-terminus of the immunoglobulin Fc region, but this corresponds to one example.
  • the "immunoglobulin Fc region” refers to a region including a heavy chain constant region 2 (CH2) and/or a heavy chain constant region 3 (CH3) portion, excluding the heavy and light chain variable regions of an immunoglobulin.
  • the immunoglobulin Fc region may be a component constituting the moiety of the conjugate of the present invention.
  • Such an immunoglobulin Fc region may include a hinge portion in the heavy chain constant region, but is not limited thereto.
  • the immunoglobulin Fc region of the present invention has substantially the same or improved effect with the native type, part or all of the heavy chain constant region 1 (CH1) and/or light chain constant region except for the heavy and light chain variable regions of the immunoglobulin It may be an expanded Fc region comprising 1 (CL1). In addition, it may be a region from which some considerably long amino acid sequences corresponding to CH2 and/or CH3 have been removed.
  • the immunoglobulin Fc region of the present invention is 1) CH1 domain, CH2 domain, CH3 domain and CH4 domain, 2) CH1 domain and CH2 domain, 3) CH1 domain and CH3 domain, 4) CH2 domain and CH3 domain, 5) Combination of one or two or more domains of the CH1 domain, CH2 domain, CH3 domain, and CH4 domain with an immunoglobulin hinge region (or part of the hinge region), 6) heavy chain constant region It may be a dimer of each domain and light chain constant region. . However, it is not limited thereto.
  • the immunoglobulin Fc region may be in a dimeric form, and one X molecule may be covalently linked to one Fc region in a dimer form, and at this time, the immunoglobulin Fc and X can be linked to each other by a non-peptidyl polymer.
  • two X molecules it is also possible for two X molecules to symmetrically bind to one Fc region in the form of a dimer. At this time, the immunoglobulin Fc and X may be linked to each other by a non-peptide linker.
  • a non-peptide linker it is not limited to the examples described above.
  • the immunoglobulin Fc region of the present invention includes a native amino acid sequence as well as a sequence derivative thereof.
  • An amino acid sequence derivative means that one or more amino acid residues in a natural amino acid sequence have a different sequence by deletion, insertion, non-conservative or conservative substitution, or a combination thereof.
  • amino acid residues 214 to 238, 297 to 299, 318 to 322, or 327 to 331, which are known to be important for binding may be used as a suitable site for modification.
  • various kinds of derivatives are possible, such as a site capable of forming a disulfide bond is removed, several amino acids at the N-terminus in a native Fc are removed, or a methionine residue may be added to the N-terminus of a native Fc.
  • the complement binding site eg, the C1q binding site
  • the ADCC antibody dependent cell mediated cytotoxicity
  • the Fc derivative described above exhibits a biological activity equivalent to that of the Fc region of the present invention, and may have increased structural stability against heat and pH of the Fc region.
  • these Fc regions may be obtained from natural types isolated in vivo from animals such as humans, cows, goats, pigs, mice, rabbits, hamsters, rats or guinea pigs, or obtained from transformed animal cells or microorganisms. It may be recombinant or a derivative thereof.
  • the method of obtaining from the natural form may be a method of obtaining the whole immunoglobulin by separating it from a human or animal body and then treating a protease. When papain is treated, it is cleaved into Fab and Fc, and when treated with pepsin, it is cleaved into pF'c and F(ab) 2 . This can be separated from Fc or pF'c using size-exclusion chromatography or the like.
  • the human-derived Fc region is a recombinant immunoglobulin Fc region obtained from a microorganism.
  • the immunoglobulin Fc region may be a natural type sugar chain, an increased sugar chain compared to the natural type, a reduced sugar chain compared to the natural type, or a form in which sugar chains are removed.
  • Conventional methods such as chemical methods, enzymatic methods, and genetic engineering methods using microorganisms may be used for the increase or decrease of such immunoglobulin Fc sugar chains.
  • the immunoglobulin Fc region from which the sugar chain has been removed from the Fc significantly decreases the binding ability with complement (c1q), and the antibody-dependent cytotoxicity or complement-dependent cytotoxicity is reduced or eliminated, so it does not induce unnecessary immune responses in vivo. Does not.
  • a form more suitable for the original purpose as a drug carrier would be referred to as an immunoglobulin Fc region in which sugar chains are removed or non-glycosylated.
  • deglycosylation refers to an Fc region from which sugar is removed by an enzyme
  • aglycosylation refers to an Fc region that is not glycosylated by producing in a prokaryotic animal, or in a more specific embodiment, E. coli. .
  • the immunoglobulin Fc region may be of human origin or animal origin, such as cattle, goats, pigs, mice, rabbits, hamsters, rats, and guinea pigs, and in more specific embodiments, human origin.
  • the immunoglobulin Fc region may be an Fc region derived from IgG, IgA, IgD, IgE, or IgM, or a combination thereof or a hybrid thereof. In a more specific embodiment, it is derived from IgG or IgM, which is most abundant in human blood, and in a more specific embodiment, it is derived from IgG known to improve the half-life of a ligand binding protein. In a more specific embodiment, the immunoglobulin Fc region is an IgG4 Fc region, and in the most specific embodiment, the immunoglobulin Fc region is a non-glycosylated Fc region derived from human IgG4, but is not limited thereto.
  • “combination” means that when forming a dimer or multimer, a polypeptide encoding a single-chain immunoglobulin Fc region of the same origin forms a bond with a single-chain polypeptide of a different origin. That is, it is possible to prepare a dimer or a multimer from two or more fragments selected from the group consisting of IgG Fc, IgA Fc, IgM Fc, IgD Fc and IgE Fc fragment.
  • conjugate may be that the persistence of the potency is increased compared to the native GLP-1, GIP, or glucagon, or F is not modified compared to X, and such conjugates are not only in the above-described form, but also biodegradable nano Includes all shapes enclosed in particles, etc.
  • a composition comprising the peptide may be used for the prevention or treatment of hyperlipidemia.
  • prevention refers to any action that suppresses or delays the onset of hyperlipidemia by administration of the peptide or a composition comprising the same
  • treatment refers to the symptoms of hyperlipidemia by administration of the peptide or a composition comprising the same. It refers to any action that improves or benefits.
  • the term "administration" means introducing a predetermined substance to the patient by any suitable method, and the route of administration of the composition is not particularly limited thereto, but any general route by which the composition can reach the target in vivo Can be administered through, for example, intraperitoneal administration, intravenous administration, intramuscular administration, subcutaneous administration, intradermal administration, oral administration, topical administration, intranasal administration, intrapulmonary administration, or rectal administration. I can.
  • the pharmaceutical composition of the present invention may further include a pharmaceutically acceptable carrier, excipient, or diluent.
  • a pharmaceutically acceptable carrier, excipient, or diluent may be non-naturally occurring.
  • the term "pharmaceutically acceptable” means a sufficient amount to exhibit a therapeutic effect and does not cause side effects, and the type of disease, the patient's age, weight, health, sex, and the patient's sensitivity to drugs ,
  • the route of administration, the method of administration, the number of times of administration, the duration of the treatment, and drugs used in combination or concurrently can be easily determined by those skilled in the art according to factors well known in the medical field.
  • the pharmaceutical composition including the peptide or conjugate of the present invention may further include a pharmaceutically acceptable carrier.
  • the carrier is not particularly limited thereto, but when administered orally, a binder, a lubricant, a disintegrant, an excipient, a solubilizing agent, a dispersing agent, a stabilizing agent, a suspending agent, a coloring agent, a flavoring agent, etc. may be used.
  • a buffer agent Preservatives, painless agents, solubilizers, isotonic agents, stabilizers, and the like can be mixed and used.
  • base agents, excipients, lubricants, preservatives, etc. can be used.
  • the formulation of the composition of the present invention can be variously prepared by mixing with a pharmaceutically acceptable carrier as described above.
  • a pharmaceutically acceptable carrier as described above.
  • it when administered orally, it can be prepared in the form of tablets, troches, capsules, elixirs, suspensions, syrups, wafers, etc., and in the case of injections, it can be prepared in the form of unit dosage ampoules or multiple dosage forms.
  • Others, solutions, suspensions, tablets, pills, capsules can be formulated as sustained-release preparations.
  • examples of carriers, excipients and diluents suitable for formulation include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, Microcrystalline cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, mineral oil, and the like may be used.
  • fillers, anti-aggregating agents, lubricants, wetting agents, flavoring agents, preservatives, and the like may additionally be included.
  • the pharmaceutical composition of the present invention is any one selected from the group consisting of tablets, pills, powders, granules, capsules, suspensions, liquid solutions, emulsions, syrups, sterilized aqueous solutions, non-aqueous solutions, freeze-dried preparations, and suppositories. It can have a formulation of.
  • composition is formulated in a unit dosage form suitable for intra-body administration of a patient according to a conventional method in the pharmaceutical field, specifically, in a form useful for administration of protein medicines, and administration commonly used in the art.
  • the peptide or conjugate may be used in combination with various carriers permitted as drugs such as physiological saline or an organic solvent, and carbohydrates such as glucose, sucrose or dextran, ascorbic acid, in order to increase stability or absorption.
  • drugs such as physiological saline or an organic solvent
  • carbohydrates such as glucose, sucrose or dextran, ascorbic acid, in order to increase stability or absorption.
  • Antioxidants such as (ascorbic acid) or glutathione, chelating agents, low-molecular proteins, or other stabilizers may be used as drugs.
  • the dosage and frequency of the pharmaceutical composition of the present invention are determined according to the type of drug as the active ingredient, along with various related factors such as the disease to be treated, the route of administration, the age, sex and weight of the patient, and the severity of the disease.
  • the total effective amount of the composition of the present invention may be administered to a patient in a single dose, and may be administered by a fractionated treatment protocol that is administered for a long period of time in multiple doses.
  • the pharmaceutical composition of the present invention may vary the content of the active ingredient according to the severity of the disease.
  • the preferred total dose of the conjugate of the present invention may be about 0.0001 mg to 500 mg per 1 kg of patient body weight per day.
  • the dose of the conjugate is determined by considering various factors such as the patient's age, weight, health status, sex, disease severity, diet and excretion rate, as well as the administration route and number of treatments of the pharmaceutical composition.
  • composition according to the present invention is not particularly limited in its formulation, route of administration, and method of administration as long as it shows the effect of the present invention.
  • the pharmaceutical composition of the present invention has excellent in vivo persistence and potency, and can significantly reduce the number and frequency of administration of the pharmaceutical formulation of the present invention.
  • Another aspect embodying the present invention provides a method for preventing or treating hyperlipidemia, comprising administering the peptide, conjugate, or composition comprising the same to an individual in need thereof.
  • the peptide, conjugate, or composition comprising the same, hyperlipidemia, prevention and treatment are as described above.
  • the subject is an individual suspected of hyperlipidemia
  • the suspected hyperlipidemia refers to mammals including mice, livestock, etc., including humans who have or may develop the disease, but the conjugate of the present invention or include the same Subjects treatable with the composition described above are included without limitation.
  • the method of the present invention may include administering a pharmaceutical composition containing the peptide or conjugate in a pharmaceutically effective amount.
  • the appropriate total daily use amount may be determined by the treating physician within the range of correct medical judgment, and may be administered once or in several divided doses.
  • a specific therapeutically effective amount for a specific patient is a specific composition, including the type and degree of reaction to be achieved, whether other agents are used in some cases, the patient's age, weight, general health status, It is preferable to apply differently according to various factors including sex and diet, administration time, administration route and secretion rate of the composition, treatment period, drugs used with or concurrently with the specific composition, and similar factors well known in the medical field.
  • Another aspect embodying the present invention is the use of the peptide, conjugate, or a composition comprising the same in the manufacture of a medicament for the prevention or treatment of hyperlipidemia.
  • the peptide, conjugate, or composition comprising the same, hyperlipidemia, prevention and treatment are as described above.
  • the amino acid indicated by X is non-natural amino acid Aib (aminoisobutyric acid), and the underlined amino acid means that the underlined amino acids form a ring with each other.
  • Aib amino acid Aib
  • CA 4-imidazoacetyl
  • Y represents tyrosine.
  • maleimide-PEG-aldehyde 10 kDa PEG having a maleimide group and an aldehyde group at both ends
  • maleimide-PEG-aldehyde 10 kDa, NOF, Japan
  • the triple active agent of Example 1 SEQ ID NOs: 21, 22, 42, 43, 50, 77, and 96
  • the molar ratio of the triactivator to the maleimide-PEG-aldehyde is 1:1 to 3
  • the protein concentration is 1 to 5 mg/ml, and 0.5 to 3 at low temperature. Reacted for hours.
  • the reaction was carried out in an environment in which 20 to 60% isopropanol was added to 50 mM Tris buffer (pH 7.5). After the reaction was completed, the reaction solution was applied to SP Sepharose HP (GE healthcare, USA) to purify a triactivator mono-pegylated to cysteine.
  • SP Sepharose HP GE healthcare, USA
  • the purified mono-pegylated triactivator and immunoglobulin Fc were reacted at a molar ratio of 1: 1 to 5, and a protein concentration of 10 to 50 mg/ml for 12 to 18 hours at 4 to 8°C. Made it.
  • the reaction was carried out in an environment in which 10 to 50 mM sodium cyanoborohydride and 10 to 30% isopropanol as a reducing agent were added to 100 mM potassium phosphate buffer (pH 6.0).
  • the reaction solution is applied to a butyl Sepharose FF purification column (GE healthcare, USA) and a Source ISO purification column (GE healthcare, USA) to purify a conjugate including a triple activator and an immunoglobulin Fc. I did.
  • the conjugate in which the triple active agent and immunoglobulin Fc prepared in this example are linked through PEG is referred to as a conjugate or long-acting conjugate including a triple conjugate and an immunoglobulin Fc.
  • conjugate in which the triple activator of SEQ ID NO: 42 and immunoglobulin Fc are linked through PEG was named as'conjugate comprising SEQ ID NO: 42 and immunoglobulin Fc' or'long-acting conjugate of SEQ ID NO: 42', These may be used interchangeably herein.
  • Each of the cell lines is transformed to express human GLP-1 receptor, human GCG receptor and human GIP receptor genes in CHO (chinese hamster ovary), and is suitable for measuring the activities of GLP-1, GCG and GIP. Therefore, the activity for each part was measured using each transformed cell line.
  • human GLP-1 was serially diluted from 50 nM to 0.000048 nM by 4 times, and prepared in Examples 1 and 2
  • the triple activator and its long-acting conjugate were serially diluted from 400nM to 0.00038nM by 4 times.
  • Remove the culture medium from the cultured human GLP-1 receptor-expressing CHO cells add 5 ⁇ l of each of the serially diluted substances to the cells, and then add 5 ⁇ l each of a buffer containing cAMP antibody for 15 minutes. During incubation at room temperature.
  • Table 3 is a table confirming the relative titer ratio of the triple active long-acting conjugate, and the novel triple-active long-acting conjugate prepared above has triple activity capable of activating all of the GLP-1 receptor, GIP receptor, and glucagon receptor. It was confirmed that it has a sieve function.
  • a hyperlipidemia model was induced by feeding Fructose for 2 weeks in a Golden syrian hamster (hereinafter referred to as hyperlipidemia hamster).
  • SEQ ID NO: 42 was selected, and the long-acting conjugate of SEQ ID NO: 42 (1.6, 3.1 nmol/kg, Q2D) was repeatedly administered subcutaneously for 3 weeks to hamsters inducing hyperlipidemia.
  • evolocumab 219.2 nmol/kg, QW
  • QW a commercially available treatment for hyperlipidemia
  • the triple active long-acting conjugate of Example 2 has excellent hyperlipidemia treatment efficacy.
  • the mode of action was confirmed in vitro .
  • HepG2 cells were treated with the control, the long-acting conjugate of SEQ ID NO: 42 (0.01, 0.1, 1, 10 ⁇ M) and evolocumab (10 ⁇ g/mL) for 48 hours, and then the amount of BODIPY-labeled LDL uptake was confirmed.
  • the results are shown in FIG. 2.
  • HMGCR 3-Hydroxy-3-methylglutaryl-CoA reductase
  • the ketone body a by-product of fatty acid decomposition, was measured.
  • the amount of the produced ketone body was measured using a commercially available ketone body measurement kit (Sigma #MAK041).
  • the triple activator and the long-acting conjugate thereof according to the present invention increases LDL absorption, inhibits cholesterol synthesis through inhibition of HMGCR activity, and reduces total cholesterol and LDL in blood through promotion of fatty acid degradation, thereby treating hyperlipidemia. It was confirmed that the effect can be exhibited.

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Abstract

The present invention relates to therapeutic use of a triple agonist or a long-lasting conjugate thereof against hyperlipidemia, the triple agonist acting on all of glucagon, GLP-1 and GIP receptors.

Description

글루카곤, GLP-1 및 GIP 수용체 모두에 활성을 갖는 삼중 활성체 또는 이의 결합체를 포함하는 고지혈증 예방 또는 치료용 약학적 조성물 및 예방 또는 치료 방법A pharmaceutical composition for preventing or treating hyperlipidemia and a method for preventing or treating hyperlipidemia comprising a triple activator having activity on both glucagon, GLP-1 and GIP receptors or a conjugate thereof
본 발명은 글루카곤, GLP-1 및 GIP 수용체 모두에 활성을 갖는 삼중 활성체 또는 이의 결합체의 고지혈증에 대한 치료적 용도에 관한 것이다. The present invention relates to the therapeutic use of a triple activator or a conjugate thereof having activity on both glucagon, GLP-1 and GIP receptors for hyperlipidemia.
지방질에는 대표적으로 콜레스테롤과 중성지방이 있고, 이 중 콜레스테롤은 저밀도지단백(LDL) 콜레스테롤과 고밀도지단백(HDL) 콜레스테롤로 나누어진다. 관련하여, 혈액 내에 콜레스테롤과 중성지방 수치가 정상범위에서 벗어나 있는(증가 및 감소 모두 포함) 상태인 이상지질혈증과 혈액 내에 총 콜레스테롤이 높은 상태인 고콜레스테롤혈증과 달리, 고지혈증(hyperlipidemia)은 혈액 내에 저밀도지단백 콜레스테롤이나 중성지방 중 한가지가 높거나 둘 다 높은 경우를 총칭하는 질환으로서 이상지질혈증 및 고콜레스테롤증과 구분되는 질환이다. 이에 따라, 이들 질환에 있어서 발병 기전 및 환자의 상태가 서로 상이하여 서로 다른 치료 및 식이조절 방법이 요구된다. 예켠대, 체내 지질 수준 조절 이상으로 발병되는 질환으로서 이상지질혈증의 경우 비정상적으로 낮은 콜레스테롤 수치가 문제가 될 수 있는데, 고지혈증과 달리 건강보조식품 섭취와 같은 식이요법을 통해 생체활동에 필수적인 콜레스테롤의 수치를 증가시키는 것이 중요하다.Lipids typically include cholesterol and triglycerides, and cholesterol is divided into low-density lipoprotein (LDL) cholesterol and high-density lipoprotein (HDL) cholesterol. Relatedly, unlike dyslipidemia, a condition in which cholesterol and triglyceride levels in the blood are out of the normal range (including both increase and decrease) and hypercholesterolemia, a condition in which total cholesterol is high in the blood, hyperlipidemia is It is a disease that is a generic term for a case in which one or both of low-density lipoprotein cholesterol or triglycerides is high, and is a disease that is distinguished from dyslipidemia and hypercholesterolosis. Accordingly, in these diseases, the pathogenesis and the patient's condition are different from each other, and different treatment and diet control methods are required. As a matter of fact, it is a disease caused by abnormal regulation of lipid levels in the body, and abnormally low cholesterol levels can be a problem in dyslipidemia.Unlike hyperlipidemia, cholesterol levels essential for biological activities through dietary regimens such as dietary supplements It is important to increase.
혈액 내에 저밀도지단백 콜레스테롤 및/또는 중성지방이 비정상적으로 높은 경우, 혈관벽에 쌓여 염증을 일으키고, 그 결과 심혈관계질환 (cardiovascular disease), 뇌혈관질환 (Cerebrovascular diseases), 말초동맥질환 (peripheral arterial disease)을 일으킬 수 있는 원인이 된다. 이중 심혈관계질환 및 뇌혈관질환은 암과 함께 우리나라 3대 사망원인이다. 고지혈증은 서구식 식습관으로 인한 비만 인구가 증가함에 따라 증가하고 있는 추세이며, 국내에서도 고지혈증 환자가 해마다 약 10%씩 증가하는 추세이다. 이러한 고지혈증의 병인으로는 비만, 음주 등이 기인한다고 알려져 있다. 그 중 가장 큰 병인으로는 비만을 꼽는다.Abnormally high levels of low-density lipoprotein cholesterol and/or triglycerides in the blood cause inflammation by accumulating on the walls of blood vessels, resulting in cardiovascular disease, cerebrovascular disease, and peripheral arterial disease. It is a possible cause. Among them, cardiovascular disease and cerebrovascular disease are the three major causes of death in Korea along with cancer. Hyperlipidemia is increasing as the obese population increases due to Western-style eating habits, and the number of hyperlipidemia patients is increasing by about 10% each year in Korea. It is known that obesity and alcohol are the causes of such hyperlipidemia. Obesity is the biggest etiology among them.
이와 같은 고지혈증을 예방 및/또는 치료하기 위해 혈중 지질을 개선시키는데 많은 노력을 하고 있다. 예를 들어, HMGCR (HMG-CoA reductase) 억제제로 작용하며 콜레스테롤의 합성을 저해하는 효과를 가진 statin과 소장에서 NPC1L1 (Niemann-Pick C1-Like 1)를 억제하여 콜레스테롤의 재흡수를 억제하는 효과를 가진 ezetimibe, 그리고 LDL 흡수를 방해하는 PCSK9 (Proprotein convertase subtilisin/kexin type 9)을 억제하는 evolocumab이 개발이 완료되어 치료제로 사용되고 있다. (US Endocrinology, 2011;7(1):23-9, Am J Pharm Benefits. 2010;2(4):267-274, N Engl J Med 2014; 370:1809-1819)In order to prevent and/or treat such hyperlipidemia, many efforts have been made to improve blood lipids. For example, statin, which acts as an HMGCR (HMG-CoA reductase) inhibitor and has an effect of inhibiting the synthesis of cholesterol, and NPC1L1 (Niemann-Pick C1-Like 1) in the small intestine, inhibit the reuptake of cholesterol. Ezetimibe, and evolocumab, which inhibits PCSK9 (Proprotein convertase subtilisin/kexin type 9), which interferes with LDL absorption, has been developed and is being used as a therapeutic agent. (US Endocrinology, 2011;7(1):23-9, Am J Pharm Benefits. 2010;2(4):267-274, N Engl J Med 2014; 370:1809-1819)
그러나 statin계열의 약물을 이용한 치료의 경우, 충분히 콜레스테롤을 조절해주지 못할 때가 많을 뿐만 아니라 고용량 또는 장기간 사용할 경우, 간 독성 및 근육 병증을 유발 할 수 있다는 단점이 있다. ezetimibe는 임상에서 혈중 지질개선에 대한 효력이 기존 약물에 비해 현저히 낮다는 단점을 가지고 있다. Evolocumab의 경우 LDLR (LDL receptor) 유전적 변이가 있는 환자에게는 효력이 미비하다 보고가 있다. 위에서 언급한 바와 같이 현재 상용화된 약물들의 경우 단일 표적에만 작용하기 때문에 유전적 변이를 가진 환자나, 초위험군 고지혈증 환자에서의 혈중 지질제거에는 효과적이지 못하다. 따라서, 폭 넓은 고지혈증 환자에게 효능을 보일 수 있는, 다중 표적을 가진 약물의 개발이 여전히 요구되고 있다. However, in the case of treatment using statin-based drugs, there are many disadvantages in that it is not possible to sufficiently control cholesterol, and when used in high doses or for a long period of time, liver toxicity and myopathy may occur. Ezetimibe has a disadvantage in that its efficacy in improving blood lipids in clinical practice is significantly lower than that of conventional drugs. Evolocumab has been reported to be ineffective in patients with genetic mutations in LDLR (LDL receptor). As mentioned above, currently commercially available drugs act only on a single target, so they are not effective in removing lipids from blood in patients with genetic mutations or in hyperlipidemia patients. Therefore, there is still a need for the development of drugs with multiple targets that can show efficacy in a wide range of hyperlipidemia patients.
한편, GLP-1 (Glucagon-like peptide-1) 및 GIP (Glucose-dependent insuliontropic polypeptide)는 대표적인 위장 호르몬이자 신경 호르몬으로서 음식물 섭취에 따른 혈중 당 농도 조절에 관여하는 물질이다. 글루카곤 (Glucagon)은 췌장에서 분비되는 펩타이드 호르몬으로 전술한 두 물질과 함께 혈중 당 농도 조절작용 및 지질 대사에 관여한다.On the other hand, GLP-1 (Glucagon-like peptide-1) and GIP (Glucose-dependent insuliontropic polypeptide) are representative gastrointestinal hormones and neurological hormones, and are substances involved in the regulation of blood sugar levels according to food intake. Glucagon is a peptide hormone secreted from the pancreas and, together with the two substances described above, is involved in blood sugar concentration regulation and lipid metabolism.
상세하게는 GLP-1은 음식물 섭취에 자극을 받아 소장에서 분비되는 호르몬으로 혈당 농도 의존적으로 췌장에서의 인슐린 분비를 촉진하고, 혈당 농도를 낮추는 작용을 돕는다. 또한, 포만 인자로 작용하여 위장의 소화작용을 늦추고 음식 소화물의 위장 통과시간을 지연시켜 음식물 섭취를 줄이는 역할을 지닌다. 더욱이 쥐에 투여 시 음식 섭취억제와 체중 감소효과가 있음이 보고되었으며, 이러한 효과는 정상과 비만상태 모두에서 동일하게 나타남이 확인되어 비만 치료제로서의 가능성을 보여주었다.In detail, GLP-1 is a hormone secreted from the small intestine when stimulated by food intake. It promotes the secretion of insulin in the pancreas in a blood sugar concentration-dependent manner and helps to lower blood sugar levels. In addition, it acts as a satiety factor, slowing the digestion of the stomach and delaying the passage time of food digests, thereby reducing food intake. Moreover, when administered to rats, it was reported that there was an effect of inhibiting food intake and reducing weight, and these effects were found to be the same in both normal and obese states, showing the potential as a treatment for obesity.
GLP-1과 더불어 음식섭취에 자극을 받아 분비되는 위장 호르몬의 하나인 GIP는 소장의 K 세포로부터 분비되는 42개 아미노산으로 구성된 호르몬으로서 소장에서 흡수 된 지질을 소장세포(enterocyte)에서 ApoB48 (Apolipoprotein B 48)을 억제 함으로써 모세혈관으로의 지질 방출을 낮추는데 도움을 주는 기능을 수행하며 GLP-1의 활성 증가 효과, 항 염증 효과 등이 보고되었다.Along with GLP-1, GIP, one of the gastrointestinal hormones secreted by food intake, is a hormone composed of 42 amino acids secreted from K cells in the small intestine. Lipids absorbed from the small intestine are transferred from the enterocyte to ApoB48 (Apolipoprotein B). 48) to help lower the release of lipids into the capillaries by inhibiting them, and the effect of increasing the activity of GLP-1 and anti-inflammatory effects have been reported.
글루카곤은 약물 치료 또는 질병, 호르몬이나 효소 결핍 등의 원인으로 혈당이 떨어지기 시작하면 췌장에서 생산된다. 글루카곤은 간에 신호하여 글리코겐을 분해하여 글루코오스를 방출하도록 유도하고, 혈당 수준을 정상 수준까지 높이는 역할을 한다. 또한 글루카곤은 혈당 상승효과 이외에 동물과 사람에서 식욕억제, 지방세포 갈색화 및 에너지대사 (energy expenditure) 촉진할 수 있음이 보고 되었다.Glucagon is produced by the pancreas when blood sugar starts to drop due to drug treatment, disease, or hormone or enzyme deficiency. Glucagon signals the liver to break down glycogen to induce glucose to be released, and to raise blood sugar levels to normal levels. In addition, it has been reported that glucagon can suppress appetite, brown fat cells, and promote energy expenditure in animals and humans in addition to synergistic effects on blood sugar.
본 발명의 하나의 목적은 글루카곤 수용체, GLP-1 (Glucagon-like peptide-1) 수용체, 및 GIP (Glucose-dependent insuliontropic polypeptide) 수용체에 대해 활성을 갖는 펩타이드 또는 결합체를 포함하는 고지혈증의 예방 또는 치료용 약학적 조성물을 제공하는 것이다. One object of the present invention is for the prevention or treatment of hyperlipidemia comprising a peptide or conjugate having activity against a glucagon receptor, a GLP-1 (Glucagon-like peptide-1) receptor, and a GIP (Glucose-dependent insuliontropic polypeptide) receptor. It is to provide a pharmaceutical composition.
본 발명의 다른 목적은 상기 펩타이드, 결합체 또는 이를 포함하는 조성물을 이를 필요로 하는 개체에 투여하는 단계를 포함하는, 고지혈증의 예방 또는 치료 방법을 제공하는 것이다. Another object of the present invention is to provide a method for preventing or treating hyperlipidemia, comprising administering the peptide, conjugate, or composition containing the same to an individual in need thereof.
본 발명의 다른 목적은 고지혈증의 예방 또는 치료를 위한 약제의 제조에 있어서 상기 펩타이드, 결합체 또는 이를 포함하는 조성물의 용도를 제공하는 것이다. Another object of the present invention is to provide the use of the peptide, conjugate or a composition comprising the same in the manufacture of a medicament for the prevention or treatment of hyperlipidemia.
본 발명을 구현하는 하나의 양태는 글루카곤 수용체, GLP-1 (Glucagon-like peptide-1) 수용체, 및 GIP (Glucose-dependent insuliontropic polypeptide) 수용체에 대해 활성을 갖는 펩타이드를 포함하는 고지혈증의 예방 또는 치료용 약학적 조성물이다. One aspect embodying the present invention is for the prevention or treatment of hyperlipidemia comprising a peptide having activity against a glucagon receptor, a GLP-1 (Glucagon-like peptide-1) receptor, and a GIP (Glucose-dependent insuliontropic polypeptide) receptor. It is a pharmaceutical composition.
하나의 구체예로서, 상기 펩타이드에서 추가되는 아미노산 서열은 천연형 GLP-1, 천연형 GIP, 또는 천연형 엑센딘-4 아미노산 서열에서 유래된 것을 특징으로 한다. In one embodiment, the amino acid sequence added from the peptide is characterized in that it is derived from a native GLP-1, a native GIP, or a native exendin-4 amino acid sequence.
다른 구체예로서, 상기 펩타이드는 하기 일반식 1로 표시되는 아미노산 서열을 포함하는 펩타이드인 것을 특징으로 한다:In another embodiment, the peptide is characterized in that it is a peptide comprising an amino acid sequence represented by the following general formula 1:
Xaa1-Xaa2-Xaa3-Gly-Thr-Phe-Xaa7-Ser-Asp-Xaa10-Ser-Xaa12-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp-Leu-Xaa27-Xaa28-Xaa29-Xaa30-R1 (일반식 1, 서열번호 103)Xaa1-Xaa2-Xaa3-Gly-Thr-Phe-Xaa7-Ser-Asp-Xaa10-Ser-Xaa12-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp- Leu-Xaa27-Xaa28-Xaa29-Xaa30-R1 (general formula 1, SEQ ID NO: 103)
상기 일반식 1에서, Xaa1은 히스티딘 (His, H), 4-이미다조아세틸 (CA), 또는 티로신 (Tyr, Y)이고, Xaa2는 글리신 (Gly, G), 알파-메틸-글루탐산, 또는 Aib (aminoisobutyric acid)이며, Xaa3은 글루탐산 (Glu, E) 또는 글루타민 (Gln, Q)이고, Xaa7은 트레오닌 (Thr, T) 또는 이소류신 (Ile, I)이며, Xaa10은 류신 (Leu, L), 티로신 (Tyr, Y), 리신 (Lys, K), 시스테인 (Cys, C), 또는 발린 (Val, V)이고, Xaa12는 리신 (Lys, K), 세린 (Ser, S), 또는 이소류신 (Ile, I)이며, Xaa13은 글루타민 (Gln, Q), 티로신 (Tyr, Y), 알라닌 (Ala, A), 또는 시스테인 (Cys, C)이고, Xaa14는 류신 (Leu, L), 메티오닌 (Met, M), 또는 티로신 (Tyr, Y)이며, Xaa15는 시스테인 (Cys, C), 아스파르트산 (Asp, D), 글루탐산 (Glu, E), 또는 류신 (Leu, L)이며, Xaa16은 글리신 (Gly, G), 글루탐산 (Glu, E), 또는 세린 (Ser, S)이고, Xaa17은 글루타민 (Gln, Q), 아르기닌 (Arg, R), 이소류신 (Ile, I), 글루탐산 (Glu, E), 시스테인 (Cys, C), 또는 리신 (Lys, K)이며, Xaa18은 알라닌 (Ala, A), 글루타민 (Gln, Q), 아르기닌 (Arg, R), 또는 히스티딘 (His, H)이고, Xaa19는 알라닌 (Ala, A), 글루타민 (Gln, Q), 시스테인 (Cys, C), 또는 발린 (Val, V)이며, Xaa20은 리신 (Lys, K), 글루타민 (Gln, Q), 또는 아르기닌 (Arg, R)이고, Xaa21은 글루탐산 (Glu, E), 글루타민 (Gln, Q), 류신 (Leu, L), 시스테인 (Cys, C), 또는 아스파르트산 (Asp, D)이며, Xaa23은 이소류신 (Ile, I) 또는 발린 (Val, V)이고, Xaa24는 알라닌 (Ala, A), 글루타민 (Gln, Q), 시스테인 (Cys, C), 아스파라긴 (Asn, N), 아스파르트산 (Asp, D), 또는 글루탐산 (Glu, E)이며, Xaa27은 발린 (Val, V), 류신 (Leu, L), 리신 (Lys, K), 또는 메티오닌 (Met, M)이고, Xaa28은 시스테인 (Cys, C), 리신 (Lys, K), 알라닌 (Ala, A), 아스파라긴 (Asn, N), 또는 아스파르트산 (Asp, D)이며, Xaa29는 시스테인 (Cys, C), 글리신 (Gly, G), 글루타민 (Gln, Q), 트레오닌 (Thr, T), 글루탐산 (Glu, E), 또는 히스티딘 (His, H)이고, Xaa30은 시스테인 (Cys, C), 글리신 (Gly, G), 리신 (Lys, K), 또는 히스티딘 (His, H)이거나, 부존재하며, In the general formula 1, Xaa1 is histidine (His, H), 4-imidazoacetyl (CA), or tyrosine (Tyr, Y), and Xaa2 is glycine (Gly, G), alpha-methyl-glutamic acid, or Aib (aminoisobutyric acid), Xaa3 is glutamic acid (Glu, E) or glutamine (Gln, Q), Xaa7 is threonine (Thr, T) or isoleucine (Ile, I), Xaa10 is leucine (Leu, L), tyrosine (Tyr, Y), lysine (Lys, K), cysteine (Cys, C), or valine (Val, V), and Xaa12 is lysine (Lys, K), serine (Ser, S), or isoleucine (Ile, I), Xaa13 is glutamine (Gln, Q), tyrosine (Tyr, Y), alanine (Ala, A), or cysteine (Cys, C), and Xaa14 is leucine (Leu, L), methionine (Met, M ), or tyrosine (Tyr, Y), Xaa15 is cysteine (Cys, C), aspartic acid (Asp, D), glutamic acid (Glu, E), or leucine (Leu, L), and Xaa16 is glycine (Gly, G), glutamic acid (Glu, E), or serine (Ser, S), Xaa17 is glutamine (Gln, Q), arginine (Arg, R), isoleucine (Ile, I), glutamic acid (Glu, E), cysteine (Cys, C), or lysine (Lys, K), Xaa18 is alanine (Ala, A), glutamine (Gln, Q), arginine (Arg, R), or histidine (His, H), and Xaa19 is alanine (Ala, A), glutamine (Gln, Q), cysteine (Cys, C), or valine (Val, V), and Xaa20 is lysine (Lys, K), glutamine (Gln, Q), or arginine (Arg, R), Xaa21 is glutamic acid (Glu, E), glutamine (Gln, Q), leucine (Leu, L), cysteine (Cys, C), or aspartic acid (Asp, D), and Xaa23 is isoleucine (Ile, I) again Is valine (Val, V), Xaa24 is alanine (Ala, A), glutamine (Gln, Q), cysteine (Cys, C), asparagine (Asn, N), aspartic acid (Asp, D), or glutamic acid ( Glu, E), Xaa27 is valine (Val, V), leucine (Leu, L), lysine (Lys, K), or methionine (Met, M), and Xaa28 is cysteine (Cys, C), lysine (Lys , K), alanine (Ala, A), asparagine (Asn, N), or aspartic acid (Asp, D), Xaa29 is cysteine (Cys, C), glycine (Gly, G), glutamine (Gln, Q) , Threonine (Thr, T), glutamic acid (Glu, E), or histidine (His, H), Xaa30 is cysteine (Cys, C), glycine (Gly, G), lysine (Lys, K), or histidine ( His, H) or absent,
R1은 시스테인 (Cys, C), GKKNDWKHNIT (서열번호 106), m-SSGAPPPS-n (서열번호 107), 또는 m-SSGQPPPS-n (서열번호 108)이거나, 부존재하며, 여기서, m은 -Cys-, -Pro-, 또는 -Gly-Pro-이고, n은 -Cys-, -Gly-, -Ser-, 또는 -His-Gly-이거나, 부존재하는 것을 특징으로 한다.R1 is cysteine (Cys, C), GKKNDWKHNIT (SEQ ID NO: 106), m-SSGAPPPS-n (SEQ ID NO: 107), or m-SSGQPPPS-n (SEQ ID NO: 108), or absent, wherein m is -Cys- , -Pro-, or -Gly-Pro-, and n is -Cys-, -Gly-, -Ser-, or -His-Gly-, or it is characterized in that it does not exist.
앞선 구체예 중 어느 하나에 따른 조성물로서, 상기 펩타이드는 상기 일반식 1에서 Xaa14는 류신 또는 메티오닌이며, Xaa15는 시스테인, 아스파르트산, 또는 류신인 것을 특징으로 한다.As the composition according to any one of the preceding embodiments, the peptide is characterized in that in the general formula 1, Xaa14 is leucine or methionine, and Xaa15 is cysteine, aspartic acid, or leucine.
앞선 구체예 중 어느 하나에 따른 조성물로서, 상기 펩타이드는 상기 일반식 1에서, Xaa2는 글리신, 알파-메틸-글루탐산, 또는 Aib이고, Xaa7은 트레오닌이며, Xaa10은 티로신, 시스테인, 또는 발린이고, Xaa12는 리신 또는 이소류신이며, Xaa13은 티로신, 알라닌, 글루타민, 또는 시스테인이고, Xaa14는 류신, 시스테인, 또는 메티오닌이며, Xaa15는 시스테인, 류신, 글루탐산, 또는 아스파르트산이고, Xaa17은 글루타민, 아르기닌, 이소류신, 시스테인, 글루탐산, 또는 리신이며, Xaa18은 알라닌, 글루타민, 아르기닌, 또는 히스티딘이고, Xaa19는 알라닌, 글루타민, 발린, 또는 시스테인이며, Xaa20은 리신, 아르기닌, 또는 글루타민이고, Xaa21은 글루탐산, 글루타민, 류신, 시스테인, 또는 아스파르트산이며, Xaa23은 이소류신 또는 발린이고, Xaa24는 시스테인, 알라닌, 글루타민, 아스파라긴, 글루탐산, 또는 아스파르트산이며, Xaa27은 류신 또는 리신인 것을 특징으로 한다. As a composition according to any one of the preceding embodiments, wherein the peptide is in the general formula 1, Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib, Xaa7 is threonine, Xaa10 is tyrosine, cysteine, or valine, and Xaa12 Is lysine or isoleucine, Xaa13 is tyrosine, alanine, glutamine, or cysteine, Xaa14 is leucine, cysteine, or methionine, Xaa15 is cysteine, leucine, glutamic acid, or aspartic acid, and Xaa17 is glutamine, arginine, isoleucine, cysteine. , Glutamic acid, or lysine, Xaa18 is alanine, glutamine, arginine, or histidine, Xaa19 is alanine, glutamine, valine, or cysteine, Xaa20 is lysine, arginine, or glutamine, and Xaa21 is glutamic acid, glutamine, leucine, cysteine , Or aspartic acid, Xaa23 is isoleucine or valine, Xaa24 is cysteine, alanine, glutamine, asparagine, glutamic acid, or aspartic acid, and Xaa27 is leucine or lysine.
앞선 구체예 중 어느 하나에 따른 조성물로서, 상기 펩타이드는 하기 일반식 2로 표시되는 아미노산 서열을 포함하는 펩타이드인 것을 특징으로 한다. As a composition according to any one of the preceding embodiments, the peptide is characterized in that it is a peptide comprising an amino acid sequence represented by the following general formula 2.
Xaa1-Xaa2-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Xaa10-Ser-Lys-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp-Leu-Leu-Xaa28-Xaa29-Xaa30-Xaa31-Ser-Ser-Gly-Gln-Pro-Pro-Pro-Ser-Xaa40 (일반식 2, 서열번호 104)Xaa1-Xaa2-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Xaa10-Ser-Lys-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp- Leu-Leu-Xaa28-Xaa29-Xaa30-Xaa31-Ser-Ser-Gly-Gln-Pro-Pro-Pro-Ser-Xaa40 (general formula 2, SEQ ID NO: 104)
상기 일반식 2에서, Xaa1은 4-이미다조아세틸, 히스티딘, 또는 티로신이고, Xaa2는 글리신, 알파-메틸-글루탐산, 또는 Aib이며, Xaa10은 티로신, 또는 시스테인이고, Xaa13은 알라닌, 글루타민, 티로신, 또는 시스테인이며, Xaa14는 류신, 메티오닌, 또는 티로신이고, Xaa15는 아스파르트산, 글루탐산, 또는 류신이며, Xaa16은 글리신, 글루탐산, 또는 세린이고, Xaa17은 글루타민, 아르기닌, 이소류신, 글루탐산, 시스테인, 또는 리신이며, Xaa18은 알라닌, 글루타민, 아르기닌, 또는 히스티딘이고, Xaa19는 알라닌, 글루타민, 시스테인, 또는 발린이며, Xaa20은 리신, 글루타민, 또는 아르기닌이고, Xaa21은 시스테인, 글루탐산, 글루타민, 류신, 또는 아스파르트산이며, Xaa23은 이소류신 또는 발린이고, Xaa24는 시스테인, 알라닌, 글루타민, 아스파라긴, 또는 글루탐산이며, Xaa28은 리신, 시스테인, 아스파라긴, 또는 아스파르트산이고, Xaa29는 글리신, 글루타민, 시스테인, 또는 히스티딘이며, Xaa30은 시스테인, 글리신, 리신, 또는 히스티딘이고, Xaa31은 프롤린 또는 시스테인이며, Xaa40은 시스테인이거나, 부존재하는 것을 특징으로 한다.In Formula 2, Xaa1 is 4-imidazoacetyl, histidine, or tyrosine, Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib, Xaa10 is tyrosine, or cysteine, and Xaa13 is alanine, glutamine, tyrosine, Or cysteine, Xaa14 is leucine, methionine, or tyrosine, Xaa15 is aspartic acid, glutamic acid, or leucine, Xaa16 is glycine, glutamic acid, or serine, and Xaa17 is glutamine, arginine, isoleucine, glutamic acid, cysteine, or lysine. , Xaa18 is alanine, glutamine, arginine, or histidine, Xaa19 is alanine, glutamine, cysteine, or valine, Xaa20 is lysine, glutamine, or arginine, and Xaa21 is cysteine, glutamic acid, glutamine, leucine, or aspartic acid, Xaa23 is isoleucine or valine, Xaa24 is cysteine, alanine, glutamine, asparagine, or glutamic acid, Xaa28 is lysine, cysteine, asparagine, or aspartic acid, Xaa29 is glycine, glutamine, cysteine, or histidine, Xaa30 is cysteine, Glycine, lysine, or histidine, Xaa31 is proline or cysteine, and Xaa40 is cysteine or absent.
앞선 구체예 중 어느 하나에 따른 조성물로서, 상기 펩타이드는 상기 일반식 1에서, Xaa2는 글리신, 알파-메틸-글루탐산, 또는 Aib이고, Xaa7은 트레오닌이며, Xaa10은 티로신, 시스테인, 또는 발린이고, Xaa12는 리신 또는 이소류신이며, Xaa13은 티로신, 알라닌, 또는 시스테인이고, Xaa14는 류신 또는 메티오닌이며, Xaa15는 시스테인 또는 아스파르트산이고, Xaa17은 글루타민, 아르기닌, 이소류신, 시스테인, 또는 리신이며, Xaa18은 알라닌, 아르기닌, 또는 히스티딘이고, Xaa19는 알라닌, 글루타민, 또는 시스테인이며, Xaa20은 리신 또는 글루타민이고, Xaa21은 글루탐산, 시스테인, 또는 아스파르트산이며, Xaa23은 발린이고, Xaa24는 알라닌, 글루타민, 시스테인, 아스파라긴, 또는 아스파르트산이며, Xaa27은 류신 또는 리신인 것을 특징으로 한다. As a composition according to any one of the preceding embodiments, wherein the peptide is in the general formula 1, Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib, Xaa7 is threonine, Xaa10 is tyrosine, cysteine, or valine, and Xaa12 Is lysine or isoleucine, Xaa13 is tyrosine, alanine, or cysteine, Xaa14 is leucine or methionine, Xaa15 is cysteine or aspartic acid, Xaa17 is glutamine, arginine, isoleucine, cysteine, or lysine, and Xaa18 is alanine, arginine. , Or histidine, Xaa19 is alanine, glutamine, or cysteine, Xaa20 is lysine or glutamine, Xaa21 is glutamic acid, cysteine, or aspartic acid, Xaa23 is valine, Xaa24 is alanine, glutamine, cysteine, asparagine, or aspartic acid. It is an acid, and Xaa27 is characterized in that it is leucine or lysine.
앞선 구체예 중 어느 하나에 따른 조성물로서, 상기 펩타이드는 상기 일반식 2에서, Xaa13은 알라닌, 티로신, 또는 시스테인이고, Xaa15는 아스파르트산 또는 글루탐산이며, Xaa17은 글루타민, 아르기닌, 시스테인, 또는 리신이고, Xaa18은 알라닌, 아르기닌, 또는 히스티딘이며, Xaa21은 시스테인, 글루탐산, 글루타민, 또는 아스파르트산이고, Xaa23은 이소류신 또는 발린이며, Xaa24는 시스테인, 글루타민, 또는 아스파라긴이고, Xaa28은 시스테인, 아스파라긴, 또는 아스파르트산이며, Xaa29는 글루타민, 시스테인, 또는 히스티딘이고, Xaa30은 시스테인, 리신, 또는 히스티딘인 것을 특징으로 한다. As a composition according to any one of the preceding embodiments, wherein the peptide is in the general formula 2, Xaa13 is alanine, tyrosine, or cysteine, Xaa15 is aspartic acid or glutamic acid, and Xaa17 is glutamine, arginine, cysteine, or lysine, Xaa18 is alanine, arginine, or histidine, Xaa21 is cysteine, glutamic acid, glutamine, or aspartic acid, Xaa23 is isoleucine or valine, Xaa24 is cysteine, glutamine, or asparagine, and Xaa28 is cysteine, asparagine, or aspartic acid. , Xaa29 is glutamine, cysteine, or histidine, and Xaa30 is cysteine, lysine, or histidine.
앞선 구체예 중 어느 하나에 따른 조성물로서, 상기 펩타이드는 상기 일반식 1에서, Xaa2는 알파-메틸-글루탐산 또는 Aib이고, Xaa7은 트레오닌이며, Xaa10은 티로신 또는 시스테인이고, Xaa12는 리신 또는 이소류신이며, Xaa13은 티로신, 알라닌, 또는 시스테인이고, Xaa14는 류신 또는 메티오닌이며, Xaa15는 시스테인 또는 아스파르트산이고, Xaa16은 글루탐산이며, Xaa17은 아르기닌, 이소류신, 시스테인, 또는 리신이고, Xaa18은 알라닌, 아르기닌, 또는 히스티딘이며, Xaa19는 알라닌, 글루타민, 또는 시스테인이고, Xaa20은 리신 또는 글루타민이며, Xaa21은 글루탐산 또는 아스파르트산이고, Xaa23은 발린이며, Xaa24는 글루타민, 아스파라긴, 또는 아스파르트산이고, Xaa27은 류신이며, Xaa28은 시스테인, 알라닌, 아스파라긴, 또는 아스파르트산인 것을 특징으로 한다. As a composition according to any one of the preceding embodiments, wherein the peptide is in the general formula 1, Xaa2 is alpha-methyl-glutamic acid or Aib, Xaa7 is threonine, Xaa10 is tyrosine or cysteine, Xaa12 is lysine or isoleucine, Xaa13 is tyrosine, alanine, or cysteine, Xaa14 is leucine or methionine, Xaa15 is cysteine or aspartic acid, Xaa16 is glutamic acid, Xaa17 is arginine, isoleucine, cysteine, or lysine, and Xaa18 is alanine, arginine, or histidine. Xaa19 is alanine, glutamine, or cysteine, Xaa20 is lysine or glutamine, Xaa21 is glutamic acid or aspartic acid, Xaa23 is valine, Xaa24 is glutamine, asparagine, or aspartic acid, Xaa27 is leucine, and Xaa28 is It is characterized in that it is cysteine, alanine, asparagine, or aspartic acid.
앞선 구체예 중 어느 하나에 따른 조성물로서, 상기 펩타이드는 상기 일반식 1에서, Xaa1은 히스티딘 또는 4-이미다조아세틸이고, Xaa2는 알파-메틸-글루탐산 또는 Aib이며, Xaa3은 글루타민이고, Xaa7은 트레오닌이며, Xaa10은 티로신이고, Xaa12는 이소류신이며, Xaa13은 알라닌 또는 시스테인이고, Xaa14는 메티오닌이며, Xaa15는 아스파르트산이고, Xaa16은 글루탐산이며, Xaa17은 이소류신 또는 리신이고, Xaa18은 알라닌 또는 히스티딘이며, Xaa19는 글루타민 또는 시스테인이고, Xaa20은 리신이며, Xaa21은 아스파르트산이고, Xaa23은 발린이며, Xaa24는 아스파라긴이고, Xaa27은 류신이며, Xaa28은 알라닌 또는 아스파라긴이고, Xaa29는 글루타민 또는 트레오닌이며, Xaa30은 시스테인 또는 리신이거나, 부존재하는 것을 특징으로 한다. As the composition according to any one of the preceding embodiments, wherein the peptide is in the general formula 1, Xaa1 is histidine or 4-imidazoacetyl, Xaa2 is alpha-methyl-glutamic acid or Aib, Xaa3 is glutamine, and Xaa7 is threonine. , Xaa10 is tyrosine, Xaa12 is isoleucine, Xaa13 is alanine or cysteine, Xaa14 is methionine, Xaa15 is aspartic acid, Xaa16 is glutamic acid, Xaa17 is isoleucine or lysine, Xaa18 is alanine or histidine, Xaa19 Is glutamine or cysteine, Xaa20 is lysine, Xaa21 is aspartic acid, Xaa23 is valine, Xaa24 is asparagine, Xaa27 is leucine, Xaa28 is alanine or asparagine, Xaa29 is glutamine or threonine, Xaa30 is cysteine or It is characterized by being lysine or absent.
앞선 구체예 중 어느 하나에 따른 조성물로서, 상기 펩타이드는 하기 일반식 3의 아미노산 서열을 포함하는 펩타이드인 것을 특징으로 한다. As a composition according to any one of the preceding embodiments, the peptide is characterized in that it is a peptide comprising the amino acid sequence of the following general formula 3.
Xaa1-Xaa2-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-Lys-Xaa13-Leu-Asp-Glu-Xaa17-Xaa18-Xaa19-Lys-Xaa21-Phe-Val-Xaa24-Trp-Leu-Leu-Xaa28-Xaa29-Xaa30-Xaa31-Ser-Ser-Gly-Gln-Pro-Pro-Pro-Ser-Xaa40 (일반식 3, 서열번호 105),Xaa1-Xaa2-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-Lys-Xaa13-Leu-Asp-Glu-Xaa17-Xaa18-Xaa19-Lys-Xaa21-Phe-Val-Xaa24-Trp- Leu-Leu-Xaa28-Xaa29-Xaa30-Xaa31-Ser-Ser-Gly-Gln-Pro-Pro-Pro-Ser-Xaa40 (general formula 3, SEQ ID NO: 105),
상기 일반식 3에서, Xaa1은 히스티딘 또는 티로신이고, Xaa2는 알파-메틸-글루탐산 또는 Aib이며, Xaa13은 알라닌, 티로신 또는 시스테인이고, Xaa17은 아르기닌, 시스테인, 또는 리신이며, Xaa18은 알라닌 또는 아르기닌이고, Xaa19는 알라닌 또는 시스테인이며, Xaa21은 글루탐산 또는 아스파르트산이고, Xaa24는 글루타민 또는 아스파라긴이며, Xaa28은 시스테인 또는 아스파르트산이고, Xaa29는 시스테인, 히스티딘, 또는 글루타민이며, Xaa30은 시스테인 또는 히스티딘이고, Xaa31은 프롤린 또는 시스테인이며, Xaa40은 시스테인 또는 부존재하는 것을 특징으로 한다.In the general formula 3, Xaa1 is histidine or tyrosine, Xaa2 is alpha-methyl-glutamic acid or Aib, Xaa13 is alanine, tyrosine or cysteine, Xaa17 is arginine, cysteine, or lysine, and Xaa18 is alanine or arginine, Xaa19 is alanine or cysteine, Xaa21 is glutamic acid or aspartic acid, Xaa24 is glutamine or asparagine, Xaa28 is cysteine or aspartic acid, Xaa29 is cysteine, histidine, or glutamine, Xaa30 is cysteine or histidine, Xaa31 is proline Or cysteine, and Xaa40 is cysteine or absent.
앞선 구체예 중 어느 하나에 따른 조성물로서, 상기 펩타이드는 상기 R1은 시스테인, GKKNDWKHNIT (서열번호 106), CSSGQPPPS (서열번호 109), GPSSGAPPPS (서열번호 110), GPSSGAPPPSC (서열번호 111), PSSGAPPPS (서열번호 112), PSSGAPPPSG (서열번호 113), PSSGAPPPSHG (서열번호 114), PSSGAPPPSS (서열번호 115), PSSGQPPPS (서열번호 116), 또는 PSSGQPPPSC (서열번호 117)이거나, 부존재하는 것을 특징으로 한다. As a composition according to any one of the preceding embodiments, wherein the peptide is a cysteine, GKKNDWKHNIT (SEQ ID NO: 106), CSSGQPPPS (SEQ ID NO: 109), GPSSGAPPPS (SEQ ID NO: 110), GPSSGAPPPSC (SEQ ID NO: 111), PSSGAPPPS (SEQ ID NO: Number 112), PSSGAPPPSG (SEQ ID NO: 113), PSSGAPPPSHG (SEQ ID NO: 114), PSSGAPPPSS (SEQ ID NO: 115), PSSGQPPPS (SEQ ID NO: 116), or PSSGQPPPSC (SEQ ID NO: 117), or absent.
앞선 구체예 중 어느 하나에 따른 조성물로서, 상기 펩타이드는 서열번호: 1 내지 102로 이루어진 군으로부터 선택된 아미노산 서열을 포함하는 것을 특징으로 한다. The composition according to any one of the preceding embodiments, wherein the peptide comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 1 to 102.
앞선 구체예 중 어느 하나에 따른 조성물로서, 상기 일반식에서 N-말단으로부터 16번 아미노산과 20번 아미노산은 서로 고리를 형성하는 것을 특징으로 한다. A composition according to any one of the preceding embodiments, wherein amino acids 16 and 20 from the N-terminus in the general formula form a ring with each other.
앞선 구체예 중 어느 하나에 따른 조성물로서, 상기 펩타이드는 이의 C-말단이 아미드화된 것을 특징으로 한다. A composition according to any one of the preceding embodiments, wherein the peptide is amidated at its C-terminus.
앞선 구체예 중 어느 하나에 따른 조성물로서, 상기 약학적 조성물은 LDL 흡수 증가, 3-Hydroxy-3-methylglutaryl-CoA reductase (HMGCR) 활성 억제, 및 지방산 분해 촉진 효과 중 어느 하나 이상의 효과를 갖는 것을 특징으로 한다. The composition according to any one of the preceding embodiments, wherein the pharmaceutical composition has any one or more of an effect of increasing LDL absorption, inhibiting 3-Hydroxy-3-methylglutaryl-CoA reductase (HMGCR) activity, and promoting fatty acid degradation. To do.
본 발명을 구현하는 다른 하나의 양태는 상기 펩타이드를 포함하는 결합체를 포함하는 고지혈증의 예방 또는 치료를 위한 약학적 조성물이다.Another aspect embodying the present invention is a pharmaceutical composition for the prevention or treatment of hyperlipidemia comprising a conjugate comprising the peptide.
하나의 구체예로서, 상기 결합체는 하기 화학식 1로 표시되는 것을 특징으로 할 수 있다: In one embodiment, the conjugate may be characterized in that it is represented by the following Formula 1:
[화학식 1][Formula 1]
X - La - FX-La-F
여기서,here,
X는, 상기 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는, 펩타이드이고,X is a peptide having activity against the glucagon receptor, GLP-1 receptor, and GIP receptor,
L은, 폴리에틸렌 글리콜이며,L is polyethylene glycol,
a는, 0 또는 자연수이며, 다만 a가 2 이상일 때, 각각의 L은 서로 독립적이고,a is 0 or a natural number, provided that when a is 2 or more, each L is independent of each other,
F는, 면역글로불린 Fc 영역임.F is an immunoglobulin Fc region.
다른 구체예로서, 상기 펩타이드는 하기 일반식 1로 표시되는 아미노산 서열을 포함하는 펩타이드인 것을 특징으로 한다:In another embodiment, the peptide is characterized in that it is a peptide comprising an amino acid sequence represented by the following general formula 1:
Xaa1-Xaa2-Xaa3-Gly-Thr-Phe-Xaa7-Ser-Asp-Xaa10-Ser-Xaa12-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp-Leu-Xaa27-Xaa28-Xaa29-Xaa30-R1 (일반식 1, 서열번호 103)Xaa1-Xaa2-Xaa3-Gly-Thr-Phe-Xaa7-Ser-Asp-Xaa10-Ser-Xaa12-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp- Leu-Xaa27-Xaa28-Xaa29-Xaa30-R1 (general formula 1, SEQ ID NO: 103)
상기 일반식 1에서,In the general formula 1,
Xaa1은 히스티딘 (His, H), 4-이미다조아세틸 (CA), 또는 티로신 (Tyr, Y)이고, Xaa1 is histidine (His, H), 4-imidazoacetyl (CA), or tyrosine (Tyr, Y),
Xaa2는 글리신 (Gly, G), 알파-메틸-글루탐산, 또는 Aib (aminoisobutyric acid)이며,Xaa2 is glycine (Gly, G), alpha-methyl-glutamic acid, or Aib (aminoisobutyric acid),
Xaa3은 글루탐산 (Glu, E) 또는 글루타민 (Gln, Q)이고, Xaa3 is glutamic acid (Glu, E) or glutamine (Gln, Q),
Xaa7은 트레오닌 (Thr, T) 또는 이소류신 (Ile, I)이며,Xaa7 is threonine (Thr, T) or isoleucine (Ile, I),
Xaa10은 류신 (Leu, L), 티로신 (Tyr, Y), 리신 (Lys, K), 시스테인 (Cys, C), 또는 발린 (Val, V)이고, Xaa10 is leucine (Leu, L), tyrosine (Tyr, Y), lysine (Lys, K), cysteine (Cys, C), or valine (Val, V),
Xaa12는 리신 (Lys, K), 세린 (Ser, S), 또는 이소류신 (Ile, I)이며, Xaa12 is lysine (Lys, K), serine (Ser, S), or isoleucine (Ile, I),
Xaa13은 글루타민 (Gln, Q), 티로신 (Tyr, Y), 알라닌 (Ala, A), 또는 시스테인 (Cys, C)이고, Xaa13 is glutamine (Gln, Q), tyrosine (Tyr, Y), alanine (Ala, A), or cysteine (Cys, C),
Xaa14는 류신 (Leu, L), 메티오닌 (Met, M), 또는 티로신 (Tyr, Y)이며,Xaa14 is leucine (Leu, L), methionine (Met, M), or tyrosine (Tyr, Y),
Xaa15는 시스테인 (Cys, C), 아스파르트산 (Asp, D), 글루탐산 (Glu, E), 또는 류신 (Leu, L)이며,Xaa15 is cysteine (Cys, C), aspartic acid (Asp, D), glutamic acid (Glu, E), or leucine (Leu, L),
Xaa16은 글리신 (Gly, G), 글루탐산 (Glu, E), 또는 세린 (Ser, S)이고, Xaa16 is glycine (Gly, G), glutamic acid (Glu, E), or serine (Ser, S),
Xaa17은 글루타민 (Gln, Q), 아르기닌 (Arg, R), 이소류신 (Ile, I), 글루탐산 (Glu, E), 시스테인 (Cys, C), 또는 리신 (Lys, K)이며,Xaa17 is glutamine (Gln, Q), arginine (Arg, R), isoleucine (Ile, I), glutamic acid (Glu, E), cysteine (Cys, C), or lysine (Lys, K),
Xaa18은 알라닌 (Ala, A), 글루타민 (Gln, Q), 아르기닌 (Arg, R), 또는 히스티딘 (His, H)이고,Xaa18 is alanine (Ala, A), glutamine (Gln, Q), arginine (Arg, R), or histidine (His, H),
Xaa19는 알라닌 (Ala, A), 글루타민 (Gln, Q), 시스테인 (Cys, C), 또는 발린 (Val, V)이며,Xaa19 is alanine (Ala, A), glutamine (Gln, Q), cysteine (Cys, C), or valine (Val, V),
Xaa20은 리신 (Lys, K), 글루타민 (Gln, Q), 또는 아르기닌 (Arg, R)이고,Xaa20 is lysine (Lys, K), glutamine (Gln, Q), or arginine (Arg, R),
Xaa21은 글루탐산 (Glu, E), 글루타민 (Gln, Q), 류신 (Leu, L), 시스테인 (Cys, C), 또는 아스파르트산 (Asp, D)이며,Xaa21 is glutamic acid (Glu, E), glutamine (Gln, Q), leucine (Leu, L), cysteine (Cys, C), or aspartic acid (Asp, D),
Xaa23은 이소류신 (Ile, I) 또는 발린 (Val, V)이고,Xaa23 is isoleucine (Ile, I) or valine (Val, V),
Xaa24는 알라닌 (Ala, A), 글루타민 (Gln, Q), 시스테인 (Cys, C), 아스파라긴 (Asn, N), 아스파르트산 (Asp, D), 또는 글루탐산 (Glu, E)이며,Xaa24 is alanine (Ala, A), glutamine (Gln, Q), cysteine (Cys, C), asparagine (Asn, N), aspartic acid (Asp, D), or glutamic acid (Glu, E),
Xaa27은 발린 (Val, V), 류신 (Leu, L), 리신 (Lys, K), 또는 메티오닌 (Met, M)이고,Xaa27 is valine (Val, V), leucine (Leu, L), lysine (Lys, K), or methionine (Met, M),
Xaa28은 시스테인 (Cys, C), 리신 (Lys, K), 알라닌 (Ala, A), 아스파라긴 (Asn, N), 또는 아스파르트산 (Asp, D)이며,Xaa28 is cysteine (Cys, C), lysine (Lys, K), alanine (Ala, A), asparagine (Asn, N), or aspartic acid (Asp, D),
Xaa29는 시스테인 (Cys, C), 글리신 (Gly, G), 글루타민 (Gln, Q), 트레오닌 (Thr, T), 글루탐산 (Glu, E), 또는 히스티딘 (His, H)이고,Xaa29 is cysteine (Cys, C), glycine (Gly, G), glutamine (Gln, Q), threonine (Thr, T), glutamic acid (Glu, E), or histidine (His, H),
Xaa30은 시스테인 (Cys, C), 글리신 (Gly, G), 리신 (Lys, K), 또는 히스티딘 (His, H)이거나, 부존재하며,Xaa30 is cysteine (Cys, C), glycine (Gly, G), lysine (Lys, K), or histidine (His, H) or absent,
R1은 시스테인 (Cys, C), GKKNDWKHNIT (서열번호 106), m-SSGAPPPS-n (서열번호 107), 또는 m-SSGQPPPS-n (서열번호 108)이거나, 부존재하며, R1 is cysteine (Cys, C), GKKNDWKHNIT (SEQ ID NO: 106), m-SSGAPPPS-n (SEQ ID NO: 107), or m-SSGQPPPS-n (SEQ ID NO: 108), or absent,
여기서, here,
m은 -Cys-, -Pro-, 또는 -Gly-Pro-이고, m is -Cys-, -Pro-, or -Gly-Pro-,
n은 -Cys-, -Gly-, -Ser-, 또는 -His-Gly-이거나, 부존재함. n is -Cys-, -Gly-, -Ser-, or -His-Gly- or absent.
앞선 구체예 중 어느 하나에 따른 조성물로서, 상기 약학적 조성물은 상기 일반식 1로 표현되는 펩타이드가 폴리에틸렌 글리콜을 통해 면역글로불린 Fc 영역과 결합한 결합체를 포함하는 것을 특징으로 한다.As the composition according to any one of the preceding embodiments, the pharmaceutical composition is characterized in that it comprises a conjugate in which the peptide represented by the general formula 1 is bound to an immunoglobulin Fc region through polyethylene glycol.
앞선 구체예 중 어느 하나에 따른 조성물로서, 상기 약학적 조성물은 LDL 흡수 증가, 3-Hydroxy-3-methylglutaryl-CoA reductase (HMGCR) 활성 억제, 및 지방산 분해 촉진 효과 중 어느 하나 이상의 효과를 갖는 것을 특징으로 한다. The composition according to any one of the preceding embodiments, wherein the pharmaceutical composition has any one or more of an effect of increasing LDL absorption, inhibiting 3-Hydroxy-3-methylglutaryl-CoA reductase (HMGCR) activity, and promoting fatty acid degradation. To do.
본 발명을 구현하는 다른 하나의 양태는 상기 펩타이드, 결합체 또는 이를 포함하는 조성물을 이를 필요로 하는 개체에 투여하는 단계를 포함하는, 고지혈증의 예방 또는 치료 방법이다. Another embodiment of the present invention is a method for preventing or treating hyperlipidemia, comprising administering the peptide, conjugate, or a composition containing the same to an individual in need thereof.
본 발명을 구현하는 다른 하나의 양태는 고지혈증의 예방 또는 치료를 위한 약제의 제조에 있어서 상기 펩타이드, 결합체 또는 이를 포함하는 조성물의 용도이다. Another aspect embodying the present invention is the use of the peptide, conjugate, or a composition comprising the same in the manufacture of a medicament for the prevention or treatment of hyperlipidemia.
본 발명에 따른 삼중 활성체 지속형 결합체는 글루카곤 수용체, GLP-1 (Glucagon-like peptide-1) 수용체, 및 GIP (Glucose-dependent insuliontropic polypeptide) 수용체에 대해 활성을 가져, 고지혈증 치료에 적용될 수 있다. The triple activator long-acting conjugate according to the present invention has activity against the glucagon receptor, the GLP-1 (Glucagon-like peptide-1) receptor, and the GIP (Glucose-dependent insuliontropic polypeptide) receptor, and can be applied to the treatment of hyperlipidemia.
도 1은 본 발명의 삼중 활성체 지속형 결합체의 고지혈증 치료 효과를 in vivo에서 확인한 도이다.1 is a diagram confirming in vivo the effect of treating hyperlipidemia of the triple activator long-acting conjugate of the present invention.
도 2는 본 발명의 삼중 활성체 지속형 결합체에 의해 LDL 흡수가 증가한 것을 확인한 도이다.Figure 2 is a diagram confirming that LDL absorption is increased by the triple-active agent long-acting conjugate of the present invention.
도 3은 본 발명의 삼중 활성체 지속형 결합체에 의해 3-Hydroxy-3-methylglutaryl-CoA reductase (HMGCR) 활성이 억제된 것을 확인한 도이다.3 is a diagram confirming that the activity of 3-Hydroxy-3-methylglutaryl-CoA reductase (HMGCR) was inhibited by the triple-active conjugate of the present invention.
도 4는 본 발명의 삼중 활성체 지속형 결합체의 지방산 분해 촉진 효과를 확인한 도이다.Figure 4 is a diagram confirming the effect of promoting fatty acid decomposition of the triple active substance long-acting conjugate of the present invention.
이하에서는, 본 발명을 더욱 상세히 설명한다.Hereinafter, the present invention will be described in more detail.
한편, 본원에서 개시되는 각각의 설명 및 실시형태는 각각의 다른 설명 및 실시 형태에도 적용될 수 있다. 즉, 본원에서 개시된 다양한 요소들의 모든 조합이 본 발명의 범주에 속한다. 또한, 하기 기술되는 구체적인 서술에 의하여 본 발명의 범주가 제한된다고 할 수 없다.On the other hand, each description and embodiment disclosed herein can be applied to each other description and embodiment. That is, all combinations of various elements disclosed herein fall within the scope of the present invention. In addition, it cannot be said that the scope of the present invention is limited by the specific description described below.
본 명세서 전반을 통하여, 천연적으로 존재하는 아미노산에 대한 통상의 1문자 및 3문자 코드가 사용될 뿐만 아니라 Aib (α-아미노이소부티르산), Sar (N-methylglycine), 알파-메틸-글루탐산 (α-methyl-glutamic acid) 등과 같은 다른 아미노산에 대해 일반적으로 허용되는 3문자 코드가 사용된다. 또한 본 명세서에서 약어로 언급된 아미노산은 IUPAC-IUB 명명법에 따라 기재되었다. Throughout this specification, not only the usual 1-letter and 3-letter codes for naturally occurring amino acids are used, but also Aib (α-aminoisobutyric acid), Sar (N-methylglycine), alpha-methyl-glutamic acid (α- For other amino acids, such as methyl-glutamic acid), the generally accepted three letter code is used. In addition, amino acids referred to as abbreviations herein have been described according to the IUPAC-IUB nomenclature.
알라닌 Ala, A 아르기닌 Arg, RAlanine Ala, A Arginine Arg, R
아스파라긴 Asn, N 아스파르트산 Asp, DAsparagine Asn, N Aspartic Acid Asp, D
시스테인 Cys, C 글루탐산 Glu, ECysteine Cys, C Glutamic acid Glu, E
글루타민 Gln, Q 글리신 Gly, GGlutamine Gln, Q Glycine Gly, G
히스티딘 His, H 이소류신 Ile, IHistidine His, H isoleucine Ile, I
류신 Leu, L 리신 Lys, KLeucine Leu, L Lysine Lys, K
메티오닌 Met, M 페닐알라닌 Phe, FMethionine Met, M Phenylalanine Phe, F
프롤린 Pro, P 세린 Ser, SProline Pro, P Serine Ser, S
트레오닌 Thr, T 트립토판 Trp, WThreonine Thr, T Tryptophan Trp, W
티로신 Tyr, Y 발린 Val, VTyrosine Tyr, Y valine Val, V
본 발명을 구현하기 위한 하나의 양태는 글루카곤 수용체, GLP-1 (Glucagon-like peptide-1) 수용체, 및 GIP (Glucose-dependent insuliontropic polypeptide) 수용체에 대해 활성을 갖는, 펩타이드를 포함하는, 고지혈증의 예방 또는 치료용 약학적 조성물이다. One aspect for embodying the present invention is the prevention of hyperlipidemia, including a peptide having activity against glucagon receptor, GLP-1 (Glucagon-like peptide-1) receptor, and GIP (Glucose-dependent insuliontropic polypeptide) receptor Or a therapeutic pharmaceutical composition.
본 발명의 삼중활성체는 고지혈증에 대한 치료 또는 예방 효과가 있는 것을 특징으로 한다. 본 발명에서, 고지혈증은 혈액 내에 콜레스테롤과 중성지방 수치가 정상범위에서 벗어나 있는(증가 및 감소 모두 포함) 상태인 이상지질혈증과 혈액 내에 총 콜레스테롤이 높은 상태인 고콜레스테롤혈증과 달리, 혈액 내에 저밀도지단백 콜레스테롤이나 중성지방 중 한가지가 높거나 둘 다 높은 경우를 총칭하는 질환으로서 이상지질혈증 및 고콜레스테롤증과 구분되는 질환이다.The triple activator of the present invention is characterized by having a therapeutic or preventive effect on hyperlipidemia. In the present invention, hyperlipidemia, unlike dyslipidemia, in which cholesterol and triglyceride levels in the blood are out of the normal range (including both increase and decrease) and hypercholesterolemia, in which total cholesterol in the blood is high, is low-density lipoprotein in the blood. It is a disease that is a generic term for a case in which one or both of cholesterol or triglycerides is high, and is a disease distinguished from dyslipidemia and hypercholesterolosis.
상기 "글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는, 펩타이드"는 본 발명에서 삼중 활성체로 혼용되어 사용될 수 있다. The "glucagon receptor, GLP-1 receptor, and peptide having activity against the GIP receptor" may be used interchangeably as a triple activator in the present invention.
이러한 펩타이드는 글루카곤, GLP-1, 및 GIP 수용체에 대해 유의한 수준의 활성을 가지는 다양한 물질, 예컨대 다양한 펩타이드를 포함한다.Such peptides include various substances, such as various peptides, with significant levels of activity against glucagon, GLP-1, and GIP receptors.
특별히 이에 제한되는 것은 아니나, 상기 글루카곤, GLP-1, 및 GIP 수용체에 대해 유의한 수준의 활성을 가지는 삼중 활성체는 글루카곤, GLP-1, 및 GIP 수용체 중 하나 또는 그 이상의 수용체, 구체적으로 둘 또는 그 이상의 수용체, 보다 구체적으로 세 개의 수용체 모두에 대해 in vitro 활성이 해당 수용체의 천연형 리간드 (천연형 글루카곤, 천연형 GLP-1, 및 천연형 GIP) 대비 0.1% 이상, 1% 이상, 2% 이상, 3 % 이상, 4% 이상, 5% 이상, 6% 이상, 7% 이상, 8% 이상, 9% 이상, 10 % 이상, 20% 이상, 30% 이상, 40% 이상, 50 % 이상, 60% 이상, 70% 이상, 80% 이상, 90 % 이상, 또는 100% 이상을 나타낼 수 있다.Although not particularly limited thereto, the triple activator having a significant level of activity against glucagon, GLP-1, and GIP receptors is one or more of glucagon, GLP-1, and GIP receptors, specifically two or more. More receptors, more specifically, in vitro activity against all three receptors is 0.1% or more, 1% or more, 2% compared to the natural ligands of the receptor (natural glucagon, natural GLP-1, and natural GIP). More than, 3% or more, 4% or more, 5% or more, 6% or more, 7% or more, 8% or more, 9% or more, 10% or more, 20% or more, 30% or more, 40% or more, 50% or more, It may represent 60% or more, 70% or more, 80% or more, 90% or more, or 100% or more.
이러한 삼중 활성체의 in vitro 활성을 측정하는 방법은 본원 명세서의 실험예 1을 참조할 수 있으나, 특별히 이에 제한되는 것은 아니다. A method of measuring the in vitro activity of such a triple activator may refer to Experimental Example 1 of the present specification, but is not particularly limited thereto.
한편, 상기 펩타이드는 하기 i) 내지 iii) 중 하나 이상, 둘 이상, 구체적으로 세 개의 활성을 보유하는 것, 구체적으로 유의한 활성을 보유하는 것을 특징으로 한다:On the other hand, the peptide is characterized by having one or more, two or more, specifically three activities, specifically significant activities of the following i) to iii):
i) GLP-1 수용체의 활성화; ii) 글루카곤 수용체의 활성화; 및 iii) GIP 수용체의 활성화.i) activation of the GLP-1 receptor; ii) activation of the glucagon receptor; And iii) activation of the GIP receptor.
여기서, 수용체를 활성화시킨다는 것은, 천연형 대비 수용체에 대한 in vitro 활성이 0.1% 이상, 1% 이상, 2% 이상, 3 % 이상, 4% 이상, 5% 이상, 6% 이상, 7% 이상, 8% 이상, 9% 이상, 10 % 이상, 20% 이상, 30% 이상, 40% 이상, 50 % 이상, 60% 이상, 70% 이상, 80% 이상, 90 % 이상, 또는 100% 이상을 나타내는 경우를 예로 들 수 있다. 그러나, 이에 제한되는 것은 아니다.Here, activating the receptor means that the in vitro activity against the receptor is 0.1% or more, 1% or more, 2% or more, 3% or more, 4% or more, 5% or more, 6% or more, 7% or more, 8% or more, 9% or more, 10% or more, 20% or more, 30% or more, 40% or more, 50% or more, 60% or more, 70% or more, 80% or more, 90% or more, or 100% or more An example is the case. However, it is not limited thereto.
본 발명에 따른 조성물은 글루카곤 수용체, GLP-1 (Glucagon-like peptide-1) 수용체, 및 GIP (Glucose-dependent insuliontropic polypeptide) 수용체에 대해 활성을 갖는 분리된 펩타이드를 포함함으로써, LDL 흡수 증가, 콜레스테롤 합성 억제, 및 지방산 분해 촉진 효과 중 어느 하나 이상의 효과를 나타낼 수 있고, 그에 따라 고지혈증에 대해 우수한 치료 효과를 나타낼 수 있다. 예컨대, 상기 조성물 투여 후 24시간 내지 48시간 후의 3-Hydroxy-3-methylglutaryl-CoA reductase (HMGCR)의 활성은 50% 미만일 수 있다.The composition according to the present invention contains an isolated peptide having activity against a glucagon receptor, a GLP-1 (Glucagon-like peptide-1) receptor, and a GIP (Glucose-dependent insuliontropic polypeptide) receptor, thereby increasing LDL absorption and synthesizing cholesterol. It may exhibit an effect of any one or more of the inhibitory and fatty acid decomposition promoting effects, thereby exhibiting an excellent therapeutic effect for hyperlipidemia. For example, the activity of 3-Hydroxy-3-methylglutaryl-CoA reductase (HMGCR) 24 to 48 hours after administration of the composition may be less than 50%.
또한, 상기 펩타이드는 천연형 GLP-1, 천연형 글루카곤 및 천연형 GIP 중 어느 하나 대비 체내 반감기가 증가된 것일 수 있으나, 특별히 이에 제한되는 것은 아니다. In addition, the peptide may have an increased half-life in the body compared to any one of natural GLP-1, natural glucagon, and natural GIP, but is not particularly limited thereto.
특별히 이에 제한되는 것은 아니나, 이러한 상기 펩타이드는 비자연적으로 발생된 (non-naturally occurring) 것일 수 있다.Although not particularly limited thereto, such a peptide may be non-naturally occurring.
구체적으로, 상기 분리된 펩타이드는 천연형 글루카곤의 아날로그일 수 있으나, 특별히 이에 제한되는 것은 아니다. Specifically, the isolated peptide may be an analog of natural glucagon, but is not particularly limited thereto.
본 발명에 따른 천연형 글루카곤 아날로그는 천연형 글루카곤과 비교하여 아미노산 서열에 하나 이상의 차이가 있는 펩타이드, 천연형 글루카곤 서열의 개질 (modification)을 통하여 변형시킨 펩타이드, 천연형 글루카곤의 모방체를 포함한다.The natural-type glucagon analog according to the present invention includes a peptide having one or more differences in amino acid sequence compared to the natural-type glucagon, a peptide modified through modification of the natural-type glucagon sequence, and a mimic of natural-type glucagon.
한편, 특별히 이에 제한되는 것은 아니나, 천연형 글루카곤은 다음의 아미노산 서열을 가질 수 있다: Meanwhile, although not particularly limited thereto, the native glucagon may have the following amino acid sequence:
His-Ser-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-Lys-Tyr-Leu-Asp-Ser-Arg-Arg-Ala-Gln-Asp-Phe-Val-Gln-Trp-Leu-Met-Asn-Thr (서열번호: 118)His-Ser-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-Lys-Tyr-Leu-Asp-Ser-Arg-Arg-Ala-Gln-Asp-Phe-Val-Gln-Trp- Leu-Met-Asn-Thr (SEQ ID NO: 118)
구체적으로, 상기 펩타이드는 천연형 글루카곤 서열에서 적어도 하나 이상의 아미노산에 치환 (substitution), 추가 (addition), 제거 (deletion), 수식 (modification) 및 이들의 조합으로 이루어지는 군에서 선택된 변형이 일어난, 천연형 글루카곤의 아날로그일 수 있으나, 특별히 이에 제한되는 것은 아니다. Specifically, the peptide is selected from the group consisting of substitution, addition, deletion, modification, and combinations thereof in at least one amino acid in the natural glucagon sequence. It may be an analog of glucagon, but is not particularly limited thereto.
또한, 상기 아미노산의 치환은 아미노산으로의 치환이나 비 천연형 화합물로의 치환을 모두 포함한다. In addition, the substitution of the amino acid includes both substitution with amino acids and substitution with non-natural compounds.
또한, 추가는 펩타이드의 N-말단 및/또는 C-말단에 이루어질 수 있다. 한편, 추가되는 아미노산의 길이는 특별히 이에 제한되지 않으며, 1 이상, 2 이상, 3 이상, 4 이상, 5 이상, 6 이상, 7 이상, 8 이상, 9 이상, 10 이상, 또는 11 이상의 아미노산이 추가될 수 있으며, 넓게는 폴리펩타이드의 추가를 포함하나, 특별히 이에 제한되는 것은 아니다. Further, addition may be made at the N-terminus and/or C-terminus of the peptide. Meanwhile, the length of the added amino acid is not particularly limited thereto, and 1 or more, 2 or more, 3 or more, 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, 9 or more, 10 or more, or 11 or more amino acids are added It may be, and broadly includes the addition of a polypeptide, but is not particularly limited thereto.
보다 구체적으로, 상기 글루카곤 아날로그는 천연형 글루카곤 아미노산 서열에서 1번, 2번, 3번, 7번, 10번, 12번, 13번, 14번, 15번, 16번, 17번, 18번, 19번, 20번, 21번, 23번, 24번, 27번, 28번 및 29번으로 이루어진 군에서 선택된, 1 이상, 2 이상, 3 이상, 4 이상, 5 이상, 6 이상, 7 이상, 8 이상, 9 이상, 10 이상, 11 이상, 12 이상, 13 이상, 14 이상, 15 이상, 16 이상, 17 이상, 18 이상, 19 이상, 또는 20개의 아미노산이 다른 아미노산이 치환된 것일 수 있으며, 또한 독립적으로 또는 추가적으로 이의 C-말단에 1 이상, 2 이상, 3 이상, 4 이상, 5 이상, 6 이상, 7 이상, 8 이상, 9 이상, 10 이상, 또는 11개 이상의 아미노산이 추가된 것일 수 있으나, 특별히 이에 제한되는 것은 아니다. More specifically, the glucagon analog is 1 times, 2 times, 3 times, 7 times, 10 times, 12 times, 13 times, 14 times, 15 times, 16 times, 17 times, 18 times in the natural glucagon amino acid sequence, 1 or more, 2 or more, 3 or more, 4 or more, 5 or more, 6 or more, 7 or more, selected from the group consisting of 19 times, 20 times, 21 times, 23 times, 24 times, 27 times, 28 times and 29 times, 8 or more, 9 or more, 10 or more, 11 or more, 12 or more, 13 or more, 14 or more, 15 or more, 16 or more, 17 or more, 18 or more, 19 or more, or 20 amino acids may be substituted with other amino acids, In addition, independently or additionally, 1 or more, 2 or more, 3 or more, 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, 9 or more, 10 or more, or 11 or more amino acids may be added to the C-terminus thereof. However, it is not particularly limited thereto.
보다 더 구체적으로 상기 글루카곤 아날로그는 천연형 글루카곤 아미노산 서열에서 1번, 2번, 3번, 10번, 12번, 13번, 14번, 15번, 16번, 17번, 18번, 19번, 20번, 21번, 23번, 24번, 27번, 28번 및 29번으로 이루어진 군에서 선택된, 1 이상, 2 이상, 3 이상, 4 이상, 5 이상, 6 이상, 7 이상, 8 이상, 9 이상, 10 이상, 11 이상, 12 이상, 13 이상, 14 이상, 15 이상, 16 이상, 17 이상, 18 이상, 또는 19개의 아미노산이 다른 아미노산이 치환된 것일 수 있으며, 또한 독립적으로 또는 추가적으로 이의 C-말단에 1 이상, 2 이상, 3 이상, 4 이상, 5 이상, 6 이상, 7 이상, 8 이상, 9 이상, 10 이상, 또는 11개 이상의 아미노산이 추가된 것일 수 있으나, 특별히 이에 제한되는 것은 아니다. More specifically, the glucagon analog is 1, 2, 3, 10, 12, 13, 14, 15, 16, 17, 18, 19, in the natural glucagon amino acid sequence, 1 or more, 2 or more, 3 or more, 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, selected from the group consisting of 20, 21, 23, 24, 27, 28 and 29, 9 or more, 10 or more, 11 or more, 12 or more, 13 or more, 14 or more, 15 or more, 16 or more, 17 or more, 18 or more, or 19 amino acids may be substituted with other amino acids, and independently or additionally 1 or more, 2 or more, 3 or more, 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, 9 or more, 10 or more, or 11 or more amino acids may be added to the C-terminus, but is particularly limited thereto It is not.
보다 더 구체적으로 상기 글루카곤 아날로그는 천연형 글루카곤 아미노산 서열에서 1번, 2번, 3번, 10번, 13번, 14번, 15번, 16번, 17번, 18번, 19번, 20번, 21번, 23번, 24번, 28번 및 29번으로 이루어진 군에서 선택된, 1 이상, 2 이상, 3 이상, 4 이상, 5 이상, 6 이상, 7 이상, 8 이상, 9 이상, 10 이상, 11 이상, 12 이상, 13 이상, 14 이상, 15 이상, 16 이상, 또는 17개의 아미노산이 다른 아미노산이 치환된 것일 수 있으며, 또한 독립적으로 또는 추가적으로 이의 C-말단에 1 이상, 2 이상, 3 이상, 4 이상, 5 이상, 6 이상, 7 이상, 8 이상, 9 이상, 10 이상, 또는 11개 이상의 아미노산이 추가된 것일 수 있으나, 특별히 이에 제한되는 것은 아니다. More specifically, the glucagon analog is 1, 2, 3, 10, 13, 14, 15, 16, 17, 18, 19, 20, in the natural glucagon amino acid sequence, 1 or more, 2 or more, 3 or more, 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, 9 or more, 10 or more, selected from the group consisting of 21 times, 23 times, 24 times, 28 times and 29 times, 11 or more, 12 or more, 13 or more, 14 or more, 15 or more, 16 or more, or 17 amino acids may be substituted with other amino acids, and also independently or additionally at the C-terminus of 1 or more, 2 or more, 3 or more , 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, 9 or more, 10 or more, or 11 or more amino acids may be added, but is not particularly limited thereto.
보다 더 구체적으로 상기 글루카곤 아날로그는 천연형 글루카곤 아미노산 서열에서 1번, 2번, 13번, 16번, 17번, 18번, 19번, 20번, 21번, 23번, 24번, 27번, 28번 및 29번으로 이루어진 군에서 선택된, 1 이상, 2 이상, 3 이상, 4 이상, 5 이상, 6 이상, 7 이상, 8 이상, 9 이상, 10 이상, 11 이상, 12이상, 13이상, 또는 14개의 아미노산이 다른 아미노산이 치환된 것일 수 있으며, 또한 독립적으로 또는 추가적으로 이의 C-말단에 1 이상, 2 이상, 3 이상, 4 이상, 5 이상, 6 이상, 7 이상, 8 이상, 9 이상, 10 이상, 또는 11개 이상의 아미노산이 추가된 것일 수 있으나, 특별히 이에 제한되는 것은 아니다. More specifically, the glucagon analog is 1, 2, 13, 16, 17, 18, 19, 20, 21, 23, 24, 27, in the natural glucagon amino acid sequence, 1 or more, 2 or more, 3 or more, 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, 9 or more, 10 or more, 11 or more, 12 or more, 13 or more, selected from the group consisting of 28 and 29, Or 14 amino acids may be substituted with other amino acids, and independently or additionally at the C-terminus of 1 or more, 2 or more, 3 or more, 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, 9 or more , 10 or more, or 11 or more amino acids may be added, but is not particularly limited thereto.
상기 천연형 글루카곤에서 도입되는 아미노산은 티로신, 알파-메틸-글루탐산, Aib, 메티오닌, 글루탐산, 히스티딘, 리신, 류신, 이소류신, 글루타민, 발린, 글리신, 알라닌, 시스테인, 세린, 알라닌, 아스파르트산, 및 아르기닌으로 이루어진 군으로부터 선택될 수 있으나, 특별히 이에 제한되는 것은 아니다. The amino acids introduced from the natural glucagon are tyrosine, alpha-methyl-glutamic acid, Aib, methionine, glutamic acid, histidine, lysine, leucine, isoleucine, glutamine, valine, glycine, alanine, cysteine, serine, alanine, aspartic acid, and arginine. It may be selected from the group consisting of, but is not particularly limited thereto.
예컨대, 상기 추가되는 아미노산 서열은 천연형 GLP-1, 천연형 GIP, 또는 천연형 엑센딘-4 아미노산 서열에서 유래되는 1개 이상의 아미노산 서열일 수 있다. For example, the added amino acid sequence may be one or more amino acid sequences derived from native GLP-1, native GIP, or native exendin-4 amino acid sequence.
이러한 글루카곤 아날로그 또는 삼중 활성체는 분자 내 가교 (intramolecular bridge)를 포함할 수 있으며 (예컨대, 공유결합적 가교 또는 비공유결합적 가교), 구체적으로 고리를 포함하는 형태일 수 있으며, 예컨대 글루카곤 아날로그 또는 삼중 활성체의 16번 및 20번 아미노산 사이에 고리가 형성된 형태일 수 있으나, 특별히 이에 제한되는 것은 아니다. These glucagon analogs or triple activators may include intramolecular bridges (e.g., covalent bridges or non-covalent bridges), and specifically may be in a form including a ring, such as glucagon analogs or triples. A ring may be formed between amino acids 16 and 20 of the active substance, but is not particularly limited thereto.
상기 고리의 비제한적인 예로 락탐 가교 (또는 락탐 고리)를 포함할 수 있다.Non-limiting examples of the ring may include a lactam bridge (or a lactam ring).
또한, 상기 글루카곤 아날로그 또는 삼중 활성체는 고리를 포함하도록, 목적하는 위치에 고리를 형성할 수 있는 아미노산을 포함하도록 변형된 것을 모두 포함한다. In addition, the glucagon analogue or triple activator includes all those modified to include an amino acid capable of forming a ring at a desired position to include a ring.
예컨대, 글루카곤 아날로그 또는 삼중 활성체의 16번 및 20번 아미노산 쌍이 각각 고리를 형성할 수 있는 글루탐산 또는 리신으로 치환된 것일 수 있으나, 이에 제한되지 않는다. For example, the pair of amino acids 16 and 20 of the glucagon analog or triple activator may be substituted with glutamic acid or lysine capable of forming a ring, but is not limited thereto.
이러한 고리는 상기 글루카곤 아날로그 또는 삼중 활성체 내의 아미노산 곁 사슬 간에 형성될 수 있으며, 그 예로 리신의 곁 사슬과 글루탐산의 곁 사슬 간에 락탐 고리가 형성되는 형태일 수 있으나, 특별히 이에 제한되는 것은 아니다. Such a ring may be formed between the side chains of amino acids in the glucagon analog or triple activator, for example, a lactam ring may be formed between the side chain of lysine and the side chain of glutamic acid, but is not particularly limited thereto.
이러한 방법들의 조합으로 제조되는 글루카곤의 아날로그의 예로, 천연형 글루카곤과 아미노산 서열이 하나 이상 다르고, N-말단의 아미노산 잔기의 알파-탄소가 제거된, 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 보유한 펩타이드 등이 있으나, 이에 제한되지 않으며, 아날로그 제조를 위한 여러 방법들의 조합으로 본 발명에 적용되는 천연형 글루카곤의 아날로그를 제조할 수 있다. An example of an analogue of glucagon prepared by a combination of these methods, in which one or more amino acid sequences are different from natural glucagon, and the alpha-carbon of the N-terminal amino acid residue has been removed, glucagon receptor, GLP-1 receptor, and GIP receptor. There are peptides that have activity against, but are not limited thereto, and an analog of natural glucagon applied to the present invention can be prepared by a combination of several methods for analog production.
또한, 특별히 이에 제한되지 않으나, 본 발명의 펩타이드는 체내 반감기의 증가를 위해 활성체 분해 효소의 인식작용을 회피하기 위하여 일부 아미노산을 타 아미노산 혹은 비 천연형 화합물로 치환할 수 있다. In addition, although not particularly limited thereto, in the peptide of the present invention, some amino acids may be substituted with other amino acids or non-natural compounds in order to avoid the recognition of activator decomposing enzymes to increase half-life in the body.
구체적으로, 상기 삼중 활성체의 아미노산 서열 중 두 번째 아미노산 서열의 치환을 통해 분해효소의 인식 작용을 회피하여 체내 반감기를 증가시킨 펩타이드일 수 있으나, 체내 분해 효소의 인식 작용을 회피하기 위한 아미노산 치환 또는 변경은 제한 없이 포함된다.Specifically, it may be a peptide having an increased half-life in the body by avoiding the recognition of a degrading enzyme through substitution of the second amino acid sequence of the amino acid sequence of the triple active agent, but amino acid substitution or Changes are included without limitation.
또한, 천연형 글루카곤의 아날로그의 제조를 위한 이러한 변형은 L-형 혹은 D-형 아미노산, 및/또는 비-천연형 아미노산을 이용한 변형; 및/또는 천연형 서열을 개질, 예를 들어 측쇄 작용기의 변형, 분자 내 공유결합, 예컨대, 측쇄 간 고리 형성, 메틸화, 아실화, 유비퀴틴화, 인산화, 아미노헥산화, 바이오틴화 등과 같이 개질함으로써 변형하는 것을 모두 포함한다. In addition, such modifications for the production of analogs of natural glucagon include modifications using L-type or D-type amino acids, and/or non-natural amino acids; And/or by modifying the native sequence, for example, modification of side chain functional groups, intramolecular covalent bonds, such as inter-side chain ring formation, methylation, acylation, ubiquitination, phosphorylation, aminohexanation, biotinylation, etc. It includes everything you do.
또한, 천연형 글루카곤의 아미노 및/또는 카르복시 말단에 하나 또는 그 이상의 아미노산이 추가된 것을 모두 포함한다. In addition, it includes all those in which one or more amino acids are added to the amino and/or carboxy terminus of the natural glucagon.
상기 치환되거나 추가되는 아미노산은 인간 단백질에서 통상적으로 관찰되는 20개의 아미노산뿐만 아니라 비정형 또는 비-자연적 발생 아미노산을 사용할 수 있다. 비정형 아미노산의 상업적 출처에는 Sigma-Aldrich, ChemPep과 Genzyme pharmaceuticals가 포함된다. 이러한 아미노산이 포함된 펩타이드와 정형적인 펩타이드 서열은 상업화된 펩타이드 합성 회사, 예를 들어 미국의 American peptide company나 Bachem, 또는 한국의 Anygen을 통해 합성 및 구매 가능하다. As the substituted or added amino acids, not only the 20 amino acids commonly observed in human proteins, but also atypical or non-naturally occurring amino acids may be used. Commercial sources of atypical amino acids include Sigma-Aldrich, ChemPep and Genzyme pharmaceuticals. Peptides containing such amino acids and typical peptide sequences can be synthesized and purchased through commercial peptide synthesis companies, such as American peptide company or Bachem in the US, or Anygen in Korea.
아미노산 유도체도 마찬가지 방식으로 입수할 수 있는데, 그 예를 일부만 들자면 4-이미다조아세트산 (4-imidazoacetic acid) 등을 사용할 수 있다. Amino acid derivatives can also be obtained in the same way, and to name only some examples, 4-imidazoacetic acid or the like can be used.
또한, 본 발명에 따른 펩타이드는 생체 내의 단백질 절단 효소들로부터 보호하고 안정성을 증가시키기 위하여 이의 N-말단 및/또는 C-말단 등이 화학적으로 수식되거나 유기단으로 보호되거나, 또는 펩타이드 말단 등에 아미노산이 추가되어 변형된 형태일 수 있다. In addition, the peptide according to the present invention has its N-terminus and/or C-terminus chemically modified or protected with an organic terminal in order to protect from protein cleavage enzymes in vivo and increase stability, or amino acid is added to the peptide terminal. It may be added and modified.
특히, 화학적으로 합성한 펩타이드의 경우, N- 및 C-말단이 전하를 띠고 있기 때문에, 이러한 전하를 제거하기 위하여 N-말단을 아세틸화 (acetylation) 및/또는 C-말단을 아미드화 (amidation)할 수 있으나, 특별히 이에 제한되지는 않는다. In particular, in the case of chemically synthesized peptides, since the N- and C-terminals are charged, the N-terminus is acetylated and/or the C-terminus is amidated to remove such charge. It can be, but is not particularly limited thereto.
또한, 본 발명에 따른 펩타이드는 펩타이드 그 자체, 이의 염 (예컨대, 상기 펩타이드의 약학적으로 허용가능한 염), 또는 이의 용매화물의 형태를 모두 포함한다. 또한, 펩타이드는 약학적으로 허용되는 임의의 형태일 수 있다. In addition, the peptide according to the present invention includes the peptide itself, a salt thereof (eg, a pharmaceutically acceptable salt of the peptide), or a solvate thereof. In addition, the peptide may be in any pharmaceutically acceptable form.
상기 염의 종류는 특별히 제한되지는 않는다. 다만, 개체, 예컨대 포유류에게 안전하고 효과적인 형태인 것이 바람직하나, 특별히 이에 제한되는 것은 아니다. The kind of the salt is not particularly limited. However, it is preferable that it is a safe and effective form for individuals, such as mammals, but is not particularly limited thereto.
상기 용어, "약학적으로 허용되는"은 의약학적 판단의 범위 내에서，과도한 독성，자극, 또는 알레르기 반응 등을 유발하지 않고 원하는 용도에 효과적으로 사용 가능한 물질을 의미한다. The term "pharmaceutically acceptable" refers to a substance that can be effectively used for a desired purpose without causing excessive toxicity, irritation, or allergic reaction within the scope of medical judgment.
본 발명에서 용어, "약학적으로 허용되는 염" 이란 약학적으로 허용되는 무기산, 유기산, 또는 염기로부터 유도된 염을 포함한다. 적합한 산의 예로는 염산, 브롬산, 황산, 질산, 과염소산, 푸마르산，말레산, 인산, 글리콜산, 락트산, 살리실산，숙신산, 톨루엔-p-설폰산, 타르타르산, 아세트산，시트르산, 메탄설폰산，포름산, 벤조산, 말론산, 나프탈렌-2-설폰산，벤젠설폰산 등을 들 수 있다. 적합한 염기로부터 유도된 염은 나트륨, 칼륨 등의 알칼리 금속，마그네슘 등의 알칼리 토금속，및 암모늄 등을 포함할 수 있다.In the present invention, the term "pharmaceutically acceptable salt" includes salts derived from pharmaceutically acceptable inorganic acids, organic acids, or bases. Examples of suitable acids include hydrochloric acid, bromic acid, sulfuric acid, nitric acid, perchloric acid, fumaric acid, maleic acid, phosphoric acid, glycolic acid, lactic acid, salicylic acid, succinic acid, toluene-p-sulfonic acid, tartaric acid, acetic acid, citric acid, methanesulfonic acid, formic acid. , Benzoic acid, malonic acid, naphthalene-2-sulfonic acid, and benzenesulfonic acid. Salts derived from suitable bases may include alkali metals such as sodium and potassium, alkaline earth metals such as magnesium, and ammonium.
또한, 본 발명에서 사용된 용어 "용매화물"은 본 발명에 따른 펩타이드 또는 이의 염이 용매 분자와 복합체를 형성한 것을 말한다. In addition, the term "solvate" as used in the present invention refers to the formation of a complex with a solvent molecule of the peptide or salt thereof according to the present invention.
다른 하나의 구체적인 양태로서, 상기 펩타이드는 하기 일반식 1로 표시된 아미노산 서열을 포함할 수 있다. As another specific embodiment, the peptide may include an amino acid sequence represented by the following general formula 1.
Xaa1-Xaa2-Xaa3-Gly-Thr-Phe-Xaa7-Ser-Asp-Xaa10-Ser-Xaa12-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp-Leu-Xaa27-Xaa28-Xaa29-Xaa30-R1 (일반식 1, 서열번호 103)Xaa1-Xaa2-Xaa3-Gly-Thr-Phe-Xaa7-Ser-Asp-Xaa10-Ser-Xaa12-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp- Leu-Xaa27-Xaa28-Xaa29-Xaa30-R1 (general formula 1, SEQ ID NO: 103)
상기 일반식 1에서, Xaa1은 히스티딘, 4-이미다조아세틸, 또는 티로신이고, Xaa2는 글리신, 알파-메틸-글루탐산, 또는 Aib이며, Xaa3은 글루탐산 또는 글루타민이고, Xaa7은 트레오닌 또는 이소류신이며, Xaa10은 류신, 티로신, 리신, 시스테인, 또는 발린이고, Xaa12는 리신, 세린, 또는 이소류신이며, Xaa13은 글루타민, 티로신, 알라닌, 또는 시스테인이고, Xaa14는 류신, 메티오닌, 또는 티로신이며, Xaa15는 시스테인, 아스파르트산, 글루탐산, 또는 류신이며, Xaa16은 글리신, 글루탐산, 또는 세린이고, Xaa17은 글루타민, 아르기닌, 이소류신, 글루탐산, 시스테인, 또는 리신이며, Xaa18은 알라닌, 글루타민, 아르기닌, 또는 히스티딘이고, Xaa19는 알라닌, 글루타민, 시스테인, 또는 발린이며, Xaa20은 리신, 글루타민, 또는 아르기닌이고, Xaa21은 글루탐산, 글루타민, 류신, 시스테인, 또는 아스파르트산이며, Xaa23은 이소류신 또는 발린이고, Xaa24는 알라닌, 글루타민, 시스테인, 아스파라긴, 아스파르트산, 또는 글루탐산이며, Xaa27은 발린, 류신, 리신, 또는 메티오닌이고, Xaa28은 시스테인, 리신, 알라닌, 아스파라긴, 또는 아스파르트산이며, Xaa29는 시스테인, 글리신, 글루타민, 트레오닌, 글루탐산, 또는 히스티딘이고, Xaa30은 시스테인, 글리신, 리신, 또는 히스티딘이거나, 부존재하며,In the general formula 1, Xaa1 is histidine, 4-imidazoacetyl, or tyrosine, Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib, Xaa3 is glutamic acid or glutamine, Xaa7 is threonine or isoleucine, and Xaa10 is Leucine, tyrosine, lysine, cysteine, or valine, Xaa12 is lysine, serine, or isoleucine, Xaa13 is glutamine, tyrosine, alanine, or cysteine, Xaa14 is leucine, methionine, or tyrosine, and Xaa15 is cysteine, aspartic acid. , Glutamic acid, or leucine, Xaa16 is glycine, glutamic acid, or serine, Xaa17 is glutamine, arginine, isoleucine, glutamic acid, cysteine, or lysine, Xaa18 is alanine, glutamine, arginine, or histidine, Xaa19 is alanine, glutamine , Cysteine, or valine, Xaa20 is lysine, glutamine, or arginine, Xaa21 is glutamic acid, glutamine, leucine, cysteine, or aspartic acid, Xaa23 is isoleucine or valine, Xaa24 is alanine, glutamine, cysteine, asparagine, aspartic acid. Acid, or glutamic acid, Xaa27 is valine, leucine, lysine, or methionine, Xaa28 is cysteine, lysine, alanine, asparagine, or aspartic acid, Xaa29 is cysteine, glycine, glutamine, threonine, glutamic acid, or histidine, Xaa30 Is cysteine, glycine, lysine, or histidine, or is absent,
R1은 시스테인, GKKNDWKHNIT (서열번호 106), m-SSGAPPPS-n (서열번호 107), 또는 m-SSGQPPPS-n (서열번호 108)이거나, 부존재하며, R1 is cysteine, GKKNDWKHNIT (SEQ ID NO: 106), m-SSGAPPPS-n (SEQ ID NO: 107), or m-SSGQPPPS-n (SEQ ID NO: 108), or absent,
여기서, m은 -Cys-, -Pro-, 또는 -Gly-Pro-이고, n은 -Cys-, -Gly-, -Ser-, 또는 -His-Gly-이거나, 부존재할 수 있다.Here, m is -Cys-, -Pro-, or -Gly-Pro-, and n is -Cys-, -Gly-, -Ser-, or -His-Gly-, or may be absent.
상기 삼중 활성체의 예로, 서열번호: 1 내지 102로 이루어진 군에서 선택된 아미노산 서열을 포함하는 것, 서열번호: 1 내지 102로 이루어진 군에서 선택된 아미노산 서열로 (필수적으로) 구성된 것일 수 있으나, 이에 제한되는 것은 아니다. Examples of the triple activator include those comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 1 to 102, and those consisting of (essentially) an amino acid sequence selected from the group consisting of SEQ ID NOs: 1 to 102, but are limited thereto. It does not become.
또한, 본원에서 '특정 서열번호로 구성되는 펩타이드'라고 기재되어 있다 하더라도, 해당 서열번호의 아미노산 서열로 이루어진 펩타이드와 동일 혹은 상응하는 활성을 가지는 경우라면 해당 서열번호의 아미노산 서열 앞뒤의 무의미한 서열 추가 또는 자연적으로 발생할 수 있는 돌연변이, 혹은 이의 잠재성 돌연변이 (silent mutation)를 제외하는 것이 아니며, 이러한 서열 추가 혹은 돌연변이를 가지는 경우에도 본원의 범위 내에 속하는 것이 자명하다.In addition, even if it is described herein as'a peptide consisting of a specific sequence number', if it has the same or corresponding activity as the peptide consisting of the amino acid sequence of the corresponding sequence number, an insignificant sequence is added before and after the amino acid sequence of the corresponding sequence number or It does not exclude mutations that may occur naturally, or silent mutations thereof, and it is obvious that even when such sequence additions or mutations are present, they fall within the scope of the present application.
이상의 내용은 본 발명의 다른 구체예 혹은 다른 양태에도 적용될 수 있으나, 이에 제한되는 것은 아니다. The above contents may be applied to other embodiments or other aspects of the present invention, but is not limited thereto.
구체적으로, 상기 일반식 1에서 Xaa14는 류신 또는 메티오닌이며, Xaa15는 시스테인, 아스파르트산, 또는 류신일 수 있다. Specifically, in General Formula 1, Xaa14 may be leucine or methionine, and Xaa15 may be cysteine, aspartic acid, or leucine.
이러한 펩타이드의 예로, 서열번호: 1 내지 12, 14 내지 17, 및 21 내지 102로 이루어진 군에서 선택된 아미노산 서열을 포함하거나, 이로 (필수적으로) 구성된 펩타이드를 들 수 있으나, 특별히 이에 제한되는 것은 아니다. Examples of such a peptide include, but are not limited to, a peptide comprising or (essentially) an amino acid sequence selected from the group consisting of SEQ ID NOs: 1 to 12, 14 to 17, and 21 to 102.
이러한 펩타이드는 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체 중 하나 이상을 유의하게 활성화시킬 수 있으나, 특별히 이에 제한되는 것은 아니다. 구체적으로, GLP-1을 유의하게 활성화시키거나, 추가로 글루카곤 수용체 및/또는 GIP 수용체를 유의하게 활성화시키는 것일 수 있으나, 특별히 이에 제한되지 않는다. These peptides may significantly activate one or more of the glucagon receptor, the GLP-1 receptor, and the GIP receptor, but are not particularly limited thereto. Specifically, it may significantly activate GLP-1, or significantly activate the glucagon receptor and/or the GIP receptor, but is not particularly limited thereto.
보다 더 구체적으로, 상기 일반식 1에서, Xaa2는 글리신, 알파-메틸-글루탐산, 또는 Aib이고, Xaa7은 트레오닌이며, Xaa10은 티로신, 시스테인, 또는 발린이고, Xaa12는 리신 또는 이소류신이며, Xaa13은 티로신, 알라닌, 글루타민, 또는 시스테인이고, Xaa14는 류신, 시스테인, 또는 메티오닌이며, Xaa15는 시스테인, 류신, 글루탐산, 또는 아스파르트산이고, Xaa17은 글루타민, 아르기닌, 이소류신, 시스테인, 글루탐산, 또는 리신이며, Xaa18은 알라닌, 글루타민, 아르기닌, 또는 히스티딘이고, Xaa19는 알라닌, 글루타민, 발린, 또는 시스테인이며, Xaa20은 리신, 아르기닌, 또는 글루타민이고, Xaa21은 글루탐산, 글루타민, 류신, 시스테인, 또는 아스파르트산이며, Xaa23은 이소류신 또는 발린이고, Xaa24는 시스테인, 알라닌, 글루타민, 아스파라긴, 글루탐산, 또는 아스파르트산이며, Xaa27은 류신 또는 리신인, 펩타이드일 수 있으나, 특별히 이에 제한되는 것은 아니다. More specifically, in the general formula 1, Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib, Xaa7 is threonine, Xaa10 is tyrosine, cysteine, or valine, Xaa12 is lysine or isoleucine, and Xaa13 is tyrosine. , Alanine, glutamine, or cysteine, Xaa14 is leucine, cysteine, or methionine, Xaa15 is cysteine, leucine, glutamic acid, or aspartic acid, Xaa17 is glutamine, arginine, isoleucine, cysteine, glutamic acid, or lysine, and Xaa18 is Alanine, glutamine, arginine, or histidine, Xaa19 is alanine, glutamine, valine, or cysteine, Xaa20 is lysine, arginine, or glutamine, Xaa21 is glutamic acid, glutamine, leucine, cysteine, or aspartic acid, and Xaa23 is isoleucine. Or valine, Xaa24 is cysteine, alanine, glutamine, asparagine, glutamic acid, or aspartic acid, and Xaa27 may be leucine or lysine, or a peptide, but is not particularly limited thereto.
보다 더 구체적으로, 상기 일반식 1에서, Xaa2는 글리신, 알파-메틸-글루탐산, 또는 Aib이고, Xaa7은 트레오닌이며, Xaa10은 티로신, 시스테인, 또는 발린이고, Xaa12는 리신 또는 이소류신이며, Xaa13은 티로신, 알라닌, 또는 시스테인이고, Xaa14는 류신 또는 메티오닌이며, Xaa15는 시스테인 또는 아스파르트산이고, Xaa17은 글루타민, 아르기닌, 이소류신, 시스테인, 또는 리신이며, Xaa18은 알라닌, 아르기닌, 또는 히스티딘이고, Xaa19는 알라닌, 글루타민, 또는 시스테인이며, Xaa20은 리신 또는 글루타민이고, Xaa21은 글루탐산, 시스테인, 또는 아스파르트산이며, Xaa23은 발린이고, Xaa24는 알라닌, 글루타민, 시스테인, 아스파라긴, 또는 아스파르트산이며, Xaa27은 류신 또는 리신일 수 있으나, 특별히 이에 제한되지는 않는다. More specifically, in the general formula 1, Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib, Xaa7 is threonine, Xaa10 is tyrosine, cysteine, or valine, Xaa12 is lysine or isoleucine, and Xaa13 is tyrosine. , Alanine, or cysteine, Xaa14 is leucine or methionine, Xaa15 is cysteine or aspartic acid, Xaa17 is glutamine, arginine, isoleucine, cysteine, or lysine, Xaa18 is alanine, arginine, or histidine, Xaa19 is alanine, Glutamine or cysteine, Xaa20 is lysine or glutamine, Xaa21 is glutamic acid, cysteine, or aspartic acid, Xaa23 is valine, Xaa24 is alanine, glutamine, cysteine, asparagine, or aspartic acid, and Xaa27 is leucine or lysine. However, it is not particularly limited thereto.
보다 더 구체적으로, 상기 일반식 1에서, Xaa2는 알파-메틸-글루탐산 또는 Aib이고, Xaa7은 트레오닌이며, Xaa10은 티로신 또는 시스테인이고, Xaa12는 리신 또는 이소류신이며, Xaa13은 티로신, 알라닌, 또는 시스테인이고, Xaa14는 류신 또는 메티오닌이며, Xaa15는 시스테인 또는 아스파르트산이고, Xaa16은 글루탐산이며, Xaa17은 아르기닌, 이소류신, 시스테인, 또는 리신이고, Xaa18은 알라닌, 아르기닌, 또는 히스티딘이며, Xaa19는 알라닌, 글루타민, 또는 시스테인이고, Xaa20은 리신 또는 글루타민이며, Xaa21은 글루탐산 또는 아스파르트산이고, Xaa23은 발린이며, Xaa24는 글루타민, 아스파라긴, 또는 아스파르트산이고, Xaa27은 류신이며, Xaa28은 시스테인, 알라닌, 아스파라긴, 또는 아스파르트산일 수 있다. More specifically, in the general formula 1, Xaa2 is alpha-methyl-glutamic acid or Aib, Xaa7 is threonine, Xaa10 is tyrosine or cysteine, Xaa12 is lysine or isoleucine, and Xaa13 is tyrosine, alanine, or cysteine. , Xaa14 is leucine or methionine, Xaa15 is cysteine or aspartic acid, Xaa16 is glutamic acid, Xaa17 is arginine, isoleucine, cysteine, or lysine, Xaa18 is alanine, arginine, or histidine, and Xaa19 is alanine, glutamine, or Cysteine, Xaa20 is lysine or glutamine, Xaa21 is glutamic acid or aspartic acid, Xaa23 is valine, Xaa24 is glutamine, asparagine, or aspartic acid, Xaa27 is leucine, Xaa28 is cysteine, alanine, asparagine, or aspartic acid. I can.
구체적으로, 상기 일반식 1에서, Xaa1은 히스티딘 또는 4-이미다조아세틸이고, Xaa2는 알파-메틸-글루탐산 또는 Aib이며, Xaa3은 글루타민이고, Xaa7은 트레오닌이며, Xaa10은 티로신이고, Xaa12는 이소류신이며, Xaa13은 알라닌 또는 시스테인이고, Xaa14는 메티오닌이며, Xaa15는 아스파르트산이고, Xaa16은 글루탐산이며, Xaa17은 이소류신 또는 리신이고, Xaa18은 알라닌 또는 히스티딘이며, Xaa19는 글루타민 또는 시스테인이고, Xaa20은 리신이며, Xaa21은 아스파르트산이고, Xaa23은 발린이며, Xaa24는 아스파라긴이고, Xaa27은 류신이며, Xaa28은 알라닌 또는 아스파라긴이고, Xaa29는 글루타민 또는 트레오닌이며, Xaa30은 시스테인 또는 리신이거나, 부존재할 수 있다. Specifically, in the general formula 1, Xaa1 is histidine or 4-imidazoacetyl, Xaa2 is alpha-methyl-glutamic acid or Aib, Xaa3 is glutamine, Xaa7 is threonine, Xaa10 is tyrosine, and Xaa12 is isoleucine. , Xaa13 is alanine or cysteine, Xaa14 is methionine, Xaa15 is aspartic acid, Xaa16 is glutamic acid, Xaa17 is isoleucine or lysine, Xaa18 is alanine or histidine, Xaa19 is glutamine or cysteine, Xaa20 is lysine, Xaa21 is aspartic acid, Xaa23 is valine, Xaa24 is asparagine, Xaa27 is leucine, Xaa28 is alanine or asparagine, Xaa29 is glutamine or threonine, and Xaa30 is cysteine or lysine, or may be absent.
보다 구체적으로, 상기 일반식 1에서, Xaa2는 글리신, 알파-메틸-글루탐산, 또는 Aib이며, Xaa3은 글루타민이고, Xaa7은 트레오닌이며, Xaa10은 티로신, 시스테인, 또는 발린이고, Xaa12는 리신이며, Xaa13은 티로신이고, Xaa14는 류신이며, Xaa15는 아스파르트산이며, Xaa16은 글리신, 글루탐산, 또는 세린이고, Xaa17은 글루타민, 아르기닌, 시스테인, 또는 리신이며, Xaa18은 알라닌, 아르기닌, 또는 히스티딘이고, Xaa19는 알라닌 또는 글루타민이며, Xaa20은 리신 또는 글루타민이고, Xaa21은 글루탐산, 시스테인, 또는 아스파르트산이며, Xaa23은 발린이고, Xaa24는 알라닌, 글루타민, 또는 시스테인이며, Xaa27은 류신 또는 리신이고, Xaa29는 글리신, 글루타민, 트레오닌, 또는 히스티딘일 수 있으나, 특별히 이에 제한되는 것은 아니다. More specifically, in the general formula 1, Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib, Xaa3 is glutamine, Xaa7 is threonine, Xaa10 is tyrosine, cysteine, or valine, Xaa12 is lysine, Xaa13 Is tyrosine, Xaa14 is leucine, Xaa15 is aspartic acid, Xaa16 is glycine, glutamic acid, or serine, Xaa17 is glutamine, arginine, cysteine, or lysine, Xaa18 is alanine, arginine, or histidine, and Xaa19 is alanine. Or glutamine, Xaa20 is lysine or glutamine, Xaa21 is glutamic acid, cysteine, or aspartic acid, Xaa23 is valine, Xaa24 is alanine, glutamine, or cysteine, Xaa27 is leucine or lysine, Xaa29 is glycine, glutamine, It may be threonine or histidine, but is not particularly limited thereto.
이러한 펩타이드는 GLP-1 수용체 및 글루카곤 수용체의 활성화 정도가 유의하고, GIP 수용체의 활성화 정도에 비해 높거나; GLP-1 수용체, 글루카곤 수용체 및 GIP 수용체의 활성화 정도가 모두 유의하거나; GLP-1 수용체 및 GIP 수용체의 활성화 정도가 유의하고, 글루카곤 수용체의 활성화에 비해 높은 경우에 해당할 수 있으나, 특별히 이에 제한되지 않는다. These peptides have a significant degree of activation of the GLP-1 receptor and the glucagon receptor, and are higher than that of the GIP receptor; The degree of activation of the GLP-1 receptor, the glucagon receptor, and the GIP receptor are all significant; The degree of activation of the GLP-1 receptor and the GIP receptor is significant, and may correspond to a case that is higher than that of the glucagon receptor, but is not particularly limited thereto.
이러한 펩타이드의 예로, 서열번호: 8, 9, 21 내지 37, 39, 42, 43, 49 내지 61, 64 내지 83, 85, 86, 88, 89, 91 내지 93, 95 내지 102로 이루어진 군에서 선택된 아미노산 서열을 포함하거나, 이로 (필수적으로) 구성된 펩타이드를 들 수 있으나, 특별히 이에 제한되는 것은 아니다. Examples of such peptides, SEQ ID NO: 8, 9, 21 to 37, 39, 42, 43, 49 to 61, 64 to 83, 85, 86, 88, 89, 91 to 93, selected from the group consisting of 95 to 102 Peptides including or (essentially) composed of an amino acid sequence may be mentioned, but are not particularly limited thereto.
구체적인 양태로서, 상기 펩타이드는 하기 일반식 2로 표시되는 아미노산 서열을 포함하는 것일 수 있다. In a specific embodiment, the peptide may include an amino acid sequence represented by the following general formula 2.
Xaa1-Xaa2-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Xaa10-Ser-Lys-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp-Leu-Leu-Xaa28-Xaa29-Xaa30-Xaa31-Ser-Ser-Gly-Gln-Pro-Pro-Pro-Ser-Xaa40 (일반식 2, 서열번호 104)Xaa1-Xaa2-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Xaa10-Ser-Lys-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp- Leu-Leu-Xaa28-Xaa29-Xaa30-Xaa31-Ser-Ser-Gly-Gln-Pro-Pro-Pro-Ser-Xaa40 (general formula 2, SEQ ID NO: 104)
상기 식에서, Xaa1은 4-이미다조아세틸, 히스티딘, 또는 티로신이고; Xaa2는 글리신, 알파-메틸-글루탐산, 또는 Aib이며; Xaa10은 티로신, 또는 시스테인이며; Xaa13은 알라닌, 글루타민, 티로신, 또는 시스테인이며; Xaa14는 류신, 메티오닌, 또는 티로신이고; Xaa15는 아스파르트산, 글루탐산, 또는 류신이며; Xaa16은 글리신, 글루탐산, 또는 세린이고; Xaa17은 글루타민, 아르기닌, 이소류신, 글루탐산, 시스테인, 또는 리신이며; Xaa18은 알라닌, 글루타민, 아르기닌, 또는 히스티딘이고; Xaa19는 알라닌, 글루타민, 시스테인, 또는 발린이며; Xaa20은 리신, 글루타민, 또는 아르기닌이고; Xaa21은 시스테인, 글루탐산, 글루타민, 류신, 또는 아스파르트산이며; Xaa23은 이소류신 또는 발린이고; Xaa24는 시스테인, 알라닌, 글루타민, 아스파라긴, 또는 글루탐산이며; Xaa28은 리신, 시스테인, 아스파라긴, 또는 아스파르트산이며; Xaa29는 글리신, 글루타민, 시스테인, 또는 히스티딘이고; Xaa30은 시스테인, 글리신, 리신, 또는 히스티딘이며; Xaa31은 프롤린 또는 시스테인이며; Xaa40은 시스테인이거나, 부존재할 수 있다.Wherein Xaa1 is 4-imidazoacetyl, histidine, or tyrosine; Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib; Xaa10 is tyrosine or cysteine; Xaa13 is alanine, glutamine, tyrosine, or cysteine; Xaa14 is leucine, methionine, or tyrosine; Xaa15 is aspartic acid, glutamic acid, or leucine; Xaa16 is glycine, glutamic acid, or serine; Xaa17 is glutamine, arginine, isoleucine, glutamic acid, cysteine, or lysine; Xaa18 is alanine, glutamine, arginine, or histidine; Xaa19 is alanine, glutamine, cysteine, or valine; Xaa20 is lysine, glutamine, or arginine; Xaa21 is cysteine, glutamic acid, glutamine, leucine, or aspartic acid; Xaa23 is isoleucine or valine; Xaa24 is cysteine, alanine, glutamine, asparagine, or glutamic acid; Xaa28 is lysine, cysteine, asparagine, or aspartic acid; Xaa29 is glycine, glutamine, cysteine, or histidine; Xaa30 is cysteine, glycine, lysine, or histidine; Xaa31 is proline or cysteine; Xaa40 may be cysteine or absent.
보다 구체적으로, 상기 일반식 2에서, Xaa13은 알라닌, 티로신, 또는 시스테인이며; Xaa15는 아스파르트산 또는 글루탐산이고; Xaa17은 글루타민, 아르기닌, 시스테인, 또는 리신이며; Xaa18은 알라닌, 아르기닌, 또는 히스티딘이고; Xaa21은 시스테인, 글루탐산, 글루타민, 또는 아스파르트산이며; Xaa23은 이소류신 또는 발린이고; Xaa24는 시스테인, 글루타민, 또는 아스파라긴이고; Xaa28은 시스테인, 아스파라긴, 또는 아스파르트산이며; Xaa29는 글루타민, 시스테인, 또는 히스티딘이고; Xaa30은 시스테인, 리신, 또는 히스티딘일 수 있다. More specifically, in Formula 2, Xaa13 is alanine, tyrosine, or cysteine; Xaa15 is aspartic acid or glutamic acid; Xaa17 is glutamine, arginine, cysteine, or lysine; Xaa18 is alanine, Arginine, or histidine; Xaa21 is cysteine, glutamic acid, glutamine, or aspartic acid; Xaa23 is isoleucine or valine; Xaa24 is cysteine, glutamine, or asparagine; Xaa28 is cysteine, asparagine, or aspartic acid; Xaa29 is glutamine, cysteine, or histidine; Xaa30 can be cysteine, lysine, or histidine.
이러한 펩타이드의 예로, 서열번호: 21, 22, 42, 43, 50, 64 내지 77, 및 95 내지 102로 이루어진 군에서 선택된 아미노산 서열, 보다 구체적으로 서열번호: 21, 22, 42, 43, 50, 64 내지 77, 및 96 내지 102로 이루어진 군에서 선택된 아미노산 서열을 포함하거나, 이로 (필수적으로) 구성된 펩타이드를 들 수 있으나, 특별히 이에 제한되는 것은 아니다. Examples of such peptides, SEQ ID NOs: 21, 22, 42, 43, 50, 64 to 77, and an amino acid sequence selected from the group consisting of 95 to 102, more specifically SEQ ID NO: 21, 22, 42, 43, 50, A peptide comprising or (essentially) composed of an amino acid sequence selected from the group consisting of 64 to 77, and 96 to 102 may be mentioned, but is not particularly limited thereto.
구체적인 양태로서, 상기 펩타이드는 하기 일반식 3의 아미노산 서열을 포함할 수 있다. As a specific embodiment, the peptide may include the amino acid sequence of the following general formula 3.
Xaa1-Xaa2-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-Lys-Xaa13-Leu-Asp-Glu-Xaa17-Xaa18-Xaa19-Lys-Xaa21-Phe-Val-Xaa24-Trp-Leu-Leu-Xaa28-Xaa29-Xaa30-Xaa31-Ser-Ser-Gly-Gln-Pro-Pro-Pro-Ser-Xaa40 (일반식 3, 서열번호 105),Xaa1-Xaa2-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-Lys-Xaa13-Leu-Asp-Glu-Xaa17-Xaa18-Xaa19-Lys-Xaa21-Phe-Val-Xaa24-Trp- Leu-Leu-Xaa28-Xaa29-Xaa30-Xaa31-Ser-Ser-Gly-Gln-Pro-Pro-Pro-Ser-Xaa40 (general formula 3, SEQ ID NO: 105),
상기 일반식 3에서, Xaa1은 히스티딘 또는 티로신이고; Xaa2는 알파-메틸-글루탐산 또는 Aib이며; Xaa13은 알라닌, 티로신 또는 시스테인이고; Xaa17은 아르기닌, 시스테인, 또는 리신이며; Xaa18은 알라닌 또는 아르기닌이고; Xaa19는 알라닌 또는 시스테인이며; Xaa21은 글루탐산 또는 아스파르트산이고; Xaa24는 글루타민 또는 아스파라긴이며; Xaa28은 시스테인 또는 아스파르트산이며; Xaa29는 시스테인, 히스티딘, 또는 글루타민이고; Xaa30은 시스테인 또는 히스티딘이며; Xaa31은 프롤린 또는 시스테인이며; Xaa40은 시스테인 또는 부존재할 수 있다. In Formula 3, Xaa1 is histidine or tyrosine; Xaa2 is alpha-methyl-glutamic acid or Aib; Xaa13 is alanine, tyrosine or cysteine; Xaa17 is arginine, cysteine, or lysine; Xaa18 is alanine or arginine; Xaa19 is alanine or cysteine; Xaa21 is glutamic acid or aspartic acid; Xaa24 is glutamine or asparagine; Xaa28 is cysteine or aspartic acid; Xaa29 is cysteine, histidine, or glutamine; Xaa30 is cysteine or histidine; Xaa31 is proline or cysteine; Xaa40 may be cysteine or absent.
이러한 펩타이드의 예로, 서열번호: 21, 22, 42, 43, 50, 64 내지 71, 75 내지 77, 및 96 내지 102로 이루어진 군에서 선택된 아미노산 서열을 포함하거나, 이로 (필수적으로) 구성된 펩타이드를 들 수 있으나, 특별히 이에 제한되는 것은 아니다. Examples of such peptides include, or (essentially) a peptide consisting of an amino acid sequence selected from the group consisting of SEQ ID NOs: 21, 22, 42, 43, 50, 64 to 71, 75 to 77, and 96 to 102. However, it is not particularly limited thereto.
또한, 상기 일반식 1에서 R1은 시스테인, GKKNDWKHNIT(서열번호 106), CSSGQPPPS (서열번호 109), GPSSGAPPPS (서열번호 110), GPSSGAPPPSC (서열번호 111), PSSGAPPPS (서열번호 112), PSSGAPPPSG (서열번호 113), PSSGAPPPSHG (서열번호 114), PSSGAPPPSS (서열번호 115), PSSGQPPPS (서열번호 116), 또는 PSSGQPPPSC (서열번호 117)이거나, 부존재할 수 있으나, 특별히 이에 제한되는 것은 아니다. In addition, in the general formula 1 R1 is cysteine, GKKNDWKHNIT (SEQ ID NO: 106), CSSGQPPPS (SEQ ID NO: 109), GPSSGAPPPS (SEQ ID NO: 110), GPSSGAPPPSC (SEQ ID NO: 111), PSSGAPPPS (SEQ ID NO: 112), PSSGAPPPSG (SEQ ID NO: 113), PSSGAPPPSHG (SEQ ID NO: 114), PSSGAPPPSS (SEQ ID NO: 115), PSSGQPPPS (SEQ ID NO: 116), or PSSGQPPPSC (SEQ ID NO: 117), or may be absent, but is not particularly limited thereto.
또한, 본 발명의 펩타이드는 그 길이에 따라 이 분야에서 잘 알려진 방법, 예를 들어 자동 펩타이드 합성기에 의해 합성할 수 있으며, 유전자 조작 기술에 의하여 생산할 수도 있다. In addition, the peptide of the present invention can be synthesized by a method well known in the art, for example, an automatic peptide synthesizer, depending on its length, or can be produced by genetic engineering techniques.
구체적으로, 본 발명의 펩타이드는 표준 합성 방법, 재조합 발현 시스템, 또는 임의의 다른 당해 분야의 방법에 의해 제조될 수 있다. 따라서, 본 발명에 따른 펩타이드는, 예를 들어 하기를 포함하는 방법을 포함하는 다수의 방법으로 합성될 수 있다:Specifically, the peptides of the present invention can be prepared by standard synthetic methods, recombinant expression systems, or any other art method. Thus, the peptides according to the invention can be synthesized by a number of methods, including, for example, methods including:
(a) 펩타이드를 고체상 또는 액체상 방법의 수단으로 단계적으로 또는 단편 조립에 의해 합성하고, 최종 펩타이드 생성물을 분리 및 정제하는 방법; 또는(a) a method of synthesizing a peptide stepwise or by fragment assembly by means of a solid or liquid phase method, and separating and purifying the final peptide product; or
(b) 펩타이드를 인코딩하는 핵산 작제물을 숙주세포 내에서 발현시키고, 발현 생성물을 숙주 세포 배양물로부터 회수하는 방법; 또는(b) a method of expressing a nucleic acid construct encoding a peptide in a host cell, and recovering the expression product from the host cell culture; or
(c) 펩타이드를 인코딩하는 핵산 작제물의 무세포 시험관 내 발현을 수행하고, 발현 생성물을 회수하는 방법; 또는 (c) a method of performing cell-free in vitro expression of a nucleic acid construct encoding a peptide and recovering the expression product; or
(a), (b) 및 (c)의 임의의 조합으로 펩타이드의 단편을 수득하고, 이어서 단편을 연결시켜 펩타이드를 수득하고, 당해 펩타이드를 회수하는 방법.A method of obtaining a fragment of a peptide by any combination of (a), (b) and (c), and then ligating the fragments to obtain a peptide, and recovering the peptide.
또한, 상기 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 펩타이트는, 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 펩타이드에 이의 생체 내 반감기를 증가시키기 위한 생체적합성 물질이 결합된, 결합체 형태일 수 있다. 본 명세서에서 상기 생체적합성 물질은 캐리어와 혼용될 수 있다. In addition, the glucagon receptor, the GLP-1 receptor, and the peptide having an activity against the GIP receptor, the glucagon receptor, the GLP-1 receptor, and the peptide having an activity against the GIP receptor, in vivo for increasing its half-life in vivo. It may be in the form of a combination, in which a suitable substance is combined. In the present specification, the biocompatible material may be mixed with a carrier.
본 발명에서 상기 펩타이드의 결합체는 캐리어가 결합되지 않은 상기 펩타이드에 비하여 증가된 효력의 지속성을 나타낼 수 있으며, 본 발명에서는 이러한 결합체를 "지속형 결합체"로 지칭한다. In the present invention, the conjugate of the peptide may exhibit increased persistence of potency compared to the peptide to which no carrier is bound, and in the present invention, the conjugate is referred to as a "persistent conjugate".
한편, 이러한 결합체는 비자연적인 (non-naturally occurring) 것일 수 있다. On the other hand, such a conjugate may be non-naturally occurring.
본 발명을 구현하기 위한 다른 하나의 양태는 글루카곤 수용체, GLP-1 (Glucagon-like peptide-1) 수용체, 및 GIP (Glucose-dependent insuliontropic polypeptide) 수용체에 대해 활성을 갖는 펩타이드를 포함하는 결합체를 포함하는, 고지혈증의 예방 또는 치료용 약학적 조성물이다. Another aspect for implementing the present invention comprises a conjugate comprising a peptide having activity against a glucagon receptor, a GLP-1 (Glucagon-like peptide-1) receptor, and a GIP (Glucose-dependent insuliontropic polypeptide) receptor. , It is a pharmaceutical composition for the prevention or treatment of hyperlipidemia.
본 발명의 하나의 구체예에서, 상기 결합체는 하기 화학식 1로 표시되는, 결합체이다:In one embodiment of the present invention, the conjugate is a conjugate represented by the following formula (1):
[화학식 1][Formula 1]
X - La - FX-La-F
여기서, X는, 상기 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는, 펩타이드이고; L은, 폴리에틸렌 글리콜이며; a는, 0 또는 자연수이며, 다만 a가 2 이상일 때, 각각의 L은 서로 독립적이고; F는, 면역글로불린 Fc 영역이다.Here, X is a peptide having activity against the glucagon receptor, GLP-1 receptor, and GIP receptor; L is polyethylene glycol; a is 0 or a natural number, provided that when a is 2 or more, each L is independent of each other; F is an immunoglobulin Fc region.
상기 결합체에서 X, 즉 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는 펩타이드에 대해서는 앞서 설명한 바와 같다.Peptides having activity against X in the conjugate, that is, glucagon receptor, GLP-1 receptor, and GIP receptor are as described above.
상기 결합체에서 F는 X, 즉 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는, 펩타이드의 반감기를 증가시킬 수 있는 물질로서, 본 발명의 상기 결합체를 구성하는 모이어티의 일 구성에 해당한다. In the conjugate, F is a substance capable of increasing the half-life of the peptide, having activity against X, that is, the glucagon receptor, the GLP-1 receptor, and the GIP receptor, and is a component of the moiety constituting the conjugate of the present invention. It corresponds.
상기 F는 X와 공유 화학결합 또는 비공유 화학결합으로 서로 결합되는 것일 수 있으며, 공유 화학결합, 비공유 화학결합 또는 이들의 조합으로 L을 통하여 F와 X가 서로 결합되는 것일 수 있다.The F may be bonded to each other by a covalent chemical bond or a non-covalent chemical bond with X, and F and X may be bonded to each other through L through a covalent chemical bond, a non-covalent chemical bond, or a combination thereof.
상기면역글로불린 Fc 영역은 구체적으로 IgG Fc 영역일 수 있으나, 특별히 이에 제한되지 않는다. The immunoglobulin Fc region may specifically be an IgG Fc region, but is not particularly limited thereto.
본 발명의 펩타이드 내의 하나 이상의 아미노산 측쇄는 생체 내에서 가용성 및/또는 반감기를 증가시키고/시키거나 생체이용율을 증가시키기 위해 이러한 생체적합성 물질에 접합될 수 있다. 이러한 변형은 또한 치료학적 단백질 및 펩타이드의 소거 (clearance)를 감소시킬 수 있다. One or more amino acid side chains in the peptides of the invention may be conjugated to such biocompatible materials to increase solubility and/or half-life and/or increase bioavailability in vivo. Such modifications can also reduce the clearance of therapeutic proteins and peptides.
상술한 생체적합성 물질은 수용성 (양친매성 또는 친수성) 및/또는 무독성 및/또는 약학적으로 허용가능한 것일 수 있다. The biocompatible material described above may be water-soluble (amphiphilic or hydrophilic) and/or non-toxic and/or pharmaceutically acceptable.
상기 F는 X와 직접적으로 연결되거나 (즉, 상기 화학식 1에서 a가 0이거나), 또는 링커 (L)를 통해 연결된 것일 수 있다. The F may be directly linked to X (ie, a is 0 in Formula 1), or linked through a linker (L).
구체적으로, 상기 L은 비펩타이드성 링커인 폴리에틸렌 글리콜일 수 있다. Specifically, L may be a non-peptidyl linker polyethylene glycol.
상기 La에서 a는 1 이상일 수 있으며, a가 2 이상일 때 각각의 L은 독립적일 수 있다. In La, a may be 1 or more, and when a is 2 or more, each L may be independent.
또한, 하나의 구체적인 실시 형태에서 상기 결합체는 양쪽 말단에 F, 구체적으로 면역글로불린 Fc 영역 및 X, 구체적으로 펩타이드 약물과 결합될 수 있는 반응기를 포함하는 비펩타이드성 링커를 통하여 F와 X가 서로 공유결합적으로 연결된 것일 수 있다. In addition, in one specific embodiment, the conjugate is shared with each other through a non-peptidyl linker including F at both ends, specifically an immunoglobulin Fc region and X, specifically a reactive group capable of binding to a peptide drug. It may be connected in combination.
상기 폴리에틸렌글리콜 뿐만 아니라, 당해 분야에 이미 알려진 이들의 유도체 및 당해 분야의 기술 수준에서 용이하게 제조할 수 있는 유도체들도 본 발명의 범위에 포함된다.In addition to the polyethylene glycol, derivatives thereof already known in the art and derivatives that can be easily prepared at the technical level in the art are included in the scope of the present invention.
본 발명에서 사용될 수 있는 비펩타이드성 링커인 폴리에틸렌 글리콜의 분자량은 1 내지 100 kDa 범위, 구체적으로 1 내지 20 kDa 범위이나, 이에 제한되지 않는다. 또한, 상기 F에 해당하는 폴리펩타이드와 결합되는 본 발명의 비펩타이드성 링커는 한 종류의 중합체뿐만 아니라 상이한 종류의 중합체들의 조합이 사용될 수도 있다. The molecular weight of polyethylene glycol, which is a non-peptidyl linker that can be used in the present invention, is in the range of 1 to 100 kDa, specifically 1 to 20 kDa, but is not limited thereto. In addition, the non-peptidyl linker of the present invention bonded to the polypeptide corresponding to F may be a combination of different types of polymers as well as one type of polymer.
하나의 구체적인 실시 형태에서 상기 비펩타이드성 링커의 양 말단은 각각 F, 예컨대 면역글로불린 Fc 영역의 아민기 또는 티올기 및 X의 아민기 또는 티올기에 결합할 수 있다.In one specific embodiment, both ends of the non-peptidyl linker may be bonded to F, such as an amine group or thiol group of an immunoglobulin Fc region, and an amine group or thiol group of X.
구체적으로, 상기 비펩타이드성 중합체는 양쪽 말단에 각각 F (예컨대, 면역글로불린 Fc 영역) 및 X와 결합될 수 있는 반응기, 구체적으로는 X, 혹은 F (예컨대, 면역글로불린 Fc 영역)의 N-말단 또는 리신에 위치한 아민기, 또는 시스테인의 티올기와 결합될 수 있는 반응기를 포함할 수 있으나, 이에 제한되지 않는다. Specifically, the non-peptidyl polymer is a reactive group capable of binding to F (eg, immunoglobulin Fc region) and X at both ends, specifically X, or the N-terminus of F (eg, immunoglobulin Fc region) Or it may include a reactive group that can be bonded to the amine group located in the lysine or the thiol group of cysteine, but is not limited thereto.
또한, F, 예컨대 면역글로불린 Fc 영역 및 X와 결합될 수 있는, 상기 비펩타이드성 중합체의 반응기는 알데히드기, 말레이미드기 및 석시니미드 유도체로 구성된 군으로부터 선택될 수 있으나, 이에 제한되지 않는다. In addition, the reactive group of the non-peptide polymer, which may be bound to F, such as an immunoglobulin Fc region and X, may be selected from the group consisting of an aldehyde group, a maleimide group, and a succinimide derivative, but is not limited thereto.
상기에서, 알데히드기로 프로피온 알데히드기 또는 부틸 알데히드기를 예로서 들 수 있으나, 이에 제한되지 않는다. In the above, the aldehyde group may be a propion aldehyde group or a butyl aldehyde group, but is not limited thereto.
상기에서, 석시니미드 유도체로는 석시니미딜 발레르에이트, 석시니미딜 메틸부타노에이트, 석시니미딜 메틸프로피온에이트, 석시니미딜 부타노에이트, 석시니미딜 프로피오네이트, N-하이드록시석시니미드, 히드록시 석시니미딜, 석시니미딜 카르복시메틸 또는 석시니미딜 카보네이트가 이용될 수 있으나, 이에 제한되지 않는다. In the above, the succinimide derivatives include succinimidyl valereate, succinimidyl methylbutanoate, succinimidyl methylpropionate, succinimidyl butanoate, succinimidyl propionate, N-hydroxysuccini Mid, hydroxy succinimidyl, succinimidyl carboxymethyl or succinimidyl carbonate may be used, but is not limited thereto.
비펩타이드성 링커는 이러한 반응기를 통하여 X와 F에 연결될 수 있으나, 특별히 이에 제한되는 것은 아니다.The non-peptidyl linker may be connected to X and F through such a reactive group, but is not particularly limited thereto.
또한, 알데히드 결합에 의한 환원성 아민화로 생성된 최종 산물은 아미드 결합으로 연결된 것보다 훨씬 안정적이다. 알데히드 반응기는 낮은 pH에서 N-말단에 선택적으로 반응하며, 높은 pH, 예를 들어 pH 9.0 조건에서는 리신 잔기와 공유결합을 형성할 수 있다. In addition, the final product produced by reductive amination by aldehyde bonds is much more stable than those linked by amide bonds. The aldehyde reactor reacts selectively to the N-terminus at a low pH, and can form a covalent bond with a lysine moiety at a high pH, for example, pH 9.0.
또한, 상기 비펩타이드성 링커의 양쪽 말단의 반응기는 서로 동일하거나 또는 서로 상이할 수 있으며, 예를 들어, 한쪽 말단에는 말레이미드기를, 다른 쪽 말단에는 알데히드기, 프로피온 알데히드기, 또는 부틸 알데히드기를 가질 수 있다. 그러나, 비펩타이드성 링커의 각 말단에 F, 구체적으로 면역글로불린 Fc 영역과 X가 결합될 수 있다면, 특별히 이에 제한되지 않는다. In addition, the reactive groups at both ends of the non-peptidyl linker may be the same or different from each other, for example, a maleimide group at one end and an aldehyde group, a propion aldehyde group, or a butyl aldehyde group at the other end. . However, if F, specifically immunoglobulin Fc region and X can be bonded to each end of the non-peptide linker, it is not particularly limited thereto.
예를 들어, 상기 비펩타이드성 링커의 한쪽 말단에는 반응기로서 말레이미드 기를 포함하고, 다른 쪽 말단에는 알데히드기, 프로피온 알데히드기 또는 부틸 알데히드기 등을 포함할 수 있다. For example, one end of the non-peptide linker may include a maleimide group as a reactive group, and the other end may include an aldehyde group, a propion aldehyde group, or a butyl aldehyde group.
양쪽 말단에 히드록시 반응기를 갖는 폴리에틸렌 글리콜을 비펩타이드성 중합체로 이용하는 경우에는 공지의 화학반응에 의해 상기 히드록시기를 상기 다양한 반응기로 활성화하거나, 상업적으로 입수 가능한 변형된 반응기를 갖는 폴리에틸렌 글리콜을 이용하여 본 발명의 지속형 단백질 결합체를 제조할 수 있다.In the case of using polyethylene glycol having a hydroxy reactive group at both ends as a non-peptidyl polymer, the hydroxy group may be activated with the various reactive groups by a known chemical reaction, or a polyethylene glycol having a commercially available modified reactive group may be used. The long-acting protein conjugate of the invention can be prepared.
하나의 구체적인 실시 형태에서 상기 비펩타이드성 중합체는 X의 시스테인 잔기, 보다 구체적으로 시스테인의 -SH 기에 연결되는 것일 수 있으나, 이에 제한되지 않는다. In one specific embodiment, the non-peptide polymer may be linked to a cysteine residue of X, more specifically to a -SH group of cysteine, but is not limited thereto.
예컨대, 상기 X에 해당하는 펩타이드에서 10번 시스테인 잔기, 13번 시스테인 잔기, 15번 시스테인 잔기, 17번 시스테인 잔기, 19번 시스테인 잔기, 21번 시스테인 잔기, 24번 시스테인 잔기, 28번 시스테인 잔기, 29번 시스테인 잔기, 30번 시스테인 잔기, 31번 시스테인 잔기, 40번 시스테인 잔기, 또는 41번 시스테인 잔기에 상기 비펩타이드성 중합체가 연결된 것일 수 있으나, 특별히 이에 제한되지 않는다. For example, in the peptide corresponding to X, cysteine residue 10, cysteine 13, cysteine 15, cysteine 17, cysteine 19, cysteine 21, cysteine 24, cysteine 28, 29 The non-peptidyl polymer may be linked to a cysteine residue, a cysteine residue 30, a cysteine 31, a cysteine 40, or a cysteine 41, but is not particularly limited thereto.
구체적으로, 상기 시스테인 잔기의 -SH 기에 비펩타이드성 중합체의 반응기가 연결될 수 있으며, 반응기에 대해서는 앞서 기술한 내용이 모두 적용된다. 만약, 말레이미드-PEG-알데히드를 사용하는 경우, 말레이미드 기는 X의 -SH 기와 티오에테르(thioether) 결합으로 연결하고, 알데히드기는 F, 구체적으로 면역글로불린 Fc의 -NH2기와 환원적 아민화 반응을 통해 연결할 수 있으나, 이에 제한되지 않으며, 이는 하나의 일례에 해당한다. Specifically, a reactive group of a non-peptidyl polymer may be connected to the -SH group of the cysteine residue, and all of the above descriptions apply to the reactive group. If maleimide-PEG-aldehyde is used, the maleimide group is linked by a -SH group of X and a thioether bond, and the aldehyde group is F, specifically, a reductive amination reaction with the -NH 2 group of immunoglobulin Fc. It can be connected through, but is not limited thereto, and this corresponds to one example.
또한, 상기 결합체에서, 비펩타이드성 중합체의 반응기가 면역글로불린 Fc 영역의 N-말단에 위치한 -NH2와 연결된 것일 수 있으나, 이는 하나의 일례에 해당한다.In addition, in the conjugate, the reactive group of the non-peptide polymer may be linked to -NH 2 located at the N-terminus of the immunoglobulin Fc region, but this corresponds to one example.
본 발명에서, "면역글로불린 Fc 영역"은, 면역글로불린의 중쇄와 경쇄 가변영역을 제외한, 중쇄 불변영역 2(CH2) 및/또는 중쇄 불변영역 3(CH3)부분을 포함하는 부위를 의미한다. 상기 면역글로불린 Fc 영역은 본 발명의 결합체의 모이어티를 이루는 일 구성일 수 있다. In the present invention, the "immunoglobulin Fc region" refers to a region including a heavy chain constant region 2 (CH2) and/or a heavy chain constant region 3 (CH3) portion, excluding the heavy and light chain variable regions of an immunoglobulin. The immunoglobulin Fc region may be a component constituting the moiety of the conjugate of the present invention.
이러한 면역글로불린 Fc 영역은 중쇄 불변영역에 힌지(hinge) 부분을 포함할 수 있으나, 이에 제한되는 것은 아니다. 또한 본 발명의 면역글로불린 Fc 영역은 천연형과 실질적으로 동등하거나 향상된 효과를 갖는 한, 면역 글로불린의 중쇄와 경쇄 가변영역만을 제외하고, 일부 또는 전체 중쇄 불변영역 1(CH1) 및/또는 경쇄불변영역 1(CL1)을 포함하는 확장된 Fc 영역일 수 있다. 또한, CH2 및/또는 CH3에 해당하는 상당히 긴 일부 아미노산 서열이 제거된 영역일 수도 있다. Such an immunoglobulin Fc region may include a hinge portion in the heavy chain constant region, but is not limited thereto. In addition, as long as the immunoglobulin Fc region of the present invention has substantially the same or improved effect with the native type, part or all of the heavy chain constant region 1 (CH1) and/or light chain constant region except for the heavy and light chain variable regions of the immunoglobulin It may be an expanded Fc region comprising 1 (CL1). In addition, it may be a region from which some considerably long amino acid sequences corresponding to CH2 and/or CH3 have been removed.
예컨대, 본 발명의 면역글로불린 Fc 영역은 1) CH1 도메인, CH2 도메인, CH3 도메인 및 CH4 도메인, 2) CH1 도메인 및 CH2 도메인, 3) CH1 도메인 및 CH3 도메인, 4) CH2 도메인 및 CH3 도메인, 5) CH1 도메인, CH2 도메인, CH3 도메인 및 CH4 도메인 중 1개 또는 2개의 이상의 도메인과 면역글로불린 힌지 영역(또는 힌지 영역의 일부)와의 조합, 6) 중쇄 불변 영역 각 도메인과 경쇄 불변영역의 이량체일 수 있다. 그러나, 이에 제한되는 것은 아니다. For example, the immunoglobulin Fc region of the present invention is 1) CH1 domain, CH2 domain, CH3 domain and CH4 domain, 2) CH1 domain and CH2 domain, 3) CH1 domain and CH3 domain, 4) CH2 domain and CH3 domain, 5) Combination of one or two or more domains of the CH1 domain, CH2 domain, CH3 domain, and CH4 domain with an immunoglobulin hinge region (or part of the hinge region), 6) heavy chain constant region It may be a dimer of each domain and light chain constant region. . However, it is not limited thereto.
또한, 하나의 구체예로서, 상기 면역글로불린 Fc 영역은 이합체 형태 (dimeric form)일 수 있으며, 이합체 형태의 하나의 Fc 영역에 X 한 분자가 공유결합적으로 연결될 수 있으며, 이때 상기 면역글로불린 Fc와 X는 비펩타이드성 중합체에 의해 서로 연결될 수 있다. 한편, 이합체 형태의 하나의 Fc 영역에 X 두 분자가 대칭적으로 결합하는 것 역시 가능하다. 이때 상기 면역글로불린 Fc와 X는 비펩타이드성 링커에 의해 서로 연결될 수 있다. 그러나, 상기 기술된 예에 제한되는 것은 아니다. In addition, as an embodiment, the immunoglobulin Fc region may be in a dimeric form, and one X molecule may be covalently linked to one Fc region in a dimer form, and at this time, the immunoglobulin Fc and X can be linked to each other by a non-peptidyl polymer. On the other hand, it is also possible for two X molecules to symmetrically bind to one Fc region in the form of a dimer. At this time, the immunoglobulin Fc and X may be linked to each other by a non-peptide linker. However, it is not limited to the examples described above.
또한, 본 발명의 면역글로불린 Fc 영역은 천연형 아미노산 서열뿐만 아니라 이의 서열 유도체를 포함한다. 아미노산 서열 유도체란 천연 아미노산 서열 중의 하나 이상의 아미노산 잔기가 결실, 삽입, 비보전적 또는 보전적 치환 또는 이들의 조합에 의하여 상이한 서열을 가지는 것을 의미한다.In addition, the immunoglobulin Fc region of the present invention includes a native amino acid sequence as well as a sequence derivative thereof. An amino acid sequence derivative means that one or more amino acid residues in a natural amino acid sequence have a different sequence by deletion, insertion, non-conservative or conservative substitution, or a combination thereof.
예를 들면, IgG Fc의 경우 결합에 중요하다고 알려진 214 내지 238, 297 내지 299, 318 내지 322 또는 327 내지 331번 아미노산 잔기들이 변형을 위해 적당한 부위로서 이용될 수 있다.For example, in the case of IgG Fc, amino acid residues 214 to 238, 297 to 299, 318 to 322, or 327 to 331, which are known to be important for binding, may be used as a suitable site for modification.
또한, 이황화 결합을 형성할 수 있는 부위가 제거되거나, 천연형 Fc에서 N-말단의 몇몇 아미노산이 제거되거나 또는 천연형 Fc의 N-말단에 메티오닌 잔기가 부가될 수도 있는 등 다양한 종류의 유도체가 가능하다. 또한, 이펙터 기능을 없애기 위해 보체결합부위, 예로 C1q 결합부위가 제거될 수도 있고, ADCC (antibody dependent cell mediated cytotoxicity) 부위가 제거될 수도 있다. 이러한 면역글로불린 Fc 영역의 서열 유도체를 제조하는 기술은 국제특허공개 제WO 97/34631호, 국제특허공개 제96/32478호 등에 개시되어 있다.In addition, various kinds of derivatives are possible, such as a site capable of forming a disulfide bond is removed, several amino acids at the N-terminus in a native Fc are removed, or a methionine residue may be added to the N-terminus of a native Fc. Do. In addition, the complement binding site, eg, the C1q binding site, may be removed to eliminate the effector function, or the ADCC (antibody dependent cell mediated cytotoxicity) site may be removed. Techniques for preparing a sequence derivative of such an immunoglobulin Fc region are disclosed in International Patent Publication No. WO 97/34631, International Patent Publication No. 96/32478, and the like.
분자의 활성을 전체적으로 변경시키지 않는 단백질 및 펩타이드에서의 아미노산 교환은 당해 분야에 공지되어 있다 (H.Neurath, R.L.Hill, The Proteins, Academic Press, New York, 1979). 가장 통상적으로 일어나는 교환은 아미노산 잔기 Ala/Ser, Val/Ile, Asp/Glu, Thr/Ser, Ala/Gly, Ala/Thr, Ser/Asn, Ala/Val, Ser/Gly, Thy/Phe, Ala/Pro, Lys/Arg, Asp/Asn, Leu/Ile, Leu/Val, Ala/Glu, Asp/Gly 간의 교환이다. 경우에 따라서는 인산화(phosphorylation), 황화(sulfation), 아크릴화(acrylation), 당화(glycosylation), 메틸화(methylation), 파네실화(farnesylation), 아세틸화(acetylation) 및 아미드화(amidation) 등으로 수식(modification)될 수도 있다.Amino acid exchanges in proteins and peptides that do not totally alter the activity of the molecule are known in the art (H. Neuroth, R.L. Hill, The Proteins, Academic Press, New York, 1979). The most commonly occurring exchanges are amino acid residues Ala/Ser, Val/Ile, Asp/Glu, Thr/Ser, Ala/Gly, Ala/Thr, Ser/Asn, Ala/Val, Ser/Gly, Thy/Phe, Ala/ It is an exchange between Pro, Lys/Arg, Asp/Asn, Leu/Ile, Leu/Val, Ala/Glu, Asp/Gly. In some cases, formulas such as phosphorylation, sulfation, acrylation, glycosylation, methylation, farnesylation, acetylation and amidation ( may be modified.
상기 기술한 Fc 유도체는 본 발명의 Fc 영역과 동등한 생물학적 활성을 나타내며 Fc 영역의 열, pH 등에 대한 구조적 안정성을 증대시킨 것일 수 있다. The Fc derivative described above exhibits a biological activity equivalent to that of the Fc region of the present invention, and may have increased structural stability against heat and pH of the Fc region.
또한, 이러한 Fc 영역은 인간, 소, 염소, 돼지, 마우스, 래빗, 햄스터, 랫트 또는 기니아 픽 등의 동물의 생체 내에서 분리한 천연형으로부터 얻어질 수도 있고, 형질전환된 동물세포 또는 미생물로부터 얻어진 재조합형 또는 이의 유도체일 수 있다. 여기서, 천연형으로부터 획득하는 방법은 전체 면역글로불린을 인간 또는 동물의 생체로부터 분리한 후, 단백질 분해효소를 처리하여 획득하는 방법일 수 있다. 파파인을 처리할 경우에는 Fab 및 Fc로 절단되고, 펩신을 처리할 경우에는 pF'c 및 F(ab)2로 절단된다. 이를 크기 배제 크로마토그래피 (size-exclusion chromatography) 등을 이용하여 Fc 또는 pF'c를 분리할 수 있다. 더 구체적인 실시 형태에서는 인간 유래의 Fc 영역을 미생물로부터 수득한 재조합형 면역글로불린 Fc 영역이다.In addition, these Fc regions may be obtained from natural types isolated in vivo from animals such as humans, cows, goats, pigs, mice, rabbits, hamsters, rats or guinea pigs, or obtained from transformed animal cells or microorganisms. It may be recombinant or a derivative thereof. Here, the method of obtaining from the natural form may be a method of obtaining the whole immunoglobulin by separating it from a human or animal body and then treating a protease. When papain is treated, it is cleaved into Fab and Fc, and when treated with pepsin, it is cleaved into pF'c and F(ab) 2 . This can be separated from Fc or pF'c using size-exclusion chromatography or the like. In a more specific embodiment, the human-derived Fc region is a recombinant immunoglobulin Fc region obtained from a microorganism.
또한, 면역글로불린 Fc 영역은 천연형 당쇄, 천연형에 비해 증가된 당쇄, 천연형에 비해 감소한 당쇄 또는 당쇄가 제거된 형태일 수 있다. 이러한 면역글로불린 Fc 당쇄의 증감 또는 제거에는 화학적 방법, 효소학적 방법 및 미생물을 이용한 유전 공학적 방법과 같은 통상적인 방법이 이용될 수 있다. 여기서, Fc에서 당쇄가 제거된 면역글로불린 Fc 영역은 보체(c1q)와의 결합력이 현저히 저하되고, 항체-의존성 세포독성 또는 보체-의존성 세포 독성이 감소 또는 제거되므로, 생체 내에서 불필요한 면역 반응을 유발하지 않는다. 이런 점에서 약물의 캐리어로서의 본래의 목적에 보다 부합하는 형태는 당쇄가 제거되거나 비당쇄화된 면역글로불린 Fc 영역이라 할 것이다.In addition, the immunoglobulin Fc region may be a natural type sugar chain, an increased sugar chain compared to the natural type, a reduced sugar chain compared to the natural type, or a form in which sugar chains are removed. Conventional methods such as chemical methods, enzymatic methods, and genetic engineering methods using microorganisms may be used for the increase or decrease of such immunoglobulin Fc sugar chains. Here, the immunoglobulin Fc region from which the sugar chain has been removed from the Fc significantly decreases the binding ability with complement (c1q), and the antibody-dependent cytotoxicity or complement-dependent cytotoxicity is reduced or eliminated, so it does not induce unnecessary immune responses in vivo. Does not. In this respect, a form more suitable for the original purpose as a drug carrier would be referred to as an immunoglobulin Fc region in which sugar chains are removed or non-glycosylated.
본 발명에서 "당쇄의 제거(Deglycosylation)"는 효소로 당을 제거한 Fc 영역을 말하며, 비당쇄화(Aglycosylation)는 원핵동물, 더 구체적인 실시 형태에서는 대장균에서 생산하여 당쇄화되지 않은 Fc 영역을 의미한다.In the present invention, "deglycosylation" refers to an Fc region from which sugar is removed by an enzyme, and aglycosylation refers to an Fc region that is not glycosylated by producing in a prokaryotic animal, or in a more specific embodiment, E. coli. .
한편, 면역글로불린 Fc 영역은 인간 또는 소, 염소, 돼지, 마우스, 래빗, 햄스터, 랫트, 기니아 픽 등의 동물기원일 수 있으며, 더 구체적인 실시 형태에서는 인간기원이다.On the other hand, the immunoglobulin Fc region may be of human origin or animal origin, such as cattle, goats, pigs, mice, rabbits, hamsters, rats, and guinea pigs, and in more specific embodiments, human origin.
또한, 면역글로불린 Fc 영역은 IgG, IgA, IgD, IgE, IgM 유래 또는 이들의 조합(combination) 또는 이들의 혼성(hybrid)에 의한 Fc 영역일 수 있다. 더 구체적인 실시 형태에서는 인간 혈액에 가장 풍부한 IgG 또는 IgM유래이며, 보다 더 구체적인 실시 형태에서는 리간드 결합 단백질의 반감기를 향상시키는 것으로 공지된 IgG 유래이다. 더욱 더 구체적인 실시 형태에서 상기 면역글로불린 Fc 영역은 IgG4 Fc 영역이며, 가장 구체적인 실시 형태에서 상기 면역글로불린 Fc 영역은 인간 IgG4 유래의 비-당쇄화된 Fc 영역이나, 이에 제한되는 것은 아니다. In addition, the immunoglobulin Fc region may be an Fc region derived from IgG, IgA, IgD, IgE, or IgM, or a combination thereof or a hybrid thereof. In a more specific embodiment, it is derived from IgG or IgM, which is most abundant in human blood, and in a more specific embodiment, it is derived from IgG known to improve the half-life of a ligand binding protein. In a more specific embodiment, the immunoglobulin Fc region is an IgG4 Fc region, and in the most specific embodiment, the immunoglobulin Fc region is a non-glycosylated Fc region derived from human IgG4, but is not limited thereto.
한편, 본 발명에서 "조합(combination)"이란 이량체 또는 다량체를 형성할 때, 동일 기원 단쇄 면역글로불린 Fc 영역을 암호화하는 폴리펩타이드가 상이한 기원의 단쇄 폴리펩타이드와 결합을 형성하는 것을 의미한다. 즉, IgG Fc, IgA Fc, IgM Fc, IgD Fc 및 IgE의 Fc 단편으로 이루어진 그룹으로부터 선택된 2개 이상의 단편으로부터 이량체 또는 다량체의 제조가 가능하다.Meanwhile, in the present invention, "combination" means that when forming a dimer or multimer, a polypeptide encoding a single-chain immunoglobulin Fc region of the same origin forms a bond with a single-chain polypeptide of a different origin. That is, it is possible to prepare a dimer or a multimer from two or more fragments selected from the group consisting of IgG Fc, IgA Fc, IgM Fc, IgD Fc and IgE Fc fragment.
또한, 상술한 결합체는 효력의 지속성이 천연형 GLP-1, GIP, 혹은 글루카곤에 비해, 또는 F가 수식되지 않은 X에 비해 증가된 것일 수 있으며, 이러한 결합체는 상술한 형태뿐만 아니라, 생분해성 나노파티클에 봉입된 형태 등을 모두 포함한다. In addition, the above-described conjugate may be that the persistence of the potency is increased compared to the native GLP-1, GIP, or glucagon, or F is not modified compared to X, and such conjugates are not only in the above-described form, but also biodegradable nano Includes all shapes enclosed in particles, etc.
상기 펩타이드 (예컨대 상기 펩타이드 자체 또는 이에 생체적합성 물질이 결합된 형태)를 포함하는 조성물은 고지혈증의 예방 또는 치료용일 수 있다. A composition comprising the peptide (eg, the peptide itself or a form in which a biocompatible material is combined) may be used for the prevention or treatment of hyperlipidemia.
본 발명에서 용어 "예방"은 상기 펩타이드 또는 이를 포함하는 조성물의 투여로 고지혈증의 발병을 억제 또는 지연시키는 모든 행위를 의미하며, "치료"는 상기 펩타이드 또는 이를 포함하는 조성물의 투여로 고지혈증의 증세가 호전되거나 이롭게 되는 모든 행위를 의미한다.In the present invention, the term "prevention" refers to any action that suppresses or delays the onset of hyperlipidemia by administration of the peptide or a composition comprising the same, and "treatment" refers to the symptoms of hyperlipidemia by administration of the peptide or a composition comprising the same. It refers to any action that improves or benefits.
본 발명에서 용어 "투여"는 어떠한 적절한 방법으로 환자에게 소정의 물질을 도입하는 것을 의미하며, 상기 조성물의 투여 경로는 특별히 이에 제한되지 않으나, 상기 조성물이 생체 내 표적에 도달할 수 있는 어떠한 일반적인 경로를 통하여 투여될 수 있으며, 예를 들어 복강 내 투여, 정맥 내 투여, 근육 내 투여, 피하 투여, 피 내 투여, 경구 투여, 국소 투여, 비 내 투여, 폐 내 투여, 또는 직장 내 투여 등이 될 수 있다.In the present invention, the term "administration" means introducing a predetermined substance to the patient by any suitable method, and the route of administration of the composition is not particularly limited thereto, but any general route by which the composition can reach the target in vivo Can be administered through, for example, intraperitoneal administration, intravenous administration, intramuscular administration, subcutaneous administration, intradermal administration, oral administration, topical administration, intranasal administration, intrapulmonary administration, or rectal administration. I can.
본 발명의 약학적 조성물은 약학적으로 허용가능한 담체, 부형제 또는 희석제를 추가로 포함할 수 있다. 이러한 약학적으로 허용가능한 담체, 부형제, 또는 희석제는 비자연적으로 발생된 것일 수 있다. The pharmaceutical composition of the present invention may further include a pharmaceutically acceptable carrier, excipient, or diluent. Such a pharmaceutically acceptable carrier, excipient, or diluent may be non-naturally occurring.
본 발명에서 용어 "약학적으로 허용가능한"이란 치료효과를 나타낼 수 있을 정도의 충분한 양과 부작용을 일으키지 않는 것을 의미하며, 질환의 종류, 환자의 연령, 체중, 건강, 성별, 환자의 약물에 대한 민감도, 투여 경로, 투여 방법, 투여횟수, 치료 기간, 배합 또는 동시 사용되는 약물 등 의학 분야에 잘 알려진 요소에 따라 당업자에 의해 용이하게 결정될 수 있다.In the present invention, the term "pharmaceutically acceptable" means a sufficient amount to exhibit a therapeutic effect and does not cause side effects, and the type of disease, the patient's age, weight, health, sex, and the patient's sensitivity to drugs , The route of administration, the method of administration, the number of times of administration, the duration of the treatment, and drugs used in combination or concurrently can be easily determined by those skilled in the art according to factors well known in the medical field.
본 발명의 펩타이드, 또는 결합체를 포함한 약학적 조성물은 약학적으로 허용가능한 담체를 추가로 포함할 수 있다. 상기 담체는 특별히 이에 제한되지는 않으나, 경구 투여 시에는 결합제, 활택제, 붕해제, 부형제, 가용화제, 분산제, 안정화제, 현탁화제, 색소, 향료 등을 사용할 수 있고, 주사제의 경우에는 완충제, 보존제, 무통화제, 가용화제, 등장화제, 안정화제 등을 혼합하여 사용할 수 있으며, 국소투여용의 경우에는 기제, 부형제, 윤활제, 보존제 등을 사용할 수 있다. The pharmaceutical composition including the peptide or conjugate of the present invention may further include a pharmaceutically acceptable carrier. The carrier is not particularly limited thereto, but when administered orally, a binder, a lubricant, a disintegrant, an excipient, a solubilizing agent, a dispersing agent, a stabilizing agent, a suspending agent, a coloring agent, a flavoring agent, etc. may be used.In the case of an injection, a buffer agent, Preservatives, painless agents, solubilizers, isotonic agents, stabilizers, and the like can be mixed and used. In the case of topical administration, base agents, excipients, lubricants, preservatives, etc. can be used.
본 발명의 조성물의 제형은 상술한 바와 같은 약학적으로 허용가능한 담체와 혼합하여 다양하게 제조될 수 있다. 예를 들어, 경구 투여 시에는 정제, 트로키, 캡슐, 엘릭서, 서스펜션, 시럽, 웨이퍼 등의 형태로 제조할 수 있으며, 주사제의 경우에는 단위 투약 앰플 또는 다수회 투약 형태로 제조할 수 있다. 기타, 용액, 현탁액, 정제, 환약, 캡슐, 서방형 제제 등으로 제형화할 수 있다.The formulation of the composition of the present invention can be variously prepared by mixing with a pharmaceutically acceptable carrier as described above. For example, when administered orally, it can be prepared in the form of tablets, troches, capsules, elixirs, suspensions, syrups, wafers, etc., and in the case of injections, it can be prepared in the form of unit dosage ampoules or multiple dosage forms. Others, solutions, suspensions, tablets, pills, capsules, can be formulated as sustained-release preparations.
한편, 제제화에 적합한 담체, 부형제 및 희석제의 예로는 락토스, 덱스트로스, 수크로스, 솔비톨, 만니톨, 자일리톨, 에리스리톨, 말티톨, 전분, 아카시아, 알지네이트, 젤라틴, 칼슘 포스페이트, 칼슘 실리케이트, 셀룰로스, 메틸 셀룰로스, 미정질 셀룰로스, 폴리비닐피롤리돈, 물, 메틸하이드록시벤조에이트, 프로필하이드록시벤조에이트, 탈크, 마그네슘 스테아레이트 또는 광물유 등이 사용될 수 있다. 또한, 충진제, 항응집제, 윤활제, 습윤제, 향료, 방부제 등을 추가로 포함할 수 있다. Meanwhile, examples of carriers, excipients and diluents suitable for formulation include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, Microcrystalline cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, mineral oil, and the like may be used. In addition, fillers, anti-aggregating agents, lubricants, wetting agents, flavoring agents, preservatives, and the like may additionally be included.
또한, 본 발명의 약학적 조성물은 정제, 환제, 산제, 과립제, 캡슐제, 현탁제, 내용액제, 유제, 시럽제, 멸균된 수용액, 비수성용제, 동결건조제제 및 좌제로 이루어진 군으로부터 선택되는 어느 하나의 제형을 가질 수 있다.In addition, the pharmaceutical composition of the present invention is any one selected from the group consisting of tablets, pills, powders, granules, capsules, suspensions, liquid solutions, emulsions, syrups, sterilized aqueous solutions, non-aqueous solutions, freeze-dried preparations, and suppositories. It can have a formulation of.
또한, 상기 조성물은 약학적 분야에서 통상의 방법에 따라 환자의 신체 내 투여에 적합한 단위 투여형의 제제, 구체적으로는 단백질 의약품의 투여에 유용한 제제 형태로 제형화시켜 당업계에서 통상적으로 사용하는 투여 방법을 이용하여 경구, 또는 피부, 정맥 내, 근육 내, 동맥 내, 골수 내, 수막강 내, 심실 내, 폐, 경피, 피하, 복 내, 비강 내, 소화관 내, 국소, 설하, 질 내 또는 직장 경로를 포함하는 비경구 투여 경로에 의하여 투여될 수 있으나, 이들에 한정되는 것은 아니다.In addition, the composition is formulated in a unit dosage form suitable for intra-body administration of a patient according to a conventional method in the pharmaceutical field, specifically, in a form useful for administration of protein medicines, and administration commonly used in the art. Orally, or skin, intravenous, intramuscular, intraarterial, intramedullary, intrameningal, intraventricular, pulmonary, transdermal, subcutaneous, intraperitoneal, intranasal, gastrointestinal, topical, sublingual, intravaginal or It may be administered by a parenteral route including a rectal route, but is not limited thereto.
또한, 상기 펩타이드 또는 결합체는 생리식염수 또는 유기용매와 같이 약제로 허용된 여러 전달체(carrier)와 혼합하여 사용될 수 있고, 안정성이나 흡수성을 증가시키기 위하여 글루코스, 수크로스 또는 덱스트란과 같은 탄수화물, 아스코르브산(ascorbic acid) 또는 글루타티온과 같은 항산화제(antioxidants), 킬레이트제, 저분자 단백질 또는 다른 안정화제(stabilizers) 등이 약제로 사용될 수 있다.In addition, the peptide or conjugate may be used in combination with various carriers permitted as drugs such as physiological saline or an organic solvent, and carbohydrates such as glucose, sucrose or dextran, ascorbic acid, in order to increase stability or absorption. Antioxidants such as (ascorbic acid) or glutathione, chelating agents, low-molecular proteins, or other stabilizers may be used as drugs.
본 발명의 약학적 조성물의 투여량과 횟수는 치료할 질환, 투여 경로, 환자의 연령, 성별 및 체중 및 질환의 중등도 등의 여러 관련 인자와 함께, 활성성분인 약물의 종류에 따라 결정된다. The dosage and frequency of the pharmaceutical composition of the present invention are determined according to the type of drug as the active ingredient, along with various related factors such as the disease to be treated, the route of administration, the age, sex and weight of the patient, and the severity of the disease.
본 발명의 조성물의 총 유효량은 단일 투여량 (single dose)으로 환자에게 투여될 수 있으며, 다중 투여량 (multiple dose)으로 장기간 투여되는 분할 치료 방법 (fractionated treatment protocol)에 의해 투여될 수 있다. 본 발명의 약학적 조성물은 질환의 정도에 따라 유효성분의 함량을 달리할 수 있다. 구체적으로, 본 발명의 결합체의 바람직한 전체 용량은 하루에 환자 체중 1 kg당 약 0.0001 mg 내지 500 mg일 수 있다. 그러나 상기 결합체의 용량은 약학적 조성물의 투여 경로 및 치료 횟수뿐만 아니라 환자의 연령, 체중, 건강 상태, 성별, 질환의 중증도, 식이 및 배설율 등 다양한 요인들을 고려하여 환자에 대한 유효 투여량이 결정되는 것이므로, 이러한 점을 고려할 때 당분야의 통상적인 지식을 가진 자라면 상기 본 발명의 조성물의 특정한 용도에 따른 적절한 유효 투여량을 결정할 수 있을 것이다. 본 발명에 따른 약학적 조성물은 본 발명의 효과를 보이는 한 그 제형, 투여 경로 및 투여 방법에 특별히 제한되지 아니한다.The total effective amount of the composition of the present invention may be administered to a patient in a single dose, and may be administered by a fractionated treatment protocol that is administered for a long period of time in multiple doses. The pharmaceutical composition of the present invention may vary the content of the active ingredient according to the severity of the disease. Specifically, the preferred total dose of the conjugate of the present invention may be about 0.0001 mg to 500 mg per 1 kg of patient body weight per day. However, the dose of the conjugate is determined by considering various factors such as the patient's age, weight, health status, sex, disease severity, diet and excretion rate, as well as the administration route and number of treatments of the pharmaceutical composition. Therefore, in consideration of these points, those of ordinary skill in the art will be able to determine an appropriate effective dosage according to the specific use of the composition of the present invention. The pharmaceutical composition according to the present invention is not particularly limited in its formulation, route of administration, and method of administration as long as it shows the effect of the present invention.
본 발명의 약학적 조성물은 생체 내 지속성 및 역가가 우수하여, 본 발명의 약학적 제제의 투여 횟수 및 빈도를 현저하게 감소시킬 수 있다.The pharmaceutical composition of the present invention has excellent in vivo persistence and potency, and can significantly reduce the number and frequency of administration of the pharmaceutical formulation of the present invention.
본 발명을 구현하는 다른 하나의 양태는 상기 펩타이드, 결합체 또는 이를 포함하는 조성물을 이를 필요로 하는 개체에 투여하는 단계를 포함하는, 고지혈증의 예방 또는 치료 방법을 제공한다. Another aspect embodying the present invention provides a method for preventing or treating hyperlipidemia, comprising administering the peptide, conjugate, or composition comprising the same to an individual in need thereof.
상기 펩타이드, 결합체 또는 이를 포함하는 조성물, 고지혈증, 예방 및 치료에 대해서는 앞서 설명한 바와 같다. The peptide, conjugate, or composition comprising the same, hyperlipidemia, prevention and treatment are as described above.
본 발명에서 상기 개체는 고지혈증이 의심되는 개체로서, 상기 고지혈증 의심 개체는 해당 질환이 발병하였거나 발병할 수 있는 인간을 포함한 쥐, 가축 등을 포함하는 포유동물을 의미하나, 본 발명의 결합체 혹은 이를 포함하는 상기 조성물로 치료 가능한 개체는 제한 없이 포함된다. In the present invention, the subject is an individual suspected of hyperlipidemia, and the suspected hyperlipidemia refers to mammals including mice, livestock, etc., including humans who have or may develop the disease, but the conjugate of the present invention or include the same Subjects treatable with the composition described above are included without limitation.
본 발명의 방법은 상기 펩타이드 또는 결합체를 포함하는 약학적 조성물을 약학적 유효량으로 투여하는 것을 포함할 수 있다. 적합한 총 1일 사용량은 올바른 의학적 판단범위 내에서 처치의에 의해 결정될 수 있으며, 1회 또는 수회로 나누어 투여할 수 있다. 그러나 본 발명의 목적상, 특정 환자에 대한 구체적인 치료적 유효량은 달성하고자 하는 반응의 종류와 정도, 경우에 따라 다른 제제가 사용되는지의 여부를 비롯한 구체적 조성물, 환자의 연령, 체중, 일반 건강 상태, 성별 및 식이, 투여 시간, 투여 경로 및 조성물의 분비율, 치료기간, 구체적 조성물과 함께 사용되거나 동시 사용되는 약물을 비롯한 다양한 인자와 의약 분야에 잘 알려진 유사 인자에 따라 다르게 적용하는 것이 바람직하다.The method of the present invention may include administering a pharmaceutical composition containing the peptide or conjugate in a pharmaceutically effective amount. The appropriate total daily use amount may be determined by the treating physician within the range of correct medical judgment, and may be administered once or in several divided doses. However, for the purposes of the present invention, a specific therapeutically effective amount for a specific patient is a specific composition, including the type and degree of reaction to be achieved, whether other agents are used in some cases, the patient's age, weight, general health status, It is preferable to apply differently according to various factors including sex and diet, administration time, administration route and secretion rate of the composition, treatment period, drugs used with or concurrently with the specific composition, and similar factors well known in the medical field.
본 발명을 구현하는 다른 하나의 양태는 고지혈증의 예방 또는 치료를 위한 약제의 제조에 있어서 상기 펩타이드, 결합체 또는 이를 포함하는 조성물의 용도이다. Another aspect embodying the present invention is the use of the peptide, conjugate, or a composition comprising the same in the manufacture of a medicament for the prevention or treatment of hyperlipidemia.
상기 펩타이드, 결합체 또는 이를 포함하는 조성물, 고지혈증, 예방 및 치료에 대해서는 앞서 설명한 바와 같다. The peptide, conjugate, or composition comprising the same, hyperlipidemia, prevention and treatment are as described above.
이하, 하기 실시예에 의하여 본 발명을 보다 상세하게 설명한다. 단, 하기 실시예는 본 발명을 예시하기 위한 것일 뿐 본 발명의 범위가 이들로 한정되는 것은 아니다.Hereinafter, the present invention will be described in more detail by the following examples. However, the following examples are only for illustrating the present invention, and the scope of the present invention is not limited thereto.
실시예 1: 삼중 활성체의 제조Example 1: Preparation of triple active substance
GLP-1, GIP 및 글루카곤 수용체에 모두 활성을 나타내는 삼중 활성체를 제조하여, 하기 표 1에 이의 서열을 나타냈다.A triple activator exhibiting activity on all of GLP-1, GIP and glucagon receptors was prepared, and their sequences are shown in Table 1 below.
Figure PCTKR2020005233-appb-T000001
Figure PCTKR2020005233-appb-T000001
Figure PCTKR2020005233-appb-I000001
Figure PCTKR2020005233-appb-I000001
Figure PCTKR2020005233-appb-I000002
Figure PCTKR2020005233-appb-I000002
Figure PCTKR2020005233-appb-I000003
Figure PCTKR2020005233-appb-I000003
Figure PCTKR2020005233-appb-I000004
Figure PCTKR2020005233-appb-I000004
Figure PCTKR2020005233-appb-I000005
Figure PCTKR2020005233-appb-I000005
상기 표 1에 기재된 서열에서 X로 표기된 아미노산은 비천연형 아미노산인 Aib (aminoisobutyric acid)이며, 밑줄로 표시된 아미노산은 밑줄로 표시된 아미노산들이 서로 고리를 형성하는 것을 의미한다. 또한, 상기 표 1에서 CA는 4-이미다조아세틸(4-imidazoacetyl)을, Y는 티로신을 의미한다. In the sequence shown in Table 1, the amino acid indicated by X is non-natural amino acid Aib (aminoisobutyric acid), and the underlined amino acid means that the underlined amino acids form a ring with each other. In addition, in Table 1, CA represents 4-imidazoacetyl, and Y represents tyrosine.
실시예 2: 삼중 활성체의 지속형 결합체 제조Example 2: Preparation of long-acting conjugates of triple active agents
양 말단에 각각 말레이미드기 및 알데히드기를 가지는 10kDa의 PEG, 즉 말레이미드-PEG-알데히드 (10kDa, NOF, 일본)를 실시예 1의 삼중활성체 (서열번호 21, 22, 42, 43, 50, 77, 및 96)의 시스테인 잔기에 페길화시키기 위하여, 삼중활성체와 말레이미드-PEG-알데히드의 몰비를 1 : 1 내지 3, 단백질의 농도를 1 내지 5 ㎎/㎖로 하여 저온에서 0.5 내지 3 시간 동안 반응시켰다. 이때, 반응은 50 mM Tris 완충액(pH 7.5)에 20 내지 60% 아이소프로판올이 첨가된 환경 하에서 수행되었다. 반응이 종료된 후, 상기 반응액을 SP 세파로스 HP (GE healthcare, 미국)에 적용하여 시스테인에 모노-페길화된 삼중활성체를 정제하였다.10 kDa PEG having a maleimide group and an aldehyde group at both ends, that is, maleimide-PEG-aldehyde (10 kDa, NOF, Japan) was used as the triple active agent of Example 1 (SEQ ID NOs: 21, 22, 42, 43, 50, 77, and 96) in order to PEGylate the cysteine residue, the molar ratio of the triactivator to the maleimide-PEG-aldehyde is 1:1 to 3, and the protein concentration is 1 to 5 mg/ml, and 0.5 to 3 at low temperature. Reacted for hours. At this time, the reaction was carried out in an environment in which 20 to 60% isopropanol was added to 50 mM Tris buffer (pH 7.5). After the reaction was completed, the reaction solution was applied to SP Sepharose HP (GE healthcare, USA) to purify a triactivator mono-pegylated to cysteine.
다음으로, 상기 정제된 모노-페길화된 삼중활성체와 면역글로불린 Fc을 몰비를 1 : 1 내지 5, 단백질의 농도를 10 내지 50㎎/㎖로 하여 4 내지 8℃에서 12 내지 18시간 동안 반응시켰다. 반응은 100 mM 인산칼륨 완충액 (pH 6.0)에 환원제인 10 내지 50 mM 소디움시아노보로하이드라이드와 10 내지 30 % 아이소프로판올이 첨가된 환경 하에서 수행되었다. 반응이 종료된 후, 상기 반응액을 부틸 세파로스 FF 정제컬럼 (GE healthcare, 미국)과 Source ISO 정제컬럼 (GE healthcare, 미국)에 적용하여, 삼중활성체와 면역글로불린 Fc를 포함하는 결합체를 정제하였다.Next, the purified mono-pegylated triactivator and immunoglobulin Fc were reacted at a molar ratio of 1: 1 to 5, and a protein concentration of 10 to 50 mg/ml for 12 to 18 hours at 4 to 8°C. Made it. The reaction was carried out in an environment in which 10 to 50 mM sodium cyanoborohydride and 10 to 30% isopropanol as a reducing agent were added to 100 mM potassium phosphate buffer (pH 6.0). After the reaction is complete, the reaction solution is applied to a butyl Sepharose FF purification column (GE healthcare, USA) and a Source ISO purification column (GE healthcare, USA) to purify a conjugate including a triple activator and an immunoglobulin Fc. I did.
제조 후 역상 크로마토그래피, 크기배제 크로마토그래피 및 이온교환 크로마토그래피로 분석한 순도는 95 % 이상이었다.After preparation, the purity analyzed by reverse phase chromatography, size exclusion chromatography and ion exchange chromatography was 95% or more.
본 실시예에서 제조된 삼중활성체 및 면역글로불린 Fc가 PEG를 통하여 연결된 결합체는 삼중결합체 및 면역글로불린 Fc를 포함하는 결합체 또는 지속형 결합체로 명명하였다.The conjugate in which the triple active agent and immunoglobulin Fc prepared in this example are linked through PEG is referred to as a conjugate or long-acting conjugate including a triple conjugate and an immunoglobulin Fc.
예를 들어, 서열번호 42의 삼중활성체 및 면역글로불린 Fc가 PEG를 통하여 연결된 결합체를, '서열번호 42과 면역글로불린 Fc를 포함하는 결합체' 혹은 '서열번호 42의 지속형 결합체'로 명명하였고, 이들은 본원에서 혼용되어 사용될 수 있다.For example, a conjugate in which the triple activator of SEQ ID NO: 42 and immunoglobulin Fc are linked through PEG was named as'conjugate comprising SEQ ID NO: 42 and immunoglobulin Fc' or'long-acting conjugate of SEQ ID NO: 42', These may be used interchangeably herein.
실험예 1: 삼중 활성체 및 이의 지속형 결합체의Experimental Example 1: Triple activator and its long-acting conjugate in vitro in vitro 활성 측정 Activity measurement
상기 실시예 1 및 2에서 제조된 삼중활성체와 이의 지속형 결합체의 활성을 측정하기 위해 GLP-1 수용체, 글루카곤(GCG) 수용체, 및 GIP 수용체가 각각 형질전환된 세포주를 이용하여 in vitro에서 세포 활성을 측정하는 방법을 이용하였다.Cells in vitro using cell lines transformed with GLP-1 receptor, glucagon (GCG) receptor, and GIP receptor, respectively, to measure the activity of the triple activator prepared in Examples 1 and 2 and its long-acting conjugate. A method of measuring activity was used.
상기 각 세포주는 CHO (chinese hamster ovary)에 인간 GLP-1 수용체, 인간 GCG 수용체 및 인간 GIP 수용체 유전자를 각각 발현하도록 형질 전환된 것으로서, GLP-1, GCG 및 GIP의 활성을 측정하기에 적합하다. 따라서, 각 부분에 대한 활성을 각각의 형질 전환 세포주를 이용하여 측정하였다.Each of the cell lines is transformed to express human GLP-1 receptor, human GCG receptor and human GIP receptor genes in CHO (chinese hamster ovary), and is suitable for measuring the activities of GLP-1, GCG and GIP. Therefore, the activity for each part was measured using each transformed cell line.
상기 실시예 1과 2에서 제조된 삼중활성체와 이의 지속형 결합체의 GLP-1 활성 측정을 위해 인간 GLP-1을 50nM 부터 4배씩 0.000048nM까지 연속적으로 희석하고, 상기 실시예 1과 2에서 제조된 삼중활성체와 이의 지속형 결합체를 400nM 부터 4배씩 0.00038nM까지 연속적으로 희석하였다. 상기 배양된 인간 GLP-1 수용체가 발현된 CHO 세포에서 배양액을 제거하고 연속적으로 희석된 각 물질들을 5㎕씩 상기 세포에 첨가한 다음, cAMP 항체가 포함된 완충액을 5㎕씩 추가 한 뒤 15분 동안 상온에서 배양하였다. 그런 다음 세포용해완충액 (cell lysis buffer)이 포함된 detection mix를 10㎕씩 가하여 세포를 용해시키고, 90분 동안 상온에서 반응시켰다. 상기 반응이 완료된 세포용해물을 LANCE cAMP kit (PerkinElmer, USA)에 적용하여 축적된 cAMP를 통해 EC50값을 산출한 후, 상호 비교하였다. 인간 GLP-1 대비 상대 역가는 하기 표 2와 표 3에 나타내었다.In order to measure the GLP-1 activity of the triple activator prepared in Examples 1 and 2 and its long-acting conjugate, human GLP-1 was serially diluted from 50 nM to 0.000048 nM by 4 times, and prepared in Examples 1 and 2 The triple activator and its long-acting conjugate were serially diluted from 400nM to 0.00038nM by 4 times. Remove the culture medium from the cultured human GLP-1 receptor-expressing CHO cells, add 5 µl of each of the serially diluted substances to the cells, and then add 5 µl each of a buffer containing cAMP antibody for 15 minutes. During incubation at room temperature. Then, 10 μl of a detection mix containing a cell lysis buffer was added each to lyse the cells, and the cells were reacted at room temperature for 90 minutes. The cell lysate for which the reaction was completed was applied to a LANCE cAMP kit (PerkinElmer, USA) to calculate an EC 50 value from the accumulated cAMP, and then compared with each other. The relative titers compared to human GLP-1 are shown in Tables 2 and 3 below.
상기 실시예 1과 2에서 제조된 삼중활성체와 이의 지속형 결합체의 GCG 활성 측정을 위해 인간 GCG을 50nM 부터 4배씩 0.000048nM까지 연속적으로 희석하고, 상기 실시예 1과 2에서 제조된 삼중활성체와 이의 지속형 결합체를 400nM 부터 4배씩 0.00038nM까지 연속적으로 희석하였다. 상기 배양된 인간 GCG 수용체가 발현된 CHO 세포에서 배양액을 제거하고 연속적으로 희석된 각 물질들을 5㎕씩 상기 세포에 첨가한 다음, cAMP 항체가 포함된 완충액을 5㎕씩 추가 한 뒤 15분 동안 상온에서 배양하였다. 그런 다음 세포용해완충액(cell lysis buffer)이 포함된 detection mix를 10㎕씩 가하여 세포를 용해시키고, 90분 동안 상온에서 반응시켰다. 상기 반응이 완료된 세포용해물을 LANCE cAMP kit (PerkinElmer, USA)에 적용하여 축적된 cAMP를 통해 EC50값을 산출한 후, 상호 비교하였다. 인간 GCG 대비 상대 역가는 하기 표2와 표3에 나타내었다.To measure the GCG activity of the triple activator prepared in Examples 1 and 2 and the long-acting conjugate thereof, human GCG was serially diluted from 50 nM to 0.000048 nM by 4 times, and the triple activator prepared in Examples 1 and 2 And its long-acting conjugates were serially diluted from 400 nM to 0.00038 nM in 4-fold increments. Remove the culture medium from the cultured human GCG receptor-expressing CHO cells, add 5 µl of each of the serially diluted substances to the cells, and then add 5 µl of cAMP antibody-containing buffer at room temperature for 15 minutes. Cultured in. Then, 10 μl of a detection mix containing a cell lysis buffer was added each to lyse the cells, and the cells were reacted at room temperature for 90 minutes. The cell lysate for which the reaction was completed was applied to a LANCE cAMP kit (PerkinElmer, USA) to calculate an EC 50 value from the accumulated cAMP, and then compared with each other. Relative titers compared to human GCG are shown in Tables 2 and 3 below.
상기 실시예 1과 2에서 제조된 삼중활성체와 이의 지속형 결합체의 GIP 활성 측정을 위해 인간 GIP을 50nM 부터 4배씩 0.000048nM까지 연속적으로 희석하고, 상기 실시예 1과 2에서 제조된 삼중활성체와 이의 지속형 결합체를 400nM 부터 4배씩 0.00038nM까지 연속적으로 희석하였다. 상기 배양된 인간 GIP 수용체가 발현된 CHO 세포에서 배양액을 제거하고 연속적으로 희석된 각 물질들을 5㎕씩 상기 세포에 첨가한 다음, cAMP 항체가 포함된 완충액을 5㎕씩 추가 한 뒤 15분 동안 상온에서 배양하였다. 그런 다음 세포용해완충액(cell lysis buffer)이 포함된 detection mix를 10㎕씩 가하여 세포를 용해시키고, 90분 동안 상온에서 반응시켰다. 상기 반응이 완료된 세포용해물을 LANCE cAMP kit (PerkinElmer, USA)에 적용하여 축적된 cAMP를 통해 EC50값을 산출한 후, 상호 비교하였다. 인간 GIP 대비 상대 역가는 하기 표 2와 표 3에 나타내었다.To measure the GIP activity of the triple activator prepared in Examples 1 and 2 and the long-acting conjugate thereof, human GIP was serially diluted from 50 nM to 0.000048 nM by 4 times, and the triple activator prepared in Examples 1 and 2 And its long-acting conjugate were serially diluted from 400 nM to 0.00038 nM in 4 fold increments. Remove the culture medium from the cultured human GIP receptor-expressing CHO cells, add 5 µl of each of the serially diluted substances to the cells, and add 5 µl of a buffer containing cAMP antibody at room temperature for 15 minutes. Cultured in. Then, 10 μl of a detection mix containing a cell lysis buffer was added each to lyse the cells, and the cells were reacted at room temperature for 90 minutes. The cell lysate for which the reaction was completed was applied to a LANCE cAMP kit (PerkinElmer, USA) to calculate an EC 50 value from the accumulated cAMP, and then compared with each other. Relative titers compared to human GIP are shown in Tables 2 and 3 below.
Figure PCTKR2020005233-appb-T000002
Figure PCTKR2020005233-appb-T000002
Figure PCTKR2020005233-appb-I000006
Figure PCTKR2020005233-appb-I000006
Figure PCTKR2020005233-appb-I000007
Figure PCTKR2020005233-appb-I000007
Figure PCTKR2020005233-appb-I000008
Figure PCTKR2020005233-appb-I000008
Figure PCTKR2020005233-appb-T000003
Figure PCTKR2020005233-appb-T000003
상기 표 3은 삼중활성체 지속형 결합체의 상대적 역가 비율을 확인한 표이며, 상기에서 제조한 신규한 삼중활성체 지속형 결합체는 GLP-1 수용체, GIP 수용체 및 글루카곤 수용체를 모두 활성화시킬 수 있는 삼중 활성체로 기능을 가지는 것을 확인하였다.Table 3 is a table confirming the relative titer ratio of the triple active long-acting conjugate, and the novel triple-active long-acting conjugate prepared above has triple activity capable of activating all of the GLP-1 receptor, GIP receptor, and glucagon receptor. It was confirmed that it has a sieve function.
실험예 2: 삼중활성체 지속형 결합체의 고지혈증 치료 효과 확인(In vivo)Experimental Example 2: Confirmation of the effect of treatment of hyperlipidemia of the triple active agent long-acting conjugate (In vivo)
Golden syrian hamster(이하 고지혈증 햄스터)에 2주간 Fructose 식이섭취를 진행하여 고지혈증 모델을 유도하였다. 실시예 1에서 제조된 삼중결합체 중 서열번호 42를 선정하여 서열번호 42의 지속형 결합체(1.6, 3.1 nmol/kg, Q2D)를 고지혈증을 유도한 햄스터에 3주간 피하를 통해 반복투여 하였다. 비교군으로는 시중의 고지혈증 치료제인 evolocumab (219.2 nmol/kg, QW)을 사용하여, 고지혈증 치료 효과를 확인하였다.A hyperlipidemia model was induced by feeding Fructose for 2 weeks in a Golden syrian hamster (hereinafter referred to as hyperlipidemia hamster). Among the triple conjugates prepared in Example 1, SEQ ID NO: 42 was selected, and the long-acting conjugate of SEQ ID NO: 42 (1.6, 3.1 nmol/kg, Q2D) was repeatedly administered subcutaneously for 3 weeks to hamsters inducing hyperlipidemia. As a control group, evolocumab (219.2 nmol/kg, QW), a commercially available treatment for hyperlipidemia, was used, and the effect of treating hyperlipidemia was confirmed.
도 1에 나타난 바와 같이, 서열번호 42의 지속형 결합체를 3주간 반복 투여한 시 혈중 총 콜레스테롤(50~60% 감소) 및 LDL (70 ~85% 감소)이 유의적으로 감소한 것을 확인한 반면, evolocumab 투여군에서는 미비한 효능을 보였다. As shown in Figure 1, when repeatedly administered the long-acting conjugate of SEQ ID NO: 42 for 3 weeks, it was confirmed that total cholesterol (50-60% reduction) and LDL (70-85% reduction) in blood significantly decreased, whereas evolocumab In the administration group, it showed insufficient efficacy.
실험예 3: 삼중활성체 지속형 결합체의 고지혈증 치료 효과 확인(In vitro)Experimental Example 3: Confirmation of the effect of treatment of hyperlipidemia of the triple active agent long-acting conjugate (In vitro)
HepG2 cell을 이용하여 LDL 흡수 증가, 3-Hydroxy-3-methylglutaryl-CoA reductase (HMGCR) 활성 억제, 및 지방산 분해 촉진 효과 확인 실험을 통해, 실시예 2의 삼중활성체 지속형 결합체의 우수한 고지혈증 치료효능 작용기전 (Mode of action)을 in vitro에서 확인하였다.By using HepG2 cells to increase LDL absorption, inhibit 3-Hydroxy-3-methylglutaryl-CoA reductase (HMGCR) activity, and promote fatty acid decomposition effects, the triple active long-acting conjugate of Example 2 has excellent hyperlipidemia treatment efficacy. The mode of action was confirmed in vitro .
(1) 삼중활성체 지속형 결합체에 의한 LDL 흡수 증가 (1) Increased LDL absorption by the triple-active long-acting conjugate
HepG2 cell에 대조군, 서열번호 42의 지속형 결합체 (0.01, 0.1, 1, 10 μM) 및 evolocumab (10 μg/mL)을 48시간 처리한 후, BODIPY 표지된 LDL의 세포 내 흡수량을 확인하였으며, 그 결과를 도 2에 나타내었다.HepG2 cells were treated with the control, the long-acting conjugate of SEQ ID NO: 42 (0.01, 0.1, 1, 10 μM) and evolocumab (10 μg/mL) for 48 hours, and then the amount of BODIPY-labeled LDL uptake was confirmed. The results are shown in FIG. 2.
도 2에 나타난 바와 같이, LDL-흡수량이 서열번호 42의 지속형 결합체에 의해 농도의존적 (control 대비 132.7~354%) 으로 증가하는 것을 확인하였다.As shown in FIG. 2, it was confirmed that the LDL-absorption amount increased in a concentration-dependent manner (132.7-354% compared to control) by the long-acting conjugate of SEQ ID NO: 42.
(2) 삼중활성체 지속형 결합체에 의한 3-Hydroxy-3-methylglutaryl-CoA reductase (HMGCR) 활성 억제(2) Inhibition of 3-Hydroxy-3-methylglutaryl-CoA reductase (HMGCR) activity by a triple-active long-acting conjugate
콜레스테롤 합성 율속 효소(rate limiting enzyme)인 3-Hydroxy-3-methylglutaryl-CoA reductase (이하 HMGCR)의 활성을 측정하기 위해, HepG2 cell에 서열번호 42의 지속형 결합체를 처리한 후 HMG-CoA를 처리조건에서 NADPH의 감소정도를 측정하였고 (HMGCR은 NADPH를 이용하여 HMG-COA를 콜레스테롤 전구체인 mevalonate로 전환한다.), 그 결과를 도 3에 나타내었다.To measure the activity of 3-Hydroxy-3-methylglutaryl-CoA reductase (hereinafter HMGCR), a rate limiting enzyme for cholesterol synthesis, HepG2 cells were treated with the long-acting conjugate of SEQ ID NO: 42 and then HMG-CoA was treated. The degree of reduction of NADPH was measured under conditions (HMGCR converts HMG-COA to cholesterol precursor mevalonate using NADPH), and the results are shown in FIG. 3.
도 3에 나타난 바와 같이, 서열번호 42의 지속형 결합체가 HMGCR의 활성을 지속적으로 억제함 (control 대비 약 80%감소)을 확인하였다.As shown in Figure 3, it was confirmed that the long-acting conjugate of SEQ ID NO: 42 continuously inhibits the activity of HMGCR (about 80% reduction compared to control).
(3) 삼중활성체 지속형 결합체에 의한 지방산 분해 촉진(3) Acceleration of fatty acid decomposition by long-acting triple active conjugate
지방산 분해의 부산물인 케톤체(ketone body)를 측정하였다. 이를 위해 지방산 (palmitate)를 처리한 HepG2 cell에 서열번호 42의 지속형 결합체를 처리한 후, 구매가능한 케톤체 측정 kit (Sigma #MAK041)를 이용하여 생성된 케톤체 양을 측정하였다.The ketone body, a by-product of fatty acid decomposition, was measured. To this end, after treatment with the long-acting conjugate of SEQ ID NO: 42 in HepG2 cells treated with fatty acid (palmitate), the amount of the produced ketone body was measured using a commercially available ketone body measurement kit (Sigma #MAK041).
측정 결과, 도 4에 나타난 바와 같이, 케톤체 생성이 서열번호 42의 지속형 결합체 농도 의존적으로 증가한 것을 확인 하였다. As a result of the measurement, as shown in FIG. 4, it was confirmed that the production of ketone bodies increased depending on the concentration of the long-acting complex of SEQ ID NO: 42.
상기와 같은 실험으로부터 본 발명에 따른 삼중활성체 및 이의 지속형 결합체는 LDL 흡수 증가, HMGCR 활성 억제를 통한 콜레스테롤 합성 저해, 및 지방산 분해 촉진을 통해 혈중 총 콜레스테롤 및 LDL을 감소시킴으로써, 고지혈증에 대한 치료 효과를 나타낼 수 있음을 확인하였다.From the above experiments, the triple activator and the long-acting conjugate thereof according to the present invention increases LDL absorption, inhibits cholesterol synthesis through inhibition of HMGCR activity, and reduces total cholesterol and LDL in blood through promotion of fatty acid degradation, thereby treating hyperlipidemia. It was confirmed that the effect can be exhibited.
이상의 설명으로부터, 본 발명이 속하는 기술분야의 당업자는 본 발명이 그 기술적 사상이나 필수적 특징을 변경하지 않고서 다른 구체적인 형태로 실시될 수 있다는 것을 이해할 수 있을 것이다. 이와 관련하여, 이상에서 기술한 실시예들은 모든 면에서 예시적인 것이며 한정적인 것이 아닌 것으로서 이해해야만 한다. 본 발명의 범위는 상기 상세한 설명보다는 후술하는 특허 청구범위의 의미 및 범위 그리고 그 등가 개념으로부터 도출되는 모든 변경 또는 변형된 형태가 본 발명의 범위에 포함되는 것으로 해석되어야 한다.From the above description, those skilled in the art to which the present invention pertains will be able to understand that the present invention can be implemented in other specific forms without changing the technical spirit or essential features thereof. In this regard, the embodiments described above are illustrative in all respects and should be understood as non-limiting. The scope of the present invention should be construed that all changes or modifications derived from the meaning and scope of the claims to be described later rather than the above detailed description, and equivalent concepts thereof, are included in the scope of the present invention.

Claims (17)

  1. 글루카곤 수용체, GLP-1 (Glucagon-like peptide-1) 수용체, 및 GIP (Glucose-dependent insuliontropic polypeptide) 수용체에 대해 활성을 가지는 분리된 펩타이드를 포함하며,It includes an isolated peptide having activity against a glucagon receptor, a GLP-1 (Glucagon-like peptide-1) receptor, and a GIP (Glucose-dependent insuliontropic polypeptide) receptor,
    상기 펩타이드는 하기 일반식 1로 표시되는 아미노산 서열을 포함하는 것인, 고지혈증의 예방 또는 치료를 위한 약학적 조성물:The peptide comprises an amino acid sequence represented by the following general formula 1, a pharmaceutical composition for the prevention or treatment of hyperlipidemia:
    Xaa1-Xaa2-Xaa3-Gly-Thr-Phe-Xaa7-Ser-Asp-Xaa10-Ser-Xaa12-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp-Leu-Xaa27-Xaa28-Xaa29-Xaa30-R1 (일반식 1, 서열번호 103)Xaa1-Xaa2-Xaa3-Gly-Thr-Phe-Xaa7-Ser-Asp-Xaa10-Ser-Xaa12-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp- Leu-Xaa27-Xaa28-Xaa29-Xaa30-R1 (general formula 1, SEQ ID NO: 103)
    상기 일반식 1에서, Xaa1은 히스티딘 (His, H), 4-이미다조아세틸 (CA), 또는 티로신 (Tyr, Y)이고, Xaa2는 글리신 (Gly, G), 알파-메틸-글루탐산, 또는 Aib (aminoisobutyric acid)이며, Xaa3은 글루탐산 (Glu, E) 또는 글루타민 (Gln, Q)이고, Xaa7은 트레오닌 (Thr, T) 또는 이소류신 (Ile, I)이며, Xaa10은 류신 (Leu, L), 티로신 (Tyr, Y), 리신 (Lys, K), 시스테인 (Cys, C), 또는 발린 (Val, V)이고, Xaa12는 리신 (Lys, K), 세린 (Ser, S), 또는 이소류신 (Ile, I)이며, Xaa13은 글루타민 (Gln, Q), 티로신 (Tyr, Y), 알라닌 (Ala, A), 또는 시스테인 (Cys, C)이고, Xaa14는 류신 (Leu, L), 메티오닌 (Met, M), 또는 티로신 (Tyr, Y)이며, Xaa15는 시스테인 (Cys, C), 아스파르트산 (Asp, D), 글루탐산 (Glu, E), 또는 류신 (Leu, L)이며, Xaa16은 글리신 (Gly, G), 글루탐산 (Glu, E), 또는 세린 (Ser, S)이고, Xaa17은 글루타민 (Gln, Q), 아르기닌 (Arg, R), 이소류신 (Ile, I), 글루탐산 (Glu, E), 시스테인 (Cys, C), 또는 리신 (Lys, K)이며, Xaa18은 알라닌 (Ala, A), 글루타민 (Gln, Q), 아르기닌 (Arg, R), 또는 히스티딘 (His, H)이고, Xaa19는 알라닌 (Ala, A), 글루타민 (Gln, Q), 시스테인 (Cys, C), 또는 발린 (Val, V)이며, Xaa20은 리신 (Lys, K), 글루타민 (Gln, Q), 또는 아르기닌 (Arg, R)이고, Xaa21은 글루탐산 (Glu, E), 글루타민 (Gln, Q), 류신 (Leu, L), 시스테인 (Cys, C), 또는 아스파르트산 (Asp, D)이며, Xaa23은 이소류신 (Ile, I) 또는 발린 (Val, V)이고, Xaa24는 알라닌 (Ala, A), 글루타민 (Gln, Q), 시스테인 (Cys, C), 아스파라긴 (Asn, N), 아스파르트산 (Asp, D), 또는 글루탐산 (Glu, E)이며, Xaa27은 발린 (Val, V), 류신 (Leu, L), 리신 (Lys, K), 또는 메티오닌 (Met, M)이고, Xaa28은 시스테인 (Cys, C), 리신 (Lys, K), 알라닌 (Ala, A), 아스파라긴 (Asn, N), 또는 아스파르트산 (Asp, D)이며, Xaa29는 시스테인 (Cys, C), 글리신 (Gly, G), 글루타민 (Gln, Q), 트레오닌 (Thr, T), 글루탐산 (Glu, E), 또는 히스티딘 (His, H)이고, Xaa30은 시스테인 (Cys, C), 글리신 (Gly, G), 리신 (Lys, K), 또는 히스티딘 (His, H)이거나, 부존재하며,In the general formula 1, Xaa1 is histidine (His, H), 4-imidazoacetyl (CA), or tyrosine (Tyr, Y), and Xaa2 is glycine (Gly, G), alpha-methyl-glutamic acid, or Aib (aminoisobutyric acid), Xaa3 is glutamic acid (Glu, E) or glutamine (Gln, Q), Xaa7 is threonine (Thr, T) or isoleucine (Ile, I), Xaa10 is leucine (Leu, L), tyrosine (Tyr, Y), lysine (Lys, K), cysteine (Cys, C), or valine (Val, V), and Xaa12 is lysine (Lys, K), serine (Ser, S), or isoleucine (Ile, I), Xaa13 is glutamine (Gln, Q), tyrosine (Tyr, Y), alanine (Ala, A), or cysteine (Cys, C), and Xaa14 is leucine (Leu, L), methionine (Met, M ), or tyrosine (Tyr, Y), Xaa15 is cysteine (Cys, C), aspartic acid (Asp, D), glutamic acid (Glu, E), or leucine (Leu, L), and Xaa16 is glycine (Gly, G), glutamic acid (Glu, E), or serine (Ser, S), Xaa17 is glutamine (Gln, Q), arginine (Arg, R), isoleucine (Ile, I), glutamic acid (Glu, E), cysteine (Cys, C), or lysine (Lys, K), Xaa18 is alanine (Ala, A), glutamine (Gln, Q), arginine (Arg, R), or histidine (His, H), and Xaa19 is alanine (Ala, A), glutamine (Gln, Q), cysteine (Cys, C), or valine (Val, V), and Xaa20 is lysine (Lys, K), glutamine (Gln, Q), or arginine (Arg, R), Xaa21 is glutamic acid (Glu, E), glutamine (Gln, Q), leucine (Leu, L), cysteine (Cys, C), or aspartic acid (Asp, D), and Xaa23 is isoleucine (Ile, I) again Is valine (Val, V), Xaa24 is alanine (Ala, A), glutamine (Gln, Q), cysteine (Cys, C), asparagine (Asn, N), aspartic acid (Asp, D), or glutamic acid ( Glu, E), Xaa27 is valine (Val, V), leucine (Leu, L), lysine (Lys, K), or methionine (Met, M), and Xaa28 is cysteine (Cys, C), lysine (Lys , K), alanine (Ala, A), asparagine (Asn, N), or aspartic acid (Asp, D), Xaa29 is cysteine (Cys, C), glycine (Gly, G), glutamine (Gln, Q) , Threonine (Thr, T), glutamic acid (Glu, E), or histidine (His, H), Xaa30 is cysteine (Cys, C), glycine (Gly, G), lysine (Lys, K), or histidine ( His, H) or absent,
    R1은 시스테인 (Cys, C), GKKNDWKHNIT (서열번호 106), m-SSGAPPPS-n (서열번호 107), 또는 m-SSGQPPPS-n (서열번호 108)이거나, 부존재하며, R1 is cysteine (Cys, C), GKKNDWKHNIT (SEQ ID NO: 106), m-SSGAPPPS-n (SEQ ID NO: 107), or m-SSGQPPPS-n (SEQ ID NO: 108), or absent,
    여기서, m은 -Cys-, -Pro-, 또는 -Gly-Pro-이고, n은 -Cys-, -Gly-, -Ser-, 또는 -His-Gly-이거나, 부존재함.Here, m is -Cys-, -Pro-, or -Gly-Pro-, and n is -Cys-, -Gly-, -Ser-, or -His-Gly- or absent.
  2. 제1항에 있어서, 상기 일반식 1에서, Xaa14는 류신 또는 메티오닌이며, Xaa15는 시스테인, 아스파르트산, 또는 류신인, 약학적 조성물. The pharmaceutical composition according to claim 1, wherein in the general formula 1, Xaa14 is leucine or methionine, and Xaa15 is cysteine, aspartic acid, or leucine.
  3. 제 1항에 있어서, 상기 일반식 1에서, Xaa2는 글리신, 알파-메틸-글루탐산, 또는 Aib이고, Xaa7은 트레오닌이며, Xaa10은 티로신, 시스테인, 또는 발린이고, Xaa12는 리신 또는 이소류신이며, Xaa13은 티로신, 알라닌, 글루타민, 또는 시스테인이고, Xaa14는 류신, 시스테인, 또는 메티오닌이며, Xaa15는 시스테인, 류신, 글루탐산, 또는 아스파르트산이고, Xaa17은 글루타민, 아르기닌, 이소류신, 시스테인, 글루탐산, 또는 리신이며, Xaa18은 알라닌, 글루타민, 아르기닌, 또는 히스티딘이고, Xaa19는 알라닌, 글루타민, 발린, 또는 시스테인이며, Xaa20은 리신, 아르기닌, 또는 글루타민이고, Xaa21은 글루탐산, 글루타민, 류신, 시스테인, 또는 아스파르트산이며, Xaa23은 이소류신 또는 발린이고, Xaa24는 시스테인, 알라닌, 글루타민, 아스파라긴, 글루탐산, 또는 아스파르트산이며, Xaa27은 류신 또는 리신인, 약학적 조성물. The method of claim 1, wherein in the general formula 1, Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib, Xaa7 is threonine, Xaa10 is tyrosine, cysteine, or valine, Xaa12 is lysine or isoleucine, and Xaa13 is Tyrosine, alanine, glutamine, or cysteine, Xaa14 is leucine, cysteine, or methionine, Xaa15 is cysteine, leucine, glutamic acid, or aspartic acid, Xaa17 is glutamine, arginine, isoleucine, cysteine, glutamic acid, or lysine, Xaa18 Is alanine, glutamine, arginine, or histidine, Xaa19 is alanine, glutamine, valine, or cysteine, Xaa20 is lysine, arginine, or glutamine, Xaa21 is glutamic acid, glutamine, leucine, cysteine, or aspartic acid, and Xaa23 is Isoleucine or valine, Xaa24 is cysteine, alanine, glutamine, asparagine, glutamic acid, or aspartic acid, and Xaa27 is leucine or lysine, a pharmaceutical composition.
  4. 제1항에 있어서, 상기 펩타이드는 하기 일반식 2로 표시되는 아미노산 서열을 포함하는 펩타이드인, 약학적 조성물:The pharmaceutical composition of claim 1, wherein the peptide is a peptide comprising an amino acid sequence represented by the following general formula 2:
    Xaa1-Xaa2-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Xaa10-Ser-Lys-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp-Leu-Leu-Xaa28-Xaa29-Xaa30-Xaa31-Ser-Ser-Gly-Gln-Pro-Pro-Pro-Ser-Xaa40 (일반식 2, 서열번호 104)Xaa1-Xaa2-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Xaa10-Ser-Lys-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp- Leu-Leu-Xaa28-Xaa29-Xaa30-Xaa31-Ser-Ser-Gly-Gln-Pro-Pro-Pro-Ser-Xaa40 (general formula 2, SEQ ID NO: 104)
    상기 일반식 2에서, Xaa1은 4-이미다조아세틸, 히스티딘, 또는 티로신이고, Xaa2는 글리신, 알파-메틸-글루탐산, 또는 Aib이며, Xaa10은 티로신, 또는 시스테인이고, Xaa13은 알라닌, 글루타민, 티로신, 또는 시스테인이며, Xaa14는 류신, 메티오닌, 또는 티로신이고, Xaa15는 아스파르트산, 글루탐산, 또는 류신이며, Xaa16은 글리신, 글루탐산, 또는 세린이고, Xaa17은 글루타민, 아르기닌, 이소류신, 글루탐산, 시스테인, 또는 리신이며, Xaa18은 알라닌, 글루타민, 아르기닌, 또는 히스티딘이고, Xaa19는 알라닌, 글루타민, 시스테인, 또는 발린이며, Xaa20은 리신, 글루타민, 또는 아르기닌이고, Xaa21은 시스테인, 글루탐산, 글루타민, 류신, 또는 아스파르트산이며, Xaa23은 이소류신 또는 발린이고, Xaa24는 시스테인, 알라닌, 글루타민, 아스파라긴, 또는 글루탐산이며, Xaa28은 리신, 시스테인, 아스파라긴, 또는 아스파르트산이고, Xaa29는 글리신, 글루타민, 시스테인, 또는 히스티딘이며, Xaa30은 시스테인, 글리신, 리신, 또는 히스티딘이고, Xaa31은 프롤린 또는 시스테인이며, Xaa40은 시스테인이거나, 부존재함.In Formula 2, Xaa1 is 4-imidazoacetyl, histidine, or tyrosine, Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib, Xaa10 is tyrosine, or cysteine, and Xaa13 is alanine, glutamine, tyrosine, Or cysteine, Xaa14 is leucine, methionine, or tyrosine, Xaa15 is aspartic acid, glutamic acid, or leucine, Xaa16 is glycine, glutamic acid, or serine, and Xaa17 is glutamine, arginine, isoleucine, glutamic acid, cysteine, or lysine. , Xaa18 is alanine, glutamine, arginine, or histidine, Xaa19 is alanine, glutamine, cysteine, or valine, Xaa20 is lysine, glutamine, or arginine, and Xaa21 is cysteine, glutamic acid, glutamine, leucine, or aspartic acid, Xaa23 is isoleucine or valine, Xaa24 is cysteine, alanine, glutamine, asparagine, or glutamic acid, Xaa28 is lysine, cysteine, asparagine, or aspartic acid, Xaa29 is glycine, glutamine, cysteine, or histidine, Xaa30 is cysteine, Glycine, lysine, or histidine, Xaa31 is proline or cysteine, and Xaa40 is cysteine or absent.
  5. 제1항에 있어서, 상기 일반식 1에서, Xaa2는 글리신, 알파-메틸-글루탐산, 또는 Aib이고, Xaa7은 트레오닌이며, Xaa10은 티로신, 시스테인, 또는 발린이고, Xaa12는 리신 또는 이소류신이며, Xaa13은 티로신, 알라닌, 또는 시스테인이고, Xaa14는 류신 또는 메티오닌이며, Xaa15는 시스테인 또는 아스파르트산이고, Xaa17은 글루타민, 아르기닌, 이소류신, 시스테인, 또는 리신이며, Xaa18은 알라닌, 아르기닌, 또는 히스티딘이고, Xaa19는 알라닌, 글루타민, 또는 시스테인이며, Xaa20은 리신 또는 글루타민이고, Xaa21은 글루탐산, 시스테인, 또는 아스파르트산이며, Xaa23은 발린이고, Xaa24는 알라닌, 글루타민, 시스테인, 아스파라긴, 또는 아스파르트산이며, Xaa27은 류신 또는 리신인, 약학적 조성물. The method of claim 1, wherein in the general formula 1, Xaa2 is glycine, alpha-methyl-glutamic acid, or Aib, Xaa7 is threonine, Xaa10 is tyrosine, cysteine, or valine, Xaa12 is lysine or isoleucine, and Xaa13 is Tyrosine, alanine, or cysteine, Xaa14 is leucine or methionine, Xaa15 is cysteine or aspartic acid, Xaa17 is glutamine, arginine, isoleucine, cysteine, or lysine, Xaa18 is alanine, arginine, or histidine, and Xaa19 is alanine. , Glutamine, or cysteine, Xaa20 is lysine or glutamine, Xaa21 is glutamic acid, cysteine, or aspartic acid, Xaa23 is valine, Xaa24 is alanine, glutamine, cysteine, asparagine, or aspartic acid, and Xaa27 is leucine or lysine. Phosphorus, pharmaceutical composition.
  6. 제2항에 있어서, 상기 일반식 2에서, Xaa13은 알라닌, 티로신, 또는 시스테인이고, Xaa15는 아스파르트산 또는 글루탐산이며, Xaa17은 글루타민, 아르기닌, 시스테인, 또는 리신이고, Xaa18은 알라닌, 아르기닌, 또는 히스티딘이며, Xaa21은 시스테인, 글루탐산, 글루타민, 또는 아스파르트산이고, Xaa23은 이소류신 또는 발린이며, Xaa24는 시스테인, 글루타민, 또는 아스파라긴이고, Xaa28은 시스테인, 아스파라긴, 또는 아스파르트산이며, Xaa29는 글루타민, 시스테인, 또는 히스티딘이고, Xaa30은 시스테인, 리신, 또는 히스티딘인, 약학적 조성물. The method of claim 2, wherein in the general formula 2, Xaa13 is alanine, tyrosine, or cysteine, Xaa15 is aspartic acid or glutamic acid, Xaa17 is glutamine, arginine, cysteine, or lysine, and Xaa18 is alanine, arginine, or histidine. , Xaa21 is cysteine, glutamic acid, glutamine, or aspartic acid, Xaa23 is isoleucine or valine, Xaa24 is cysteine, glutamine, or asparagine, Xaa28 is cysteine, asparagine, or aspartic acid, and Xaa29 is glutamine, cysteine, or Histidine and Xaa30 is cysteine, lysine, or histidine.
  7. 제1항에 있어서, 상기 일반식 1에서, Xaa2는 알파-메틸-글루탐산 또는 Aib이고, Xaa7은 트레오닌이며, Xaa10은 티로신 또는 시스테인이고, Xaa12는 리신 또는 이소류신이며, Xaa13은 티로신, 알라닌, 또는 시스테인이고, Xaa14는 류신 또는 메티오닌이며, Xaa15는 시스테인 또는 아스파르트산이고, Xaa16은 글루탐산이며, Xaa17은 아르기닌, 이소류신, 시스테인, 또는 리신이고, Xaa18은 알라닌, 아르기닌, 또는 히스티딘이며, Xaa19는 알라닌, 글루타민, 또는 시스테인이고, Xaa20은 리신 또는 글루타민이며, Xaa21은 글루탐산 또는 아스파르트산이고, Xaa23은 발린이며, Xaa24는 글루타민, 아스파라긴, 또는 아스파르트산이고, Xaa27은 류신이며, Xaa28은 시스테인, 알라닌, 아스파라긴, 또는 아스파르트산인, 약학적 조성물. The method of claim 1, wherein in the general formula 1, Xaa2 is alpha-methyl-glutamic acid or Aib, Xaa7 is threonine, Xaa10 is tyrosine or cysteine, Xaa12 is lysine or isoleucine, and Xaa13 is tyrosine, alanine, or cysteine. , Xaa14 is leucine or methionine, Xaa15 is cysteine or aspartic acid, Xaa16 is glutamic acid, Xaa17 is arginine, isoleucine, cysteine, or lysine, Xaa18 is alanine, arginine, or histidine, and Xaa19 is alanine, glutamine, Or cysteine, Xaa20 is lysine or glutamine, Xaa21 is glutamic acid or aspartic acid, Xaa23 is valine, Xaa24 is glutamine, asparagine, or aspartic acid, Xaa27 is leucine, Xaa28 is cysteine, alanine, asparagine, or aspartic acid. Sanin, pharmaceutical composition.
  8. 제1항에 있어서, 상기 일반식 1에서, Xaa1은 히스티딘 또는 4-이미다조아세틸이고, Xaa2는 알파-메틸-글루탐산 또는 Aib이며, Xaa3은 글루타민이고, Xaa7은 트레오닌이며, Xaa10은 티로신이고, Xaa12는 이소류신이며, Xaa13은 알라닌 또는 시스테인이고, Xaa14는 메티오닌이며, Xaa15는 아스파르트산이고, Xaa16은 글루탐산이며, Xaa17은 이소류신 또는 리신이고, Xaa18은 알라닌 또는 히스티딘이며, Xaa19는 글루타민 또는 시스테인이고, Xaa20은 리신이며, Xaa21은 아스파르트산이고, Xaa23은 발린이며, Xaa24는 아스파라긴이고, Xaa27은 류신이며, Xaa28은 알라닌 또는 아스파라긴이고, Xaa29는 글루타민 또는 트레오닌이며, Xaa30은 시스테인 또는 리신이거나, 부존재하는, 약학적 조성물. The method of claim 1, wherein in the general formula 1, Xaa1 is histidine or 4-imidazoacetyl, Xaa2 is alpha-methyl-glutamic acid or Aib, Xaa3 is glutamine, Xaa7 is threonine, Xaa10 is tyrosine, and Xaa12 Is isoleucine, Xaa13 is alanine or cysteine, Xaa14 is methionine, Xaa15 is aspartic acid, Xaa16 is glutamic acid, Xaa17 is isoleucine or lysine, Xaa18 is alanine or histidine, Xaa19 is glutamine or cysteine, Xaa20 is Lysine, Xaa21 is aspartic acid, Xaa23 is valine, Xaa24 is asparagine, Xaa27 is leucine, Xaa28 is alanine or asparagine, Xaa29 is glutamine or threonine, Xaa30 is cysteine or lysine, or is absent, pharmaceutical Composition.
  9. 제1항에 있어서, 상기 펩타이드는 하기 일반식 3의 아미노산 서열을 포함하는 펩타이드인, 약학적 조성물:The pharmaceutical composition of claim 1, wherein the peptide is a peptide comprising the amino acid sequence of the following general formula 3:
    Xaa1-Xaa2-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-Lys-Xaa13-Leu-Asp-Glu-Xaa17-Xaa18-Xaa19-Lys-Xaa21-Phe-Val-Xaa24-Trp-Leu-Leu-Xaa28-Xaa29-Xaa30-Xaa31-Ser-Ser-Gly-Gln-Pro-Pro-Pro-Ser-Xaa40 (일반식 3, 서열번호 105),Xaa1-Xaa2-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Tyr-Ser-Lys-Xaa13-Leu-Asp-Glu-Xaa17-Xaa18-Xaa19-Lys-Xaa21-Phe-Val-Xaa24-Trp- Leu-Leu-Xaa28-Xaa29-Xaa30-Xaa31-Ser-Ser-Gly-Gln-Pro-Pro-Pro-Ser-Xaa40 (general formula 3, SEQ ID NO: 105),
    상기 일반식 3에서, Xaa1은 히스티딘 또는 티로신이고, Xaa2는 알파-메틸-글루탐산 또는 Aib이며, Xaa13은 알라닌, 티로신 또는 시스테인이고, Xaa17은 아르기닌, 시스테인, 또는 리신이며, Xaa18은 알라닌 또는 아르기닌이고, Xaa19는 알라닌 또는 시스테인이며, Xaa21은 글루탐산 또는 아스파르트산이고, Xaa24는 글루타민 또는 아스파라긴이며, Xaa28은 시스테인 또는 아스파르트산이고, Xaa29는 시스테인, 히스티딘, 또는 글루타민이며, Xaa30은 시스테인 또는 히스티딘이고, Xaa31은 프롤린 또는 시스테인이며, Xaa40은 시스테인 또는 부존재함.In the general formula 3, Xaa1 is histidine or tyrosine, Xaa2 is alpha-methyl-glutamic acid or Aib, Xaa13 is alanine, tyrosine or cysteine, Xaa17 is arginine, cysteine, or lysine, and Xaa18 is alanine or arginine, Xaa19 is alanine or cysteine, Xaa21 is glutamic acid or aspartic acid, Xaa24 is glutamine or asparagine, Xaa28 is cysteine or aspartic acid, Xaa29 is cysteine, histidine, or glutamine, Xaa30 is cysteine or histidine, Xaa31 is proline Or cysteine, and Xaa40 is cysteine or absent.
  10. 제1항에 있어서, 상기 R1은 시스테인, GKKNDWKHNIT (서열번호 106), CSSGQPPPS (서열번호 109), GPSSGAPPPS (서열번호 110), GPSSGAPPPSC (서열번호 111), PSSGAPPPS (서열번호 112), PSSGAPPPSG (서열번호 113), PSSGAPPPSHG (서열번호 114), PSSGAPPPSS (서열번호 115), PSSGQPPPS (서열번호 116), 또는 PSSGQPPPSC (서열번호 117)이거나, 부존재하는, 약학적 조성물.The method of claim 1, wherein R1 is cysteine, GKKNDWKHNIT (SEQ ID NO: 106), CSSGQPPPS (SEQ ID NO: 109), GPSSGAPPPS (SEQ ID NO: 110), GPSSGAPPPSC (SEQ ID NO: 111), PSSGAPPPS (SEQ ID NO: 112), PSSGAPPPSG (SEQ ID NO: 113), PSSGAPPPSHG (SEQ ID NO: 114), PSSGAPPPSS (SEQ ID NO: 115), PSSGQPPPS (SEQ ID NO: 116), or PSSGQPPPSC (SEQ ID NO: 117), or absent, pharmaceutical composition.
  11. 제1항에 있어서, 상기 펩타이드는 서열번호 1 내지 102로 이루어진 군으로부터 선택된 아미노산 서열을 포함하는, 약학적 조성물. The pharmaceutical composition of claim 1, wherein the peptide comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 1 to 102.
  12. 제1항 내지 제9항 중 어느 한 항에 있어서, 상기 일반식에서 N-말단으로부터 16번 아미노산과 20번 아미노산은 서로 고리를 형성하는, 약학적 조성물. The pharmaceutical composition according to any one of claims 1 to 9, wherein amino acids 16 and 20 form a ring with each other from the N-terminus in the general formula.
  13. 제1항 내지 제11항 중 어느 한 항에 있어서, 상기 펩타이드는 이의 C-말단이 아미드화된, 약학적 조성물. The pharmaceutical composition according to any one of claims 1 to 11, wherein the peptide is amidated at its C-terminus.
  14. 제1항 내지 제11항 중 어느 한 항에 있어서, 상기 약학적 조성물은 LDL 흡수 증가, 3-Hydroxy-3-methylglutaryl-CoA reductase (HMGCR) 활성 억제, 및 지방산 분해 촉진 효과 중 어느 하나 이상의 효과를 갖는 것인, 약학적 조성물.The method of any one of claims 1 to 11, wherein the pharmaceutical composition exhibits any one or more of an effect of increasing LDL absorption, inhibiting 3-Hydroxy-3-methylglutaryl-CoA reductase (HMGCR) activity, and promoting fatty acid degradation. Having, a pharmaceutical composition.
  15. 하기 화학식 1로 표시되는 결합체를 포함하는 고지혈증의 예방 또는 치료를 위한 약학적 조성물로서,As a pharmaceutical composition for the prevention or treatment of hyperlipidemia comprising a conjugate represented by the following formula (1),
    [화학식 1][Formula 1]
    X - La - FX-La-F
    여기서,here,
    X는, 상기 글루카곤 수용체, GLP-1 수용체, 및 GIP 수용체에 대해 활성을 갖는, 펩타이드이고,X is a peptide having activity against the glucagon receptor, GLP-1 receptor, and GIP receptor,
    L은, 폴리에틸렌 글리콜이며,L is polyethylene glycol,
    a는, 0 또는 자연수이며, 다만 a가 2 이상일 때, 각각의 L은 서로 독립적이고,a is 0 or a natural number, provided that when a is 2 or more, each L is independent of each other,
    F는, 면역글로불린 Fc 영역이며;F is an immunoglobulin Fc region;
    상기 펩타이드는 하기 일반식 1로 표시되는 아미노산 서열을 포함하는 펩타이드임:The peptide is a peptide comprising an amino acid sequence represented by the following general formula 1:
    Xaa1-Xaa2-Xaa3-Gly-Thr-Phe-Xaa7-Ser-Asp-Xaa10-Ser-Xaa12-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp-Leu-Xaa27-Xaa28-Xaa29-Xaa30-R1 (일반식 1, 서열번호 103)Xaa1-Xaa2-Xaa3-Gly-Thr-Phe-Xaa7-Ser-Asp-Xaa10-Ser-Xaa12-Xaa13-Xaa14-Xaa15-Xaa16-Xaa17-Xaa18-Xaa19-Xaa20-Xaa21-Phe-Xaa23-Xaa24-Trp- Leu-Xaa27-Xaa28-Xaa29-Xaa30-R1 (general formula 1, SEQ ID NO: 103)
    상기 일반식 1에서,In the general formula 1,
    Xaa1은 히스티딘 (His, H), 4-이미다조아세틸 (CA), 또는 티로신 (Tyr, Y)이고, Xaa1 is histidine (His, H), 4-imidazoacetyl (CA), or tyrosine (Tyr, Y),
    Xaa2는 글리신 (Gly, G), 알파-메틸-글루탐산, 또는 Aib (aminoisobutyric acid)이며,Xaa2 is glycine (Gly, G), alpha-methyl-glutamic acid, or Aib (aminoisobutyric acid),
    Xaa3은 글루탐산 (Glu, E) 또는 글루타민 (Gln, Q)이고, Xaa3 is glutamic acid (Glu, E) or glutamine (Gln, Q),
    Xaa7은 트레오닌 (Thr, T) 또는 이소류신 (Ile, I)이며,Xaa7 is threonine (Thr, T) or isoleucine (Ile, I),
    Xaa10은 류신 (Leu, L), 티로신 (Tyr, Y), 리신 (Lys, K), 시스테인 (Cys, C), 또는 발린 (Val, V)이고, Xaa10 is leucine (Leu, L), tyrosine (Tyr, Y), lysine (Lys, K), cysteine (Cys, C), or valine (Val, V),
    Xaa12는 리신 (Lys, K), 세린 (Ser, S), 또는 이소류신 (Ile, I)이며, Xaa12 is lysine (Lys, K), serine (Ser, S), or isoleucine (Ile, I),
    Xaa13은 글루타민 (Gln, Q), 티로신 (Tyr, Y), 알라닌 (Ala, A), 또는 시스테인 (Cys, C)이고, Xaa13 is glutamine (Gln, Q), tyrosine (Tyr, Y), alanine (Ala, A), or cysteine (Cys, C),
    Xaa14는 류신 (Leu, L), 메티오닌 (Met, M), 또는 티로신 (Tyr, Y)이며,Xaa14 is leucine (Leu, L), methionine (Met, M), or tyrosine (Tyr, Y),
    Xaa15는 시스테인 (Cys, C), 아스파르트산 (Asp, D), 글루탐산 (Glu, E), 또는 류신 (Leu, L)이며,Xaa15 is cysteine (Cys, C), aspartic acid (Asp, D), glutamic acid (Glu, E), or leucine (Leu, L),
    Xaa16은 글리신 (Gly, G), 글루탐산 (Glu, E), 또는 세린 (Ser, S)이고, Xaa16 is glycine (Gly, G), glutamic acid (Glu, E), or serine (Ser, S),
    Xaa17은 글루타민 (Gln, Q), 아르기닌 (Arg, R), 이소류신 (Ile, I), 글루탐산 (Glu, E), 시스테인 (Cys, C), 또는 리신 (Lys, K)이며,Xaa17 is glutamine (Gln, Q), arginine (Arg, R), isoleucine (Ile, I), glutamic acid (Glu, E), cysteine (Cys, C), or lysine (Lys, K),
    Xaa18은 알라닌 (Ala, A), 글루타민 (Gln, Q), 아르기닌 (Arg, R), 또는 히스티딘 (His, H)이고,Xaa18 is alanine (Ala, A), glutamine (Gln, Q), arginine (Arg, R), or histidine (His, H),
    Xaa19는 알라닌 (Ala, A), 글루타민 (Gln, Q), 시스테인 (Cys, C), 또는 발린 (Val, V)이며,Xaa19 is alanine (Ala, A), glutamine (Gln, Q), cysteine (Cys, C), or valine (Val, V),
    Xaa20은 리신 (Lys, K), 글루타민 (Gln, Q), 또는 아르기닌 (Arg, R)이고,Xaa20 is lysine (Lys, K), glutamine (Gln, Q), or arginine (Arg, R),
    Xaa21은 글루탐산 (Glu, E), 글루타민 (Gln, Q), 류신 (Leu, L), 시스테인 (Cys, C), 또는 아스파르트산 (Asp, D)이며,Xaa21 is glutamic acid (Glu, E), glutamine (Gln, Q), leucine (Leu, L), cysteine (Cys, C), or aspartic acid (Asp, D),
    Xaa23은 이소류신 (Ile, I) 또는 발린 (Val, V)이고,Xaa23 is isoleucine (Ile, I) or valine (Val, V),
    Xaa24는 알라닌 (Ala, A), 글루타민 (Gln, Q), 시스테인 (Cys, C), 아스파라긴 (Asn, N), 아스파르트산 (Asp, D), 또는 글루탐산 (Glu, E)이며,Xaa24 is alanine (Ala, A), glutamine (Gln, Q), cysteine (Cys, C), asparagine (Asn, N), aspartic acid (Asp, D), or glutamic acid (Glu, E),
    Xaa27은 발린 (Val, V), 류신 (Leu, L), 리신 (Lys, K), 또는 메티오닌 (Met, M)이고,Xaa27 is valine (Val, V), leucine (Leu, L), lysine (Lys, K), or methionine (Met, M),
    Xaa28은 시스테인 (Cys, C), 리신 (Lys, K), 알라닌 (Ala, A), 아스파라긴 (Asn, N), 또는 아스파르트산 (Asp, D)이며,Xaa28 is cysteine (Cys, C), lysine (Lys, K), alanine (Ala, A), asparagine (Asn, N), or aspartic acid (Asp, D),
    Xaa29는 시스테인 (Cys, C), 글리신 (Gly, G), 글루타민 (Gln, Q), 트레오닌 (Thr, T), 글루탐산 (Glu, E), 또는 히스티딘 (His, H)이고,Xaa29 is cysteine (Cys, C), glycine (Gly, G), glutamine (Gln, Q), threonine (Thr, T), glutamic acid (Glu, E), or histidine (His, H),
    Xaa30은 시스테인 (Cys, C), 글리신 (Gly, G), 리신 (Lys, K), 또는 히스티딘 (His, H)이거나, 부존재하며,Xaa30 is cysteine (Cys, C), glycine (Gly, G), lysine (Lys, K), or histidine (His, H) or absent,
    R1은 시스테인 (Cys, C), GKKNDWKHNIT (서열번호 106), m-SSGAPPPS-n (서열번호 107), 또는 m-SSGQPPPS-n (서열번호 108)이거나, 부존재하며, R1 is cysteine (Cys, C), GKKNDWKHNIT (SEQ ID NO: 106), m-SSGAPPPS-n (SEQ ID NO: 107), or m-SSGQPPPS-n (SEQ ID NO: 108), or absent,
    여기서, here,
    m은 -Cys-, -Pro-, 또는 -Gly-Pro-이고, m is -Cys-, -Pro-, or -Gly-Pro-,
    n은 -Cys-, -Gly-, -Ser-, 또는 -His-Gly-이거나, 부존재함. n is -Cys-, -Gly-, -Ser-, or -His-Gly- or absent.
  16. 제15항에 있어서, 상기 면역글로불린 Fc 영역은 IgG Fc 영역인, 약학적 조성물. The pharmaceutical composition of claim 15, wherein the immunoglobulin Fc region is an IgG Fc region.
  17. 제15항에 있어서, 상기 약학적 조성물은 LDL 흡수 증가, 3-Hydroxy-3-methylglutaryl-CoA reductase (HMGCR) 활성 억제, 및 지방산 분해 촉진 효과 중 어느 하나 이상의 효과를 갖는 것인, 약학적 조성물.The pharmaceutical composition of claim 15, wherein the pharmaceutical composition has one or more of an effect of increasing LDL absorption, inhibiting 3-Hydroxy-3-methylglutaryl-CoA reductase (HMGCR) activity, and promoting fatty acid degradation.
PCT/KR2020/005233 2019-04-19 2020-04-20 Preventive or therapeutic pharmaceutical composition for hyperlipidemia comprising triple agonist acting on all of glucagon, glp-1 and gip receptors, or conjugate thereof, and preventive or therapeutic method WO2020214012A1 (en)

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