GLP-1 / GIP dual agonist

Abstract

The present invention relates to long-acting glucagon-like peptide-1 and human glucose-dependent insulinotropic polypeptide (GIP) dual agonist polypeptides that may be useful in the treatment of type 2 diabetes (T2D), diabetes associated with obesity, obesity, and hyperlipidemia.

Description

[Technical Field]

[0001] The present invention relates to a long-acting glucagon-like peptide-1 (GLP-1) and human glucose-dependent insulinotropic polypeptide / gastrointestinal peptide (GIP) dual agonist polypeptide that may be useful in the treatment of type 2 diabetes (T2D), diabetes with obesity, obesity, and hyperlipidemia. [Background technology]

[0002] Treatment of type 2 diabetes (T2DM) with glucagon-like peptide-1 receptor agonists (GLP-1RAs) results in improved glycemic control, weight loss, and improvement of several cardiovascular risk factors. These benefits are mediated by the glucagon-like peptide-1 receptor (GLP-1R), a member of the class B family of G protein-coupled receptors, expressed in pancreatic beta cells, various cell types of the gastrointestinal tract, and neurons in both the central (CNS) and peripheral nervous systems. Activation of GLP-1R signaling by GLP-1RAs improves glucose homeostasis by enhancing glucose-stimulated insulin secretion, slowing gastric emptying, and decreasing plasma glucagon levels, and reduces weight by activating the anorexia pathway in the brain. Due to the glucose dependence of beta cell activation, GLP-1RAs are not associated with an increased risk of hypoglycemia. The broad metabolic benefits of GLP-1RAs have established this class in the T2DM treatment paradigm; however, many patients fail to reach their HbA1c / glycemic targets, and the weight loss achieved with these drugs, therefore, requires higher doses, which also increases the risk of adverse events, remaining far lower than what can be achieved with bariatric surgery, the most powerful clinical intervention for obesity. Thus, there is a great opportunity to improve the existing GLP-1RA class.

[0003] One novel approach is to combine basic GLP-1RA therapy with pharmacological strategies that target additional pathways involved in nutrition and energy metabolism, such as the glucose-dependent insulinotropic polypeptide (GIP) pathway. GIP is an incretin secreted from K cells in the upper small intestine and duodenum in response to food. Under normal physiological conditions, postprandial GIP levels are approximately four times higher than GLP-1 levels. GIP is responsible for the majority of the insulinopro-incretin effect in humans and has important additional functions that differ from GLP-1. Unlike GLP-1, GIP is both glucagon-secreting and insulin-secreting in a glucose-dependent manner, stimulating glucagon secretion in a dose-dependent manner under hypoglycemic conditions and insulin secretion under hyperglycemic conditions, with the released glucagon promoting insulin secretion. Both the GIP receptor (GIPR) and GLP-1R are present in beta cells, but GIPR expression is distributed differently in extrapancreatic tissues because GIPR is abundant in adipose tissue and found in many non-overlapping regions of the CNS. GIP is involved in adipose tissue carbohydrate and lipid metabolism through its actions of regulating glucose uptake, lipolysis, and lipoprotein lipase activity. These findings suggest that pharmacological activation of GIPR may have therapeutic benefits in peripheral energy metabolism. Recently, a monomolecule multifunctional peptide combining GLP-1RA activity and GIP activity has been proposed as a novel therapeutic agent for blood glucose and weight management.

[0004] U.S. Patent No. 9474780 discloses a dual GLP-1 and GIP receptor agonist containing tilzepatide.

[0005] [ka] Chilzepatide is currently undergoing Phase III clinical trials for T2DM and obesity.

[0006] WIPO Publication Nos. WO2017 / 74714A1, WO2020 / 23386A1, WO2020 / 023388A1, WO2015 / 067715A2, WO2016 / 111971A1 and WO2013 / 164483A1 disclose GLP-1 R and GIP R dual agonist compounds. SUMMARY OF THE INVENTION

[0007] The present invention provides a polypeptide comprising the following amino acid sequence or a pharmaceutically acceptable salt thereof, Y-X1-E-G-T-F-T-S-D-Y-S-I-X2-L-Xaa15-K-I-A-Xaa19-X3-Xaa21-F-V-Xaa24-W-L-X4-A-G-G-P-S-S-G-A-P-P-P-S-X5-X6-X7-X8-X9-X10-X11 (SEQ ID NO: 1) wherein X1 is Aib, (L)-norvaline or (D)-norvaline, X2 is selected from Aib, Leu, (D)-Leu, Val, (D)-Val, Ile, (D)-Ile, and the L or D isomers of the amino acids of the following formula,

Chemical formula

Chemical formula

[0008] Abbreviation A1: 2-aminoisobutyric acid DIPEA: N,N'-di-isopropylethylamine HOBt: 1-hydroxybenzotriazole DIPC: N,N'-di-isopropylcarbodiimide THF: Tetrahydrofuran DCM: Dichloromethane [Modes for carrying out the invention]

[0009] The “pharmaceutically acceptable salt” according to the present invention includes an acid addition salt formed with either an organic or inorganic acid. Suitable pharmaceutically acceptable salts of the compounds of the present invention include acid addition salts that may be, for example, salts of inorganic acids such as hydrochloric acid, hydrobromic acid, or phosphoric acid, or salts of organic acids such as acetic acid, fumaric acid, methanesulfonic acid, benzoic acid, citric acid, lactic acid, fumaric acid, succinic acid, adipic acid, pimelic acid, suberic acid, azelaic acid, malic acid, tartaric acid, glutamic acid, or amino acids such as aspartic acid. The pharmaceutically acceptable acid addition salts of the compounds include, for example, salts formed by the addition of one or more acid equivalents such as monohydrochloride or dihydrochloride. The salts can be prepared by any process at the responsibility of those skilled in the art. (See Berge et al., J. Pharm. Sci. 1977, 66, 1-19; and Handbook of Pharmaceutical Salts, Properties, and Use; Stahl and Wermuth, Ed.; Wiley-VCH and VHCA: Zurich, Switzerland, 2002.) As used herein, the term “alkyl” refers to a saturated hydrocarbon chain radical whose backbone consists only of carbon and hydrogen atoms, which, unless otherwise defined, has 1 to 6 carbon atoms, including both, and is either linear or branched, attached to other molecules by single bonds. Suitable non-limiting examples of alkyl groups include, for example, methyl, ethyl, n-propyl, 1-methylethyl (isopropyl), n-pentyl, and n-hexyl.

[0010] As used herein, the term “haloalkyl” means any “alkyl” having one or more hydrogen atoms substituted with a halogen atom, the halogen atom may be selected from fluorine, chlorine, bromine, or iodine.

[0011] "C 2-5 The numbers in phrases like "C" indicate the number of carbon atoms in the chain. For example, "C 2-5 The term "alkyl" refers to an alkyl chain that has 2 to 5 carbon atoms.

[0012] As used herein, the term “alkenyl” refers to a hydrocarbon chain containing at least one carbon-carbon double bond, which may have an (E) or (Z) configuration. Unless otherwise specified, an alkenyl group may contain 2 to 8 carbon atoms. Unless otherwise stated or enumerated, all alkenyl groups described herein may form part of a linear or branched chain. Suitable non-limiting examples of alkenyl groups include, for example, ethylene, 2-propenyl(allyl), 2-methyl-2-propenyl, and 2-butenyl.

[0013] The term "alkynyl" refers to a hydrocarbon chain having at least one carbon-carbon triple bond. Unless otherwise specified, an alkynyl group may contain 2 to 8 carbon atoms. Unless otherwise stated or enumerated, all alkynyl groups described or claimed herein may form part of a linear or branched chain. Non-limiting examples of alkynyl groups include 2-propynyl, 3-butynyl, and propargyl.

[0014] As used herein, the term “cycloalkyl” refers to a non-aromatic monocyclic ring system of 3 to 7 carbon atoms, unless otherwise specified. Cycloalkyl rings include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and cycloheptyl. The term "cycloalkenyl" refers to a non-aromatic monocyclic 5- to 7-membered cycloalkyl ring system having at least one carbon-carbon double bond. Non-restrictive examples of cycloalkenylmethyl groups include cyclopentenylmethyl and cyclohexenylmethyl.

[0015] As used herein, the term “effective dose” or “effective amount” refers to the amount of a compound that is sufficient to a subject in a single or multiple doses to cure, alleviate, reduce, or partially address the clinical symptoms of a given disease or condition and its complications beyond those expected in the absence of such treatment. Thus, the result may be a reduction and / or alleviation of the signs, symptoms, or causes of the disease, or any other desirable change in the biological system. It is understood that the “therapeutic effective dose” may vary from subject to subject, depending on age, weight, the subject’s overall condition, the condition being treated, the severity of the condition being treated, and the judgment of the prescribing physician.

[0016] As used herein, the amino acid "norvaline" may also be represented by its structure. [ka] It may also be defined as "2-aminopentanoic acid" by its chemical name. The terms (L)-norvaline and (D)-norvaline refer to the "L" and "D" isomers of norvaline, respectively.

[0017] As used herein, the amino acid "norleucine" may also be represented by its structure. [ka] It may also be defined as "2-aminohexanoic acid" by its chemical name. The terms (L)-norleucine and (D)-norleucine refer to the "L" and "D" isomers of norleucine, respectively.

[0018] As used herein, the amino acid "homoalanine" may also be represented by its structure. [ka] It may also be defined as "2-aminobutyric acid" by its chemical name. The terms (L)-homoalanine and (D)-homoalanine refer to the "L" and "D" isomers of homoalanine, respectively.

[0019] The present invention provides a stable, long-acting GLP-1 / GIP agonist polypeptide that may be useful in the treatment of type 2 diabetes (T2D), diabetes mellitus with obesity, obesity, and hyperlipidemia. The polypeptide of the present invention is long-acting, which may eliminate the need for frequent administration to patients who require it.

[0020] Accordingly, in one embodiment, the present invention provides a polypeptide comprising the following amino acid sequence or a pharmaceutically acceptable salt thereof, Y-X1-EGTFTSDYSI-X2-L-Xaa15-KIA-Xaa19-X3-Xaa21-FV-Xaa24-WL-X4-AGGPSSGAPPPS-X5-X6-X7-X8-X9-X10-X11 (Sequence number 1) In the formula, X1 is Aib, (L)-norvaline, or (D)-norvaline. X2 is selected from Aib, Leu, (D)-Leu, Val, (D)-Val, Ile, (D)-Ile, and the L or D isomers of the following amino acids: [ka] During the ceremony, " [ka] " represents the attachment point to Leu, and R is C 2-5 Alkyl, C 3-7 Cycloalkyl, C 3-7 Cyclolalkyl-C 1-3 Alkyl-, C 3-5 Alkenil, C 3-5 Alkinyl, C 5-7 Cycloalkenyl-CH2-, and C 1-3 Selected from haloalkyl-, or R is C along with the carbon it is attached to. 3-6 Forming a cycloalkyl ring, X3 is either Gln or Lys, and in the formula, if X3 is Lys, the amino(ε-amino) group in the side chain of Lys is acylated at the following part: [ka] In the formula, U is -C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-}, where} is the attachment point to the group W. W is -C(O)-NH-(CH2) p Selected from the group consisting of -NH-], -C(O)-C(CH3)2-NH-], and -C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-], where p is 3 or 4, and where ] is the attachment point to group Y. Y is -C(O)-(CH2)2-CH(COOH)NH--, where -- is the attachment point to group Z. Z is -C(O)-(CH2). n -COOH or -C(O)-(CH2) n -CH3, where n is an integer between 14 and 20. X4 is Leu, Ile, or Glu. X5 is either absent or is Arg or Lys. If X5 is Lys, the amino(ε-amino) side chain of Lys is acylated at the following point: [ka] In the formula, U' is -C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-}, where} is the attachment point to the group W'. W' is -C(O)-NH-(CH2) q Selected from the group consisting of -NH-], -C(O)-C(CH3)2-NH-], and -C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-], where p is 3 or 4, and where ] is the attachment point to the group Y', Y' is -C(O)-(CH2)2-CH(COOH)NH--, where -- is the attachment point to group Z'. Z' is -C(O)-(CH2). m -COOH or -C(O)-(CH2) m -CH3, where m is an integer between 14 and 20. X6 does not exist, or it is Lys. X7 does not exist, or it is Lys. X8 does not exist, or it is Lys. X9 does not exist, or it is Lys. X10 does not exist, or it is Lys. X11 does not exist, or it is Lys. Xaa15 is either Asp or Glu. Xaa19 is either Gln or Ala. Xaa21 is either Ala or Glu. Xaa24 is either Gln or Asn. In the formula, the acid group of the C-terminal amino acid is either a free carboxylic acid group or is amidated as a C-terminal primary amide, and at least one of X3 and X5 is Lys, However, if X1 is Aib, then X2 is not Aib.

[0021] In one embodiment of the present invention, X1 is Aib.

[0022] In another embodiment of the present invention, X2 is Aib.

[0023] In another embodiment of the present invention, X4 is Ile.

[0024] In another embodiment of the present invention, X1 is (L)-norvaline.

[0025] In another embodiment of the present invention, X2 is Leu.

[0026] In another embodiment of the present invention, X2 is Ile.

[0027] In another embodiment of the present invention, X2 is selected from the L or D isomers of the following amino acid formula: [ka] During the ceremony, " [ka] " represents the attachment point to Leu, and R is C 2-5 Alkyl, C 3-7 Cycloalkyl, C 3-7 Cyclolalkyl-C 1-3 Alkyl-, C 3-5 Alkenil, C 3-5 Alkinyl, C 5-7 Cycloalkenyl-CH2-, and C 1-3 Selected from haloalkyl-, or R is C along with the carbon it is attached to. 3-6 It forms a cycloalkyl ring.

[0028] In another embodiment of the present invention, X2 is an amino acid of the following formula: [ka] It exists in an "L" configuration.

[0029] In another embodiment of the present invention, X2 is an amino acid of the following formula: [ka] It exists in a "D" configuration.

[0030] In another embodiment of the present invention, X2 is an amino acid of the following formula: [ka] In the formula, R is C 2-5 It is alkyl. In another embodiment, R is selected from ethyl, n-propyl, isopropyl, and n-butyl.

[0031] In yet another embodiment of the present invention, X2 is an amino acid of the following formula: [ka] In the formula, R is n-propyl, and therefore forms X2 as norvaline. In another embodiment, X2 is (L)-norvaline. In another embodiment, X2 is (D)-norvaline.

[0032] In yet another embodiment of the present invention, X2 is an amino acid of the following formula: [ka] In the formula, R is n-butyl, and therefore forms X2 as norleucine. In another embodiment, X2 is (L)-norleucine. In another embodiment, X2 is (D)-norleucine.

[0033] In yet another embodiment of the present invention, X2 is an amino acid of the following formula: [ka] In the formula, R is ethyl, and thus forms X2 as homoalanine. In another embodiment, X2 is (L)-homoalanine. In another embodiment, X2 is (D)-homoalanine.

[0034] In another embodiment of the present invention, X2 is an amino acid of the following formula: [ka] In the formula, R is C 3-7 Cyclolalkyl-C 1-3 It is alkyl-. In another embodiment, R is selected from cyclopropylmethyl-, cyclopentylmethyl-, and cyclohexylmethyl-.

[0035] In another embodiment of the present invention, X2 is an amino acid of the following formula: [ka]

[0036] In another embodiment of the present invention, X2 is an amino acid of the following formula: [ka]

[0037] In another embodiment of the present invention, X2 is an amino acid of the following formula: [ka]

[0038] In another embodiment of the present invention, X1 is Aib, and X2 is the L-isomer of the amino acid of the following formula: [ka] In the formula, R is n-propyl, that is, X2 is (L)-norvaline.

[0039] In another embodiment of the present invention, X1 is Aib, and X2 is the L-isomer of the amino acid of the following formula: [ka] In the formula, R is ethyl, and therefore X2 is (L)-homoalanine.

[0040] In another embodiment of the present invention, X1 is Aib, and X2 is the L-isomer of the amino acid of the following formula: [ka] In the formula, R is n-butyl, and therefore X2 is (L)-norleucine.

[0041] In another embodiment of the present invention, X1 is Aib and X2 is Leu.

[0042] In another embodiment of the present invention, X1 is Aib and X2 is Ile.

[0043] In another embodiment of the present invention, X1 is (L)-norvaline and X2 is Aib.

[0044] In another embodiment of the present invention, X1 is (L)-norvaline, and X2 is the L-isomer of the amino acid of the following formula: [ka] In the formula, R is n-propyl, that is, X2 is (L)-norvaline.

[0045] In one embodiment of the present invention, X1 is Aib or (L)-norvaline, and X2 is Aib, Leu, Ile, or an L-isomer of an amino acid with the following formula: [ka] In the formula, R is n-propyl, and neither X1 nor X2 is Aib. In one embodiment of the present invention, X1 is Aib or (L)-norvaline, and X2 is Aib, Leu, Ile, or an L-isomer of an amino acid with the following formula: [ka] In the formula, R is n-butyl, and neither X1 nor X2 is Aib.

[0046] In one embodiment of the present invention, X1 is Aib or (L)-norvaline, and X2 is Aib, Leu, Ile, or an L-isomer of an amino acid with the following formula: [ka] In the formula, R is ethyl, and neither X1 nor X2 is Aib.

[0047] In one embodiment of the present invention, X1 is Aib or (L)-norvaline, and X2 is Aib, Leu, Ile, or an L-isomer of an amino acid with the following formula: [ka] In the formula, R is n-propyl, X4 is Ile, and neither X1 nor X2 is Aib.

[0048] In one embodiment of the present invention, X1 is Aib or (L)-norvaline, and X2 is Aib, Leu, Ile, or an L-isomer of an amino acid with the following formula: [ka] In the formula, R is n-propyl, X5 is Arg, and neither X1 nor X2 is Aib.

[0049] In one embodiment of the present invention, X3 is Lys, and the amino(ε-amino) group of the Lys side chain is acylated at the following point.

[0050] TIFF0007830441000034.tif1665 In another embodiment of the present invention, W is -C(O)-C(CH3)2-NH-].

[0051] In another embodiment of the present invention, W is -C(O)-NH-(CH2) p -NH-], where p is 3 or 4.

[0052] In another embodiment of the present invention, W is -C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-].

[0053] In another embodiment of the present invention, Z is -C(O)-(CH2) n It is -COOH, and n is 16, 17, 18, 19, or 20. In a preferred embodiment, n is 16, 18, or 20. In yet another preferred embodiment, n is 18 or 20.

[0054] In another preferred embodiment of the present invention, Z is -C(O)-(CH2) n It is -COOH, and n is 16 or 18. In yet another preferred embodiment, n is 18.

[0055] In another embodiment of the present invention, W is -C(O)-C(CH3)2-NH- and Z is -C(O)-(CH2) n It is -COOH, and in the formula, n is 18.

[0056] In another embodiment of the present invention, W is -C(O)-C(CH3)2-NH- and Z is -C(O)-(CH2) n It is -COOH, and in the formula, n is 16.

[0057] In another embodiment of the present invention, W is -C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-] and Z is -C(O)-(CH2) n It is -COOH, and in the formula, n is 16.

[0058] In another embodiment of the present invention, W is -C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-] and Z is -C(O)-(CH2) n It is -COOH, and in the formula, n is 18.

[0059] In one embodiment of the present invention, X3 is Gln and X5 is Lys, and in the formula, the amino(ε-amino) group of the Lys side chain is acylated at the following part.

[0060] [ka]

[0061] In one embodiment of the present invention, X1 is Aib or (L)-norvaline, and X2 is Aib, Leu, Ile, or an L-isomer of an amino acid with the following formula: [ka] In the formula, R is n-propyl, X3 is Gln, X5 is Lys, and both X1 and X2 are not Aib. In the formula, the amino (amino) group of the Lys side chain is acylated at the following part. [ka]

[0062] In another embodiment of the present invention, W' is -C(O)-C(CH3)2-NH-].

[0063] In another embodiment of the present invention, W' is -C(O)-NH-(CH2) q -NH-], where p is 3 or 4.

[0064] In another embodiment of the present invention, W' is -C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-].

[0065] In another embodiment of the present invention, Z' is -C(O)-(CH2) m -COOH, and m is 16, 17, 18, 19, or 20. In a preferred embodiment, m is 16, 18, or 20. In another, more preferred embodiment, m is 18 or 20.

[0066] In another preferred embodiment of the present invention, Z' is -C(O)-(CH2) m -COOH, and m is 16 or 18. In another, more preferred embodiment, m is 18.

[0067] In another embodiment of the present invention, W' is -C(O)-C(CH3)2-NH- and Z' is -C(O)-(CH2) m It is -COOH, and in the formula, n is 18.

[0068] In another embodiment of the present invention, W' is -C(O)-C(CH3)2-NH- and Z' is -C(O)-(CH2) m It is -COOH, and in the formula, n is 16.

[0069] In another embodiment of the present invention, W' is -C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-] and Z' is -C(O)-(CH2) m It is -COOH, where m is 16.

[0070] In another embodiment of the present invention, W' is -C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-] and Z' is -C(O)-(CH2) m -COOH, where m is 18.

[0071] In one embodiment of the present invention, X1 is Aib, and X2 is the L-isomer of the amino acid given by the following formula: [ka] In the formula, R is n-propyl, Xaa15 is Glu, Xaa19 is Ala, X3 is Gln, Xaa21 is Glu, Xaa24 is Asn, X4 is Leu, and X5 is Lys. In the formula, the amino (ε-amino) side chain group of Lys is acylated at the following part: [ka] In the formula, W' is -C(O)-C(CH3)2-NH-, Z' is -C(O)-(CH2). m -COOH, where m is 18.

[0072] In another embodiment of the present invention, the C-terminal amino acid is amidated as a C-terminal primary amide.

[0073] In another embodiment of the present invention, X5, X6, X7, X8, X9, X10, and X11 are all absent.

[0074] In another embodiment, the present invention provides a polypeptide comprising the following amino acid sequence or a pharmaceutically acceptable salt thereof, Y-Aib-EGTFTSDYSI-X2-LDKIAQ-X3-AFVQWL-X4-AGGPSSGAPPPS-X5-X6-X7-X8-X9-X10-X11 (SEQ ID NO: 2), In the formula, X2 is Leu, Ile, (L)-norvaline, (L)-homoalanine, or (L)-norleucine. X4 is Ile, X5 does not exist, or it is Arg. X6 does not exist, or it is Lys. X7 does not exist, or it is Lys. X8 does not exist, or it is Lys. X9 does not exist, or it is Lys. X10 does not exist, or it is Lys. X11 does not exist, or it is Lys. X3 is Lys, and in the formula, the amino(ε-amino) group of the Lys side chain is acylated at the following part: [ka] In the formula, U is -C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-}, where} is the attachment point to the group W. W is -C(O)-NH-(CH2) p Selected from the group consisting of -NH-], -C(O)-C(CH3)2-NH-], and -C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-], where p is 3 or 4, and where ] is the attachment point to group Y. Y is -C(O)-(CH2)2-CH(COOH)NH--, where -- is the attachment point to group Z. Z is -C(O)-(CH2). n -COOH or -C(O)-(CH2) n The formula is -CH3, where n is an integer between 14 and 20, and where the acid group of the C-terminal amino acid is either a free carboxylic acid group or is amidated as a primary amide at the C-terminus.

[0075] In one embodiment of the present invention, X2 is (L)-norvaline.

[0076] In one embodiment of the present invention, X2 is (L)-homoalanine.

[0077] In one embodiment of the present invention, X2 is (L)-norleucine.

[0078] In another embodiment of the present invention, X2 is Leu.

[0079] In another embodiment of the present invention, X2 is Ile.

[0080] In another embodiment of the present invention, X2 is (L)-norvaline and X5 is Arg.

[0081] In another embodiment of the present invention, W is -C(O)-C(CH3)2-NH-].

[0082] In another embodiment of the present invention, W is -C(O)-NH-(CH2) 3-4 -NH-]

[0083] In another embodiment of the present invention, W is -C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-].

[0084] In another embodiment of the present invention, Z is -C(O)-(CH2) n It is -COOH, and n is 16, 17, 18, 19, or 20. In a preferred embodiment, n is 16, 18, or 20. In yet another preferred embodiment, n is 18 or 20.

[0085] In another preferred embodiment of the present invention, Z is -C(O)-(CH2) n It is -COOH, and n is 16 or 18. In yet another preferred embodiment, n is 18.

[0086] In another embodiment of the present invention, W is -C(O)-C(CH3)2-NH- and Z is -C(O)-(CH2) n It is -COOH, and in the formula, n is 18.

[0087] In another embodiment of the present invention, W is -C(O)-C(CH3)2-NH- and Z is -C(O)-(CH2) n It is -COOH, and in the formula, n is 16.

[0088] In another embodiment of the present invention, W is -C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-] and Z is -C(O)-(CH2) n It is -COOH, and in the formula, n is 16.

[0089] In another embodiment of the present invention, W is -C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-] and Z is -C(O)-(CH2) n It is -COOH, and in the formula, n is 18.

[0090] In another embodiment of the present invention, the C-terminal amino acid is amidated as a C-terminal primary amide.

[0091] In another embodiment of the present invention, X5, X6, X7, X8, X9, X10, and X11 are all absent.

[0092] In a preferred embodiment, X2 is (L)-norvaline, X4 is Ile, X5, X6, X7, X8, X9, X10 and X11 are absent, W is -C(O)-C(CH3)2-NH-, and Z is -C(O)-(CH2) n It is -COOH, and in the formula, n is 18.

[0093] In another embodiment, the present invention provides a polypeptide comprising the following amino acid sequence or a pharmaceutically acceptable salt thereof, Y-X1-EGTFTSDYSI-X2-LDKIAQ-X3-AFVQWL-X4-AGGPSSGAPPPS (Sequence ID 3) In the formula, X1 is Aib or (L)-norvaline, and X2 is Aib, Leu, Ile, (L)-norvaline, (L)-homoalanine, or (L)-norleucine. X4 is Ile, X3 is Lys, and in the formula, the amino(ε-amino) group of the Lys side chain is acylated at the following part: [ka] In the formula, U is -C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-}, where} is the attachment point to the group W. W is -C(O)-NH-(CH2) p Selected from the group consisting of -NH-], -C(O)-C(CH3)2-NH-], and -C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-], where p is 3 or 4, and where ] is the attachment point to group Y. Y is -C(O)-(CH2)2-CH(COOH)NH--, where -- is the attachment point to group Z. Z is -C(O)-(CH2). n -COOH or -C(O)-(CH2) n -CH3, where n is an integer between 14 and 20. In the formula, the acid group of the C-terminal amino acid is either a free carboxylic acid group or is amidated as a C-terminal primary amide. However, if X1 is Aib, then X2 is not Aib.

[0094] In one embodiment of the present invention, X1 is Aib.

[0095] In another embodiment of the present invention, X1 is (L)-norvaline.

[0096] In another embodiment of the present invention, X2 is Aib.

[0097] In another embodiment of the present invention, X2 is (L)-norvaline.

[0098] In another embodiment of the present invention, X2 is (L)-norleucine.

[0099] In another embodiment of the present invention, X2 is (L)-homoalanine.

[0100] In another embodiment of the present invention, X2 is Leu.

[0101] In another embodiment of the present invention, X2 is Ile.

[0102] In another embodiment of the present invention, X1 is Aib and X2 is (L)-norvaline.

[0103] In another embodiment of the present invention, X1 is Aib and X2 is (L)-norleucine.

[0104] In another embodiment of the present invention, X1 is Aib and X2 is (L)-homoalanine.

[0105] In another embodiment of the present invention, X1 is (L)-norvaline and X2 is Aib.

[0106] In another embodiment of the present invention, X1 is Aib and X2 is Leu.

[0107] In another embodiment of the present invention, X1 is Aib and X2 is Ile.

[0108] In another embodiment of the present invention, X1 is (L)-norvaline and X2 is (L)-norvaline.

[0109] In another embodiment of the present invention, W is -C(O)-C(CH3)2-NH-].

[0110] In another embodiment of the present invention, W is -C(O)-NH-(CH2) 3-4 -NH-]

[0111] In another embodiment of the present invention, W is -C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-].

[0112] In another embodiment of the present invention, Z is -C(O)-(CH2) nIt is -COOH, and n is 16, 17, 18, 19, or 20. In a preferred embodiment, n is 16, 18, or 20. In yet another preferred embodiment, n is 18 or 20.

[0113] In another preferred embodiment of the present invention, Z is -C(O)-(CH2) n It is -COOH, and n is 16 or 18. In yet another preferred embodiment, n is 18.

[0114] In another embodiment of the present invention, W is -C(O)-C(CH3)2-NH- and Z is -C(O)-(CH2) n It is -COOH, and in the formula, n is 18.

[0115] In another embodiment of the present invention, W is -C(O)-C(CH3)2-NH- and Z is -C(O)-(CH2) n It is -COOH, and in the formula, n is 16.

[0116] In another embodiment of the present invention, W is -C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-] and Z is -C(O)-(CH2) n It is -COOH, and in the formula, n is 16.

[0117] In another embodiment of the present invention, W is -C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-] and Z is -C(O)-(CH2) n It is -COOH, and in the formula, n is 18.

[0118] In another embodiment of the present invention, X1 is Aib, X2 is (L)-norvaline, and X3 is Lys, where the amino(ε-amino) side chain group of Lys is acylated at the following part: [ka] In the formula, W is -C(O)-C(CH3)2-NH- and Z is -C(O)-(CH2) n It is -COOH, and n is 18.

[0119] In another embodiment of the present invention, the C-terminal amino acid is amidated as a C-terminal primary amide.

[0120] In another embodiment, the present invention provides a polypeptide comprising an amino acid sequence selected from the group consisting of the following, or a pharmaceutically acceptable salt thereof. i.) Tyr Aib Glu Gly Thr Phe Thr Ser Asp Tyr Ser Ile L-Norvaline Leu Asp Lys Ile Ala Gln Lys Ala Phe Val Gln Trp Leu Ile Ala Gly Gly Pro Ser Ser Gly Ala Pro Pro Pro Ser-NH2 (SEQ ID NO: 04) ii.) Tyr L-Norvaline Glu Gly Thr Phe Thr Ser Asp Tyr Ser Ile Aib Leu Asp Lys Ile Ala Gln Lys Ala Phe Val Gln Trp Leu Ile Ala Gly Gly Pro Ser Ser Gly Ala Pro Pro Pro Ser-NH2 (SEQ ID NO: 05) iii.) Tyr Aib Glu Gly Thr Phe Thr Ser Asp Tyr Ser Ile Leu Leu Asp Lys Ile Ala Gln Lys Ala Phe Val Gln Trp Leu Ile Ala Gly Gly Pro Ser Ser Gly Ala Pro Pro Pro Ser-NH2 (SEQ ID NO: 06) iv.) Tyr L-norvaline Glu Gly Thr Phe Thr Ser Asp Tyr Ser Ile L-norvaline Leu Asp Lys Ile Ala Gln Lys Ala Phe Val Gln Trp Leu Ile Ala Gly Gly Pro Ser Ser Gly Ala Pro Pro Pro Ser-NH2 (Sequence ID 07) v.) Tyr Aib Glu Gly Thr Phe Thr Ser Asp Tyr Ser Ile L-Norvaline Leu Glu Lys Ile Ala Ala Gln Glu Phe Val Asn Trp Leu Leu Ala Gly Gly Pro Ser Ser Gly Ala Pro Pro Pro Ser Lys-NH2 (SEQ ID NO: 08)[[ID=!]] vi.) Tyr Aib Glu Gly Thr Phe Thr Ser Asp Tyr Ser Ile L-Norvaline Leu Asp Lys Ile Ala Gln Lys Ala Phe Val Gln Trp Leu Ile Ala Gly Gly Pro Ser Ser Gly Ala Pro Pro Pro Ser Arg (SEQ ID NO: 09) vii.) Tyr Aib Glu Gly Thr Phe Thr Ser Asp Tyr Ser Ile L-Homoalanine Leu Asp Lys Ile Ala Gln Lys Ala Phe Val Gln Trp Leu Ile Ala Gly Gly Pro Ser Ser Gly Ala Pro Pro Pro Ser-NH2 (SEQ ID NO: 10) viii.) Tyr Aib Glu Gly Thr Phe Thr Ser Asp Tyr Ser Ile L-Norleucine Leu Asp Lys Ile Ala Gln Lys Ala Phe Val Gln Trp Leu Ile Ala Gly Gly Pro Ser Ser Gly Ala Pro Pro Pro Ser-NH2 (SEQ ID NO: 11) ix.) Tyr Aib Glu Gly Thr Phe Thr Ser Asp Tyr Ser Ile Ile Leu Asp Lys Ile Ala Gln Lys Ala Phe Val Gln Trp Leu Ile Ala Gly Gly Pro Ser Ser Gly Ala Pro Pro Pro Ser-NH2 (SEQ ID NO: 12)

[0121] In another embodiment, the present invention provides polypeptides selected from representative compounds disclosed in Table 1 or pharmaceutically acceptable salts thereof.

[0122] In embodiments of the present invention, the bases U, W, Y and Z in the following parts, [ka] Or the bases U', W', Y', and Z' in the following section are: [ka] It has the meaning defined herein and should not be interpreted or confused with a single-letter code for an amino acid.

[0123] In embodiments of the present invention, the base {-UWYZ and / or {-U'-W'-Y'-Z' is selected from representative structures of parts A, B, C, and D disclosed in Table 2.

[0124] Polypeptide sequences referred to herein are represented by single-letter amino acid codes approved by IUPAC.

[0125] Unless otherwise specified, this specification is intended to cover both L and D isomers of amino acids in a sequence. However, in preferred embodiments, unless otherwise indicated, all amino acids are in the "L" form.

[0126] Tables 1 and 2 provide some representative compounds of the present invention.

[0127] [Table 1] TIFF0007830441000046.tif55167

[0128] [Table 2]

[0129] In another embodiment, the present invention provides a method for treating or preventing hyperglycemia, type 2 diabetes, impaired glucose tolerance, type 1 diabetes, obesity, hypertension, hyperlipidemia, syndrome X, dyslipidemia, cognitive impairment, atherosclerosis, myocardial infarction, coronary heart disease, stroke, inflammatory bowel syndrome, dyspepsia, alcoholism, and gastric ulcers in patients, comprising administering to patients in need thereof an effective amount of the polypeptide of the present invention or a pharmaceutically acceptable salt thereof.

[0130] In another embodiment, the present invention provides a method for treating type 2 diabetes in a patient, comprising administering an effective amount of the compound of the present invention or a pharmaceutically acceptable salt thereof to a patient requiring such treatment.

[0131] In another embodiment, the present invention provides a method for treating obesity in a patient, comprising administering an effective amount of the compound of the present invention or a pharmaceutically acceptable salt thereof to a patient requiring such treatment.

[0132] In another embodiment, the present invention provides a method for treating hyperlipidemia in a patient, comprising administering an effective amount of the compound of the present invention or a pharmaceutically acceptable salt thereof to a patient requiring such treatment.

[0133] In one embodiment, the present invention provides a pharmaceutical composition comprising a polypeptide of the present invention or a pharmaceutically acceptable salt thereof, comprising one or more pharmaceutically acceptable carriers, diluents, or excipients.

[0134] The compounds of the present invention are preferably formulated as pharmaceutical compositions administered via parenteral routes (e.g., subcutaneous, intravenous, intraperitoneal, intramuscular, or transdermal). Such pharmaceutical compositions and processes for preparing them are well known in the art. (See, for example, Remington: The Science and 50 Practices of Pharmacy (DB Troy, Editor, 21st Edition, Lippincott, Williams & Wilkins, 2006)).

[0135] In another embodiment, the polypeptide of the present invention or a pharmaceutically acceptable salt thereof is used as a pharmaceutical agent.

[0136] In another embodiment, the polypeptide of the present invention or a pharmaceutically acceptable salt thereof used for the treatment or prevention of a patient's disease, wherein the disease is arbitrarily selected from the group consisting of hyperglycemia, type 2 diabetes, impaired glucose tolerance, type 1 diabetes, obesity, hypertension, hyperlipidemia, syndrome X, dyslipidemia, cognitive impairment, atherosclerosis, myocardial infarction, coronary heart disease, stroke, inflammatory bowel syndrome, dyspepsia, alcoholism, and gastric ulcer.

[0137] In another embodiment, the polypeptide of the present invention or a pharmaceutically acceptable salt thereof may be provided simultaneously, separately, or sequentially in combination with an effective amount of one or more additional therapeutic agents.

[0138] In another embodiment, the pharmaceutical composition according to the present invention comprises a polypeptide of the present invention or a pharmaceutically acceptable salt thereof, to be used as a pharmaceutical agent.

[0139] In another embodiment, a pharmaceutical composition according to the present invention comprises a polypeptide of the present invention or a pharmaceutically acceptable salt thereof for use in the treatment or prevention of a patient's disease, wherein the aforementioned disease is arbitrarily selected from the group consisting of hyperglycemia, type 2 diabetes, impaired glucose tolerance, type 1 diabetes, obesity, hypertension, hyperlipidemia, syndrome X, dyslipidemia, cognitive impairment, atherosclerosis, myocardial infarction, coronary heart disease, stroke, inflammatory bowel syndrome, dyspepsia, alcoholism, and gastric ulcer.

[0140] In another embodiment, the pharmaceutical composition according to the present invention comprises a polypeptide of the present invention or a pharmaceutically acceptable salt thereof, provided simultaneously, separately, or sequentially in combination with an effective amount of one or more additional therapeutic agents.

[0141] The present invention may be accompanied by one or more embodiments. It should be understood that the following embodiments are illustrative of the invention and are not intended to limit the claims to any particular embodiment illustrated. It should also be understood that the embodiments defined herein may be used independently or in conjunction with any other definition or embodiment defined herein. Accordingly, the present invention intends all possible combinations and permutations of various independently described embodiments.

[0142] Other features of the present invention will become apparent from the following examples. In general, the present invention extends to any novel features or any novel combination of the features disclosed herein (including the appended claims and drawings). Accordingly, any features, integers, features, compounds, or chemical parts described in conjunction with a particular aspect, embodiment, or example of the present invention should be understood to apply to any other aspect, embodiment, or example described herein, insofar as they do not conflict.

[0143] Furthermore, unless otherwise stated, any feature disclosed herein may be replaced by an alternative feature that serves the same or similar purpose. [Examples]

[0144] Instruments and analytical methods: The instrument used for characterizing and analyzing the compounds of the present invention is HPLC (Waters e2695 Alliance; Detector Waters (2489 UV / Visible)).

[0145] Equipment: HPLC: Waters e2695 Alliance; Detector: Acquity-QDa.

[0146] The final compound of this disclosure was purified by a preparative HPLC procedure, as outlined below.

[0147] Fractional HPLC: WATERS 2555 Quaternary gradient module (maximum total flow rate: 300 mL / min, maximum pressure: 3000 psi) or Shimadzu LC-8A (maximum total flow rate: 150 mL, maximum pressure: 30 Mpa), column: phenyl, 10 μ, flow rate: 75 mL / min

[0148]

Table 3

[0149] The purity of the compounds of the present disclosure was analyzed by one of the RP-HPLC methods outlined below.

[0150] HPLC Method A: Column: Xbridge Peptide BEH C18 (4.6 mm × 250 mm, 3.5 u) Eluent: Mobile phase A: Buffer: Acetonitrile (900:100) Mobile phase B: Buffer: Acetonitrile (300:700) Buffer: Potassium dihydrogen phosphate in water adjusted to pH 3.0 ± 0.1 with orthophosphoric acid Flow rate: 0.8 mL / min Detection: UV detection at 210 nm Column temperature: 65 °C Sample tray temperature: 5 °C Run time: 90 minutes

[0151]

Table 4

[0152] HPLC Method B: Column: XSelect CSH C18 (4.6 mm × 150 mm, 2.5 μ) Eluent: Mobile phase A: Buffer: Acetonitrile (900:100) Mobile phase B: Buffer: Acetonitrile (300:700) Buffer: Potassium dihydrogen phosphate in water adjusted to pH 3.0 ± 0.1 with orthophosphoric acid Flow rate: 0.8 mL / min Detection: UV detection at 210 nm Column temperature: 65 °C Sample tray temperature: 5 °C Run time: 90 min

[0153]

Table 5

[0154] HPLC method C: Column: Xbridge Peptide BEH C18 (4.6 mm × 250 mm, 3.5 u) Eluent: Mobile phase A: Buffer: Acetonitrile (900:100) Mobile phase B: Buffer: Acetonitrile (300:700) Buffer: Potassium dihydrogen phosphate in water, pH adjusted to 3.0 ± 0.1 with orthophosphoric acid Flow rate: 1.0 mL / min Detection: UV detection at 210 nm Column temperature: 65 °C Sample tray temperature: 5 °C Run time: 60 min

[0155]

Table 6

[0156] HPLC method D: Column: XSelect CSH C18 (4.6 mm × 150 mm, 2.5 μ) Eluent: Mobile phase A: Buffer: Acetonitrile (900:100) Mobile phase B: Buffer: Acetonitrile (300:700) Buffer: Potassium dihydrogen phosphate in water, pH adjusted to 2.5 ± 0.1 with orthophosphoric acid and triethylamine added Flow rate: 0.5 mL / min Detection: UV detection at 214 nm Column temperature: 60 °C Sample tray temperature: 10 °C Execution time: 90 minutes

[0157] [Table 7]

[0158] Preparation method: Example 1: Preparation of 2-[2-[[2-[[(4S)-5-tert-butoxy-4-[(20-tert-butoxy-20-oxo-icosanoyl)amino]-5-oxo-pentanoyl]amino]-2-methyl-propanoyl]amino]ethoxy]ethoxy]acetic acid (partially A-di-tert-butyl ester) [ka] Partially A-di-tert-butyl esters were prepared using solid-phase synthesis with 2-chlorotrityl chloride resin. 2-[2-(2-Fmoc-aminoethoxy)ethoxy]acetic acid was attached to 2-chlorotrityl chloride resin in the presence of DIPEA to obtain 2-[2-(2-Fmoc-aminoethoxy)ethoxy]acetic acid-2-Cl-Trt-resin. The Fmoc protecting group was removed by selective deblockage of the amino group using piperidine, and subsequently bonded to Fmoc-Aib-OH in THF using DIPC and HOBt to obtain 2-[2-[2-[(2-Fmoc-amino-2-methyl-propanoyl)amino]ethoxy]ethoxy]acetic acid-2-Cl-Trt-resin. The Fmoc group was removed by selective deblocking using piperidine, and then the free amino group was bonded to Fmoc-Glu-OtBu using HOBt and DIPC to obtain 2-[2-[2-[[2-[[(4S)-4-Fmoc-amino-5-tert-butoxy-5-oxo-pentanoyl]amino]-2-methyl-propanoyl]amino]ethoxy]ethoxy]acetic acid-2-Cl-Trt resin. The Fmoc group of the obtained compound was selectively deblocked using piperidine, and then the free amino group was bonded to 20-(tert-butoxy)-20-oxoeicosanoic acid to obtain 2-[2-[2-[[2-[[(4S)-5-tert-butoxy-4-[(20-tert-butoxy-20-oxo-icosanoyl)amino]-5-oxo-pentanoyl]amino]-2-methyl-propanoyl]-amino]ethoxy]ethoxy]acetic acid-2-Cl-Trt-resin. Next, this intermediate was cleaved from the 2-Cl-Trt resin using trifluoroethanol:DCM (1:1) to obtain 2-[2-[2-[[2-[[(4S)-5-tert-butoxy-4-[(20-tert-butoxy-20-oxo-icosanoyl)amino]-5-oxo-pentanoyl]amino]-2-methyl-propanoyl]amino]ethoxy]ethoxy]acetic acid. (Partially A-di-tert-butyl ester) (LCMS=m / z:814.10(M+H) + ))

[0159] Example 2: Preparation of 2-[2-[[2-[[(4S)-5-tert-butoxy-4-[(18-tert-butoxy-18-oxo-octadecanoyl)amino]-5-oxo-pentanoyl]amino]-2-methyl-propanoyl]amino]ethoxy]ethoxy]acetic acid [ka] Partially B-di-tert-butyl ester The partial B-di-tert-butyl ester was prepared using solid-phase synthesis with 2-chlorotrityl chloride resin. 2-[2-(2-Fmoc-aminoethoxy)ethoxy]acetic acid was attached to 2-chlorotrityl chloride resin in the presence of DIPEA to obtain 2-[2-(2-Fmoc-aminoethoxy)ethoxy]acetic acid-2-Cl-Trt-resin. The Fmoc protecting group was removed by selective deblocking of the amino group using piperidine, followed by coupling with Fmoc-Aib-OH in THF using DIPC and HOBt to obtain 2-[2-[2-[(2-Fmoc-amino-2-methyl-propanoyl)amino]ethoxy]ethoxy]acetic acid-2-Cl-Trt-resin. The Fmoc group was removed by selective deblocking using piperidine, and the free amino group was coupled with Fmoc-Glu-OtBu using HOBt and DIPC to obtain 2-[2-[2-[[2-[[(4S)-4-Fmoc-amino-5-tert-butoxy-5-oxo-pentanoyl]amino]-2-methyl-propanoyl]amino]ethoxy]ethoxy]acetic acid-2-Cl-Trt-resin. The Fmoc group of the resulting compound was selectively deblocked using piperidine, and then the free amino group was coupled with octadecanedioic acid monotert-butyl ester to obtain 2-[2-[2-[[2-[[(4S)-5-tert-butoxy-4-[(18-tert-butoxy-18-oxo-octadecanoyl)amino]-5-oxo-pentanoyl]amino]-2-methyl-propanoyl]-amino]ethoxy]ethoxy]acetic acid-2-Cl-Trt-resin. The intermediate was then cleaved from the 2-Cl-Trt-resin using trifluoroethanol:DCM (1:1) to obtain 2-[2-[2-[[2-[[(4S)-5-tert-butoxy-4-[(18-tert-butoxy-18-oxo-octadecanoyl)amino]-5-oxo-pentanoyl]amino]-2-methyl-propanoyl]amino]ethoxy]ethoxy]acetic acid. (Partial B-di-tert-butyl ester) (LCMS = m / z: 786.39 (M+H + ))

[0160] Example 3: Preparation of 2-[2-[2-[[2-[2-[2-[[5-tert-butoxy-4-[(18-tert-butoxy-18-oxo-octadecanoyl)amino]-5-oxo-pentanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetic acid (partially C-di-tert-butyl ester) [ka] Partially C-di-tert-butyl esters were prepared using solid-phase synthesis with 2-chlorotrityl chloride resin. 2-[2-(2-Fmoc-aminoethoxy)ethoxy]acetic acid was attached to 2-chlorotrityl chloride resin in the presence of DIPEA to obtain 2-[2-(2-Fmoc-aminoethoxy)ethoxy]acetic acid-2-Cl-Trt-resin. The Fmoc protecting group was removed by selective deblocking of the amino group using piperidine, and subsequently bonded to 2-[2-(2-Fmoc-aminoethoxy)ethoxy]acetic acid in THF using DIPC and HOBt to obtain {(Fmoc-aminoethoxy)-ethoxy}acetyl-{(-aminoethoxy)-ethoxy}acetic acid-2-Cl-Trt-resin. The Fmoc group was removed by selective deblocking using piperidine, and the free amino group was bonded to Fmoc-Glu-OtBu using HOBt and DIPC to obtain Fmoc-Glu({(amino-ethoxy)-ethoxy}-acetyl-{(-amino-ethoxy)-ethoxy}-acetic acid-2-Cl-Trt-resin)-OtBu. The Fmoc group of the obtained compound was selectively deblocked using piperidine, and the free amino group was then bonded to octadecanediic acid monotert-butyl ester to obtain 2-[2-[2-[[2-[2-[2-[[5-tert-butoxy-4-[(18-tert-butoxy-18-oxo-octadecanoyl)amino]-5-oxo-pentanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetic acid-2-Cl-Trt-resin. Next, using trifluoroethanol:DCM (1:1), the intermediate is cleaved from the 2-Cl-Trt resin to obtain 2-[2-[2-[[2-[2-[2-[[5-tert-butoxy-4-[(18-tert-butoxy-18-oxo-octadecanoyl)amino]-5-oxo-pentanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetic acid (partially C-di-tert-butyl ester) (LCMS=m / z:846.10(M+H + )) was obtained.

[0161] Example 4: Preparation of 2-[2-[2-[[2-[2-[2-[[5-tert-butoxy-4-[(20-tert-butoxy-20-oxo-icosanoyl)amino]-5-oxo-pentanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetic acid (Partially D-di-tert-butyl ester) [ka] Partially D-di-tert-butyl esters were prepared using solid-phase synthesis with 2-chlorotrityl chloride resin, as schematically shown below. 2-[2-(2-Fmoc-aminoethoxy)ethoxy]acetic acid was attached to 2-chlorotrityl chloride resin in the presence of DIPEA to obtain 2-[2-(2-Fmoc-aminoethoxy)ethoxy]acetic acid-2-Cl-Trt-resin. The Fmoc protecting group was removed by selective deblocking of the amino group using piperidine, and then 2-[2-(2-Fmoc-aminoethoxy)ethoxy]acetic acid in THF was bonded using DIPC and HOBt to obtain {(Fmoc-aminoethoxy)-ethoxy}acetyl-{(-aminoethoxy)-ethoxy}acetic acid-2-Cl-Trt-resin. The Fmoc group was removed by selective deblocking using piperidine, and the free amino group was bonded to Fmoc-Glu-OtBu using HOBt and DIPC to obtain Fmoc-Glu({(amino-ethoxy)-ethoxy}acetyl-{(-amino-ethoxy)-ethoxy}acetic acid-2-Cl-Trt-resin)-OtBu. The Fmoc group of the obtained compound was selectively deblocked using piperidine, and the free amino group was subsequently bonded to 20-(tert-butoxy)-20-oxoicosanoic acid. [2-[2-[[2-[2-[2-[2-[[5-tert-butoxy-4-[(20-tert-butoxy-20-oxo-icosanoyl)amino]-5-oxo-pentanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetic acid]-2-Cl-Trt-resin was obtained. Then, the intermediate was cleaved from the 2-Cl-Trt-resin using trifluoroethanol:DCM (1:1), 2-[2-[2-[[2-[2-[2-[5-tert-butoxy-4-[(20-tert-butoxy-20-oxo-icosanoyl)amino]-5-oxo-pentanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetic acid (partially D-di-tert-butyl ester) (LCMS=m / z:874.15(M+H + )) was obtained.

[0162] Example 5: Preparation of Compound 1 The parent peptide was synthesized by a solid-phase method. The starting resin used for synthesis was Fmoc-Rink amide resin. Selective deblockage of the Fmoc-protecting amino group of the Rink amide resin was performed using piperidine to obtain Rink amide resin, which was then coupled with Fmoc-Ser(tBu)-OH to obtain Fmoc-Ser(tBu)-Rink amide resin. This coupling reaction was carried out using diisopropylcarbodiimide and N-hydroxybenzotriazole (DIPC-HOBt) as coupling reagents. This completed one cycle. Unbound amino groups at each amino acid bond were terminated / capped using acetic anhydride and diisopropylethylamine / pyridine. Selective deblockage of the amino group of the Fmoc-Ser(tBu)-Rink amide resin was performed using piperidine. Subsequently, coupling with Fmoc-Pro-OH using HOBt and DIPC was performed to obtain Fmoc-Pro-Ser(tBu)-rink amide resin. This completed the second cycle. Acetic anhydride and diisopropylethylamine / pyridine were used to terminate the unbound amino groups in each amino acid bond.

[0163] The three steps described above—selective capping, deblocking of Fmoc protection of amino acids attached to the resin, and binding to the Fmoc-protected amino group of an adjacent amino acid residue in the sequence—were repeated for the remaining 37 amino acid residues. Selective deblocking, i.e., capping of unbound amino groups, was performed using acetic anhydride and diisopropylethylamine / pyridine, deprotection of the Fmoc group was performed using piperidine, and binding to the adjacent Fmoc-protected amino acid was performed using HOBt / DIPC. The side chains of Fmoc-protected amino acids are protected at right angles; for example, the hydroxyl group of serine, tyrosine, or threonine are protected with a tert-butyl (-tBu) group, the amino group of lysine is protected with a tert-butyloxycarbonyl (-Boc) and (4,4-dimethyl-2,6-dioxocyclohex-1-ylidene)-3-methylbutyl (IVDde) group, respectively, the carboxylic acid group of aspartic acid or glutamic acid is protected with a -tBu group, and the amide group of glutamine is protected with a trityl (-Trt) group. The three steps described above—selective capping, deblocking, and subsequent binding with adjacent Fmoc-protecting amino acids—were used to obtain Fmoc-Tyr(tBu)-Aib-Glu(OtBu)-Gly-Thr(tBu)-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Tyr(tBu)-Ser(tBu)-Ile-[L-norvaline]-Leu-Asp(OtBu)-Lys(Boc)-Ile-Ala-Gln(Trt)-Lys(IVDde)-Ala-Phe-Val-Gln(Trt)-Trp-Leu-Ile-Ala-Gly-Gly-Pro-Ser(tBu)-Ser(tBu)-Gly-Ala-Pro-Pro-Pro-Ser(tBu)-Resin.

[0164] Following deblocking of the Fmoc-Tyr(tBu)-Aib-Glu(OtBu)-Gly-Thr(tBu)-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Tyr(tBu)-Ser(tBu)-Ile-[L-norvaline]-Leu-Asp(OtBu)-Lys(Boc)-Ile-Ala-Gln(Trt)-Lys(IVDde)-Ala-Phe-Val-Gln(Trt)-Trp-Leu-Ile-Ala-Gly-Gly-Pro-Ser(tBu)-Ser(tBu)-Gly-Ala-Pro-Pro-Pro-Ser(tBu)-resin with piperidine, Boc By protecting the peptide resin using anhydrous material with Boc, we obtained a Boc-Tyr(tBu)-Aib-Glu(OtBu)-Gly-Thr(tBu)-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Tyr(tBu)-Ser(tBu)-Ile-[L-norvaline]-Leu-Asp(OtBu)-Lys(Boc)-Ile-Ala-Gln(Trt)-Lys(IVDde)-Ala-Phe-Val-Gln(Trt)-Trp-Leu-Ile-Ala-Gly-Gly-Pro-Ser(tBu)-Ser(tBu)-Gly-Ala-Pro-Pro-Pro-Ser(tBu)-resin.The IVDde group of the peptide resin is deprotected using hydrazine hydrate, and then it is coupled to a partial A-di-tert-butyl ester using diisopropylcarbodiimide and N-hydroxybenzotriazole (DIPC-HOBt) as coupling reagents to form an intermediate compound resin: Boc-Tyr(tBu)-Aib-Glu(OtBu)-Gly-Thr(tBu)-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Tyr(tBu)-Ser(tBu)-Ile-[L-norvaline]-Leu-Asp(OtBu)-Lys(Boc)-Ile-Ala-Gln(Trt)-Lys(NH-partial A A resin was obtained from di-tert-butylester)-Ala-Phe-Val-Gln(Trt)-Trp-Leu-Ile-Ala-Gly-Gly-Pro-Ser(tBu)-Ser(tBu)-Gly-Ala-Pro-Pro-Pro-Ser(tBu)-resin. This resin was then cleaved and deprotected with ethane-1,2-dithiol and triisopropylsilane using trifluoroacetic acid, followed by purification by preparative HPLC to obtain compound 1.

[0165] Mass (LCMS): m / z=1192.7(MH4 4+ ), Calculated mass = 4766.77, HPLC purity: (Method B) 98.60%, RT = 33.8 min

[0166] Example 6: Synthesis of Compound 2: Compound 2 was prepared by a solid-phase method according to a similar process described in Example 5, but Fmoc-[L-norvaline]-OH was used at position 2 instead of Fmoc-Aib-OH, and Fmoc-Aib-OH was used at position 13 instead of Fmoc-[L-norvaline]-OH, and Boc-Tyr(tBu)-[L-norvaline]-Glu(OtBu)-Gly-Thr(tBu)-Phe-Thr(tBu)-Ser(tB u)-Asp(OtBu)-Tyr(tBu)-Ser(tBu)-Ile-Aib-Leu-Asp(OtBu)-Lys(Boc)-Ile-Ala-Gln(Trt)-Lys(IVDde)-Ala-Ph e-Val-Gln(Trt)-Trp-Leu-Ile-Ala-Gly-Gly-Pro-Ser(tBu)-Ser(tBu)-Gly-Ala-Pro-Pro-Pro-Ser(tBu)-resin was obtained.

[0167] Next, compound 2 was obtained by bonding it with a partial A-di-tertbutyl ester and then, according to Example 5, by subsequent cleavage, deprotection, and preparative purification using HPLC.

[0168] Mass (LCMS): m / z=1192.6(MH4 4+ ), Calculated mass = 4766.4, HPLC purity: (Method B) 96.09%, RT = 25.6 min Example 7: Synthesis of Compound 3: Compound 3 was prepared by a solid-phase method according to a similar process described in Example 5, but Fmoc-Leu-OH was used at position 13 instead of Fmoc-[L-norvaline]-OH to obtain Boc-Tyr(tBu)-Aib-Glu(OtBu)-Gly-Thr(tBu)-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Tyr(tBu)-Ser(tBu)-Ile-Leu-Leu-Asp(OtBu)-Lys(Boc)-Ile-Ala-Gln(Trt)-Lys(IVDde)-Ala-Phe-Val-Gln(Trt)-Trp-Leu-Ile-Ala-Gly-Gly-Pro-Ser(tBu)-Ser(tBu)-Gly-Ala-Pro-Pro-Pro-Ser(tBu)-Resin.

[0169] Next, compound 3 was obtained by bonding it with a partial A-di-tertbutyl ester and then, according to Example 5, by subsequent cleavage, deprotection, and preparative purification using HPLC.

[0170] Mass (LCMS): m / z=1196.1(MH4 4+ ), calculated mass = 4780.4, HPLC purity: (Method A) 95.27%, RT = 39.1 min.

[0171] Example 8: Synthesis of Compound 4: Compound 4 was prepared by a solid-phase method according to a similar process described in Example 5, but Fmoc-[L-norvaline]-OH was used at position 2 instead of Fmoc-Aib-OH to obtain Boc-Tyr(tBu)-[L-norvaline]-Glu(OtBu)-Gly-Thr(tBu)-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Tyr(tBu)-Ser(tBu)-Ile-[L-norvaline]-Leu-Asp(OtBu)-Lys(Boc)-Ile-Ala-Gln(Trt)-Lys(IVDde)-Ala-Phe-Val-Gln(Trt)-Trp-Leu-Ile-Ala-Gly-Gly-Pro-Ser(tBu)-Ser(tBu)-Gly-Ala-Pro-Pro-Pro-Ser(tBu)-Resin.

[0172] Next, compound 4 was obtained by bonding it with a partial A-di-tertbutyl ester and then, according to Example 5, by subsequent cleavage, deprotection, and preparative purification using HPLC.

[0173] Mass (LCMS): m / z=1196.32(MH4 4+ ), calculated mass = 4781.25, HPLC purity: (Method A) 94.21%, RT = 29.8 min.

[0174] Example 9: Synthesis of Compound 5: Deprotection of the IVDde group of the peptide resin: Boc-Tyr(tBu)-Aib-Glu(OtBu)-Gly-Thr(tBu)-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Tyr(tBu)-Ser(tBu)-Ile-[L-norvaline]-Leu-Asp(OtBu)-Lys(Boc)-Ile-Ala-Gln(Trt)-Lys(IVDde)-Ala-Phe-Val-Gln(Trt)-Trp-Leu-Ile-Ala-Gly-Gly-Pro-Ser(tBu)-Ser(tBu)-Gly-Ala-Pro-Pro-Pro-Ser(tBu)-resin (prepared according to Example 5) was carried out using hydrazine hydrate, and diisopropylcarbodiimide, N-hydroxybenzotriazole (DIPC-HOBt) was used as a coupling reagent to conjugate with the partial C-di-tert-butyl ester, and the intermediate compound resin Boc-Tyr(tBu)-Aib-Glu(OtBu)-Gly-Thr(tBu)-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Tyr(tBu)-Ser(tBu)-Ile-[L-norvaline]-Leu-Asp(OtBu)-Lys(Boc)-Ile-Ala-Gln(Trt)-Lys(NH-partial C di-tert-butyl ester)-Ala-Phe-Val-Gln(Trt)-Trp-Leu-Ile-Ala-Gly-Gly-Pro-Ser(tBu)-Ser(tBu)-Gly-Ala-Pro-Pro-Pro-Ser(tBu)-resin was obtained. This was cleaved and deprotected using trifluoroacetic acid together with ethan-1,2-dithiol and triisopropylsilane, and then purified through preparative HPLC to obtain Compound 5.

[0175] Mass (LCMS): m / z = 1600.80 (MH3 3+ ), calculated mass = 4799.376, HPLC purity: (Method A) 98.64%, RT = 15.9 minutes

[0176] Example 10: Synthesis of Compound 6: Compound 6 was prepared by a solid-phase method according to a similar process described in Example 9, followed by bonding with a partial D-di-tert-butyl ester, and then cleavage, deprotection, and preparative HPLC purification as used in Example 9 to obtain compound 6.

[0177] Mass (LCMS): m / z=1609.98(MH3 3+ ), calculated mass = 4826.916, HPLC purity (method A): 96.31%, RT=26.7 min

[0178] Example 11: Synthesis of Compound 7: Compound 7 was prepared by a solid-phase method according to a similar process described in Example 9, bonded with a partial β-di-tert-butyl ester, followed by cleavage, deprotection, and preparative HPLC purification as described in Example 9 to obtain compound 7.

[0179] Mass (LCMS): m / z=1580.64(MH3 3+ ), calculated mass = 4738.896, HPLC purity (method A): 98.43%, RT=18.5 min Example 12: Synthesis of Compound 8: The parent peptide was synthesized by a solid-phase method. The starting resin used for synthesis was Fmoc-Rink amide resin. The Fmoc-protecting amino group of the Rink amide resin was selectively deblocked using piperidine, followed by the bonding of Fmoc-Lys(IVDde)-OH to the Rink amide resin. The bonding was carried out using DIPC-HOBt to obtain Fmoc-Lys(IVDde)-Rink amide resin, completing one cycle. Acetic anhydride and diisopropylethylamine / pyridine were used to terminate / cap the unbonded amino group at the end of each amino acid bond. Selective deblocking of the Fmoc amino group of the Fmoc-Lys(IVDde)-Rink amide resin using piperidine, followed by bonding with the second amino acid using HOBt and DIPC, yielded Fmoc-Ser(tBu)-Lys(IVDde)-rink amide resin. This completed the second cycle. As previously mentioned, acetic anhydride and diisopropylethylamine / pyridine were used to terminate the unbound amino group [capping] after each amino acid bond.

[0180] The three steps described above—deblocking the Fmoc protection of amino acids attached to the resin, binding the Fmoc-protected amino group to an adjacent amino acid residue in the sequence, and selective capping—were repeated for the remaining 38 amino acid residues. The side chains of the Fmoc-protected amino acids used were protected at right angles; for example, the hydroxyl group of serine, tyrosine, or threonine were protected with a tert-butyl (-tBu) group, the amino group of lysine was protected with a tert-butyloxycarbonyl (-Boc) and (4,4-dimethyl-2,6-dioxocyclohex-1-ylidene)-3-methylbutyl (IVDde) group, respectively, the carboxylic acid group of aspartic acid or glutamic acid was protected with a tBu group, and the amide groups of glutamine and asparagine were protected with a trityl (-Trt) group. The three steps described above—selective capping, deblocking, and subsequent binding with adjacent Fmoc-protecting amino acids—were performed to obtain the Fmoc-Tyr(tBu)-Aib-Glu(OtBu)-Gly-Thr(tBu)-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Tyr(tBu)-Ser(tBu)-Ile-L-norvaline-Leu-Glu(OtBu)-Lys(Boc)-Ile-Ala-Ala-Gln(Trt)-Glu(OtBu)-Phe-Val-Asn(Trt)-Trp-Leu-Leu-Ala-Gly-Gly-Pro-Ser(tBu)-Ser(tBu)-Gly-Ala-Pro-Pro-Pro-Ser(tBu)-Lys(IVDde)-Rink amide resin.

[0181] Fmoc-Tyr(tBu)-Aib-Glu(OtBu)-Gly-Thr(tBu)-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Tyr(tBu)-Ser(tBu)-Ile-L-Norvaline-Leu-Glu(OtBu)-Lys(Boc)-Ile-Al a-Ala-Gln(Trt)-Glu(OtBu)-Phe-Val-Asn(Trt)-Trp-Leu-Leu-Ala-Gly-Gly-Pro-Ser(tBu)-Ser(tBu)-Gly-Ala-Pro-Pro-Pro-Ser(tBu)-Lys(IVDde)-Rink Deblocking of the Fmoc group from the amide resin was performed using piperidine, followed by Boc protection of the peptide resin using Boc anhydride, to obtain the Boc-Tyr(tBu)-Aib-Glu(OtBu)-Gly-Thr(tBu)-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Tyr(tBu)-Ser(tBu)-Ile-L-norvaline-Leu-Glu(OtBu)-Lys(Boc)-Ile-Ala-Ala-Gln(Trt)-Glu(OtBu)-Phe-Val-Asn(Trt)-Trp-Leu-Leu-Ala-Gly-Gly-Pro-Ser(tBu)-Ser(tBu)-Gly-Ala-Pro-Pro-Pro-Ser(tBu)-Lys(IVDde)-Rink amide resin.

[0182] Following the deprotection of the IVDde group of the peptide resin using hydrazine hydrate, a partial A-di-tert butyl ester was bonded using diisopropylcarbodiimide and N-hydroxybenzotriazole (DIPC-HOBt) as coupling reagents to obtain compound 8 resin.

[0183] Compound 8 was obtained by cleavage and deprotection of the Boc-Tyr(tBu)-Aib-Glu(OtBu)-Gly-Thr(tBu)-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Tyr(tBu)-Ser(tBu)-Ile-L-norvaline-Leu-Glu(OtBu)-Lys(Boc)-Ile-Ala-Ala-Gln(Trt)-Glu(OtBu)-Phe-Val-Asn(Trt)-Trp-Leu-Leu-Ala-Gly-Gly-Pro-Ser(tBu)-Ser(tBu)-Gly-Ala-Pro-Pro-Pro-Ser(tBu)-Lys(NH molar A-di-tert butyl ester)-Rink amide resin using trifluoroacetic acid with ethane-1,2-dithiol and triisopropylsilane, followed by purification by preparative HPLC. The HPLC purity of compound 8 was evaluated by the following method: mass (LCMS): m / z = 980.42 (MH5 5+) Calculated mass = 4897.06 HPLC purity (Method D): 94.55%, RT = 44.9 min

[0184] Example 13: Synthesis of Compound 9: The parent peptide was synthesized by solid-phase method. The starting resin used in the synthesis was Wang resin. Fmoc-protected Arg(Pbf) was used for bonding with the Wang resin. Bonding was carried out in the presence of 4-dimethylaminopyridine (DMAP) using diisopropylcarbodiimide and N-hydroxybenzotriazole (DIC-HOBt) as bonding reagents to obtain Fmoc-Arg(Pbf)-Wang resin. Selective deblocking of the amino groups of the Fmoc-Arg(Pbf)-Wang resin using piperidine, followed by bonding with Fmoc-Ser(tBu)-OH using HOBt / DIPC, obtained Fmoc-Ser(tBu)-Arg(Pbf)-Wang resin. This completes one cycle. Unbound amino groups at each amino acid bond were terminated using acetic anhydride and diisopropylethylamine / pyridine.

[0185] The two steps described above, namely selective deblocking of the Fmoc protection of amino acids attached to the resin, and the binding of adjacent amino acid residues in the sequence to the Fmoc-protected amino group, were repeated for the remaining 38 amino acid residues to protect the side chains of the Fmoc-protected amino acids at right angles. For example, the hydroxyl group of serine, tyrosine, or threonine was protected with a tert-butyl (-tBu) group, the amino group of lysine was protected with a tert-butyloxycarbonyl (-Boc) and a (4,4-dimethyl-2,6-dioxocyclohex-1-ylidene)-3-methylbutyl (IVDde) group, respectively, and the carboxylic acid group of aspartic acid or glutamic acid was protected with a -tBu group, respectively. The three steps described above—selective capping, deblocking, and subsequent binding with adjacent Fmoc-protecting amino acids—were performed to obtain the Fmoc-Tyr(tBu)-Aib-Glu(OtBu)-Gly-Thr(tBu)-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Tyr(tBu)-Ser(tBu)-Ile-L-norvaline-Leu-Asp(OtBu)-Lys(Boc)-Ile-Ala-Gln(Trt)-Lys(IVDde)-Ala-Phe-Val-Gln(Trt)-Trp-Leu-Ile-Ala-Gly-Gly-Pro-Ser(tBu)-Ser(tBu)-Gly-Ala-Pro-Pro-Pro-Ser(tBu)-Arg(Pbf)-Wang resin.

[0186] Following deblocking of Fmoc-Tyr(tBu)-Aib-Glu(OtBu)-Gly-Thr(tBu)-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Tyr(tBu)-Ser(tBu)-Ile-L-norvaline-Leu-Asp(OtBu)-Lys(Boc)-Ile-Ala-Gln(Trt)-Lys(IVDde)-Ala-Phe-Val-Gln(Trt)-Trp-Leu-Ile-Ala-Gly-Gly-Pro-Ser(tBu)-Ser(tBu)-Gly-Ala-Pro-Pro-Pro-Ser(tBu)-Arg(Pbf)-Wang resin with piperidine, Boc By protecting the peptide resin using anhydrous peptides with Boc, we obtained Boc-Tyr(tBu)-Aib-Glu(OtBu)-Gly-Thr(tBu)-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Tyr(tBu)-Ser(tBu)-Ile-L-norvaline-Leu-Asp(OtBu)-Lys(Boc)-Ile-Ala-Gln(Trt)-Lys(IVDde)-Ala-Phe-Val-Gln(Trt)-Trp-Leu-Ile-Ala-Gly-Gly-Pro-Ser(tBu)-Ser(tBu)-Gly-Ala-Pro-Pro-Pro-Ser(tBu)-Arg(Pbf)-Wang resin. Following the deprotection of the IVDde group of the peptide resin using hydrazine hydrate, a partial A-di-tert butyl ester was bonded using diisopropylcarbodiimide and N-hydroxybenzotriazole (DIPC-HOBt) as coupling reagents to obtain compound 9-Wang resin.

[0187] Boc-Tyr(tBu)-Aib-Glu(OtBu)-Gly-Thr(tBu)-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Tyr(tBu)-Ser(tBu)-Ile-L-norvaline-Leu-Asp(OtBu)-Lys(Boc)-Ile-Ala-Gln(Trt)-Lys(NH partial A-di-tert butyl ester)-Ala-Phe-Val-Gln(Trt)-Trp-Leu-Ile-Ala-Gly-Gly-Pro-Ser(tBu)-Ser(tBu)-Gly-Ala-Pro-Pro-Pro-Ser(tBu)-Arg(Pbf)-Wang resin.

[0188] Cleavage and deprotection from the resin using trifluoroacetic acid with ethane-1,2-dithiol and triisopropylsilane, followed by purification by preparative HPLC, yielded compound 9. Mass (LCMS): m / z = 985.88 (MH5 5+ ), Calculated mass = 4924.36, HPLC purity: (Method C) 93.52%, RT = 27.8 min

[0189] Example 14: Synthesis of Compound 10: Compound 10 was prepared by a solid-phase method according to a similar process described in Example 5, but Fmoc-[2-aminobutyric acid] was used at position 13 instead of Fmoc-[L-norvaline]-OH to obtain Boc-Tyr(tBu)-Aib-Glu(OtBu)-Gly-Thr(tBu)-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Tyr(tBu)-Ser(tBu)-Ile-2-aminobutyric acid-Leu-Asp(OtBu)-Lys(Boc)-Ile-Ala-Gln(Trt)-Lys(IVDde)-Ala-Phe-Val-Gln(Trt)-Trp-Leu-Ile-Ala-Gly-Gly-Pro-Ser(tBu)-Ser(tBu)-Gly-Ala-Pro-Pro-Pro-Ser(tBu)-Resin.

[0190] Next, compound 10 was obtained by bonding it with a partial A-di-tertbutyl ester and then, according to Example 5, by subsequent cleavage, deprotection, and preparative purification using HPLC.

[0191] Mass (LCMS): m / z=1189.57(MH4 4+ ), Calculated mass = 4754.248, HPLC purity: (Method D) 96.79%, RT = 42.6 min

[0192] Example 15: Synthesis of Compound 11: Compound 11 was prepared by a solid-phase method according to a similar process described in Example 5, but Fmoc-norleucine was used at position 13 instead of Fmoc-[L-norvaline]-OH to obtain Boc-Tyr(tBu)-Aib-Glu(OtBu)-Gly-Thr(tBu)-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Tyr(tBu)-Ser(tBu)-Ile-norleucine-Leu-Asp(OtBu)-Lys(Boc)-Ile-Ala-Gln(Trt)-Lys(IVDde)-Ala-Phe-Val-Gln(Trt)-Trp-Leu-Ile-Ala-Gly-Gly-Pro-Ser(tBu)-Ser(tBu)-Gly-Ala-Pro-Pro-Pro-Ser(tBu)-Resin.

[0193] Next, compound 11 was obtained by bonding with a partial A-di-tert-butyl ester and then cleaving, deprotecting, and preparative HPLC purification according to Example 5.

[0194] Mass (LCMS): m / z=1196.66(MH4 4+ ), Calculated mass = 4782.608, HPLC purity: (Method D) 95.43%, RT = 58.7 min

[0195] Example 16: Synthesis of Compound 12: Compound 12 was prepared by a solid-phase method according to a similar process described in Example 5, except that Fmoc-Ile-OH was used at position 13 instead of Fmoc-[L-norvaline]-OH, to obtain Boc-Tyr(tBu)-Aib-Glu(OtBu)-Gly-Thr(tBu)-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Tyr(tBu)-Ser(tBu)-Ile-Ile-Leu-Asp(OtBu)-Lys(Boc)-Ile-Ala-Gln(Trt)-Lys(IVDde)-Ala-Phe-Val-Gln(Trt)-Trp-Leu-Ile-Ala-Gly-Gly-Pro-Ser(tBu)-Ser(tBu)-Gly-Ala-Pro-Pro-Pro-Ser(tBu)-resin.

[0196] Next, it was coupled with the partial A-di-tert-butyl ester, and according to Example 5, followed by cleavage, deprotection, and preparative HPLC purification to obtain Compound 12.

[0197] Mass (LCMS): m / z = 1196.32 (MH4 4+ ), calculated mass = 4781.25, HPLC purity: (Method D) 94.38%, RT = 47.7 minutes.

[0198] Biological studies Example 12: Efficacy test in db / db mice at a dose of 10 nM / kg The effects of the compounds of the present invention on blood glucose levels, food intake, and body weight were studied in mice. This study was conducted in a type 2 diabetic mouse (db / db) model. The animals were divided into four treatment groups (n=6), a diabetic control group, compound 1 (10 nM / kg), compound 2 (10 nM / kg), and tilzepatide (10 nM / kg). Baseline blood glucose levels were measured in all animals. All animals were administered the test compound subcutaneously. Blood glucose levels were measured at 4, 8, 12, 24, 48, 72, and 96 hours post-treatment. Delta blood glucose levels (mM) were calculated. The results are provided in Table 3. Similarly, body weight change and cumulative food consumption were measured at 48 and 96 hours post-treatment. The results for body weight change are shown in Table 4, and cumulative food consumption is shown in Table 5. Similarly, the efficacy of compounds 3, 4, 5, 6, and 7 in db / db mice at 10 nM / kg doses was performed in separate studies. The animals were divided into seven treatment groups (n=6), a diabetes control group, and compound 1, compound 3, compound 4, compound 5, compound 6, and compound 7. Baseline blood glucose levels were measured in all animals. All animals received subcutaneous administration of the test compound. Blood glucose levels were measured at 4, 8, 12, 24, 48, 72, and 96 hours post-treatment. Delta blood glucose levels (mM) were calculated. The results are also provided in Table 3. Similarly, body weight change and cumulative food consumption were measured at 48 and 96 hours post-treatment. The results for body weight change are also provided in Table 4, and cumulative food consumption is provided in Table 5. Similarly, the efficacy of compounds 8 and 9 in db / db mice at a dose of 10 nM / kg was conducted in a separate study. The animals were divided into three treatment groups (n=5), a diabetes control group, compound 8, and compound 9. Baseline blood glucose levels were measured in all animals. All animals received subcutaneous administration of the test compound. Blood glucose levels were measured at 4, 8, 12, 24, 48, 72, and 96 hours post-treatment. Delta blood glucose levels (mM) were calculated. The results are provided in Table 3. Similarly, body weight change and cumulative food consumption were measured at 48 and 96 hours post-treatment. The results for body weight change are shown in Table 4, and cumulative food consumption is shown in Table 5. Separate efficacy studies were conducted for compounds 10, 11, and 12 in db / db mice at a dose of 10 nM / kg.The animals were divided into four treatment groups (n=5), a diabetes control group, compound 10, compound 11, and compound 12. Baseline blood glucose levels were measured in all animals. All animals received subcutaneous administration of the test compound. Blood glucose levels were measured at 4, 8, 12, 24, 48, 72, and 96 hours post-treatment. Delta blood glucose levels (mM) were calculated. The results are provided in Table 3. Similarly, body weight change and cumulative food consumption were measured at 48 and 96 hours post-treatment. The results for body weight change are shown in Table 4, and cumulative food consumption is shown in Table 5.

[0199] [Table 8] TIFF0007830441000058.tif37168

[0200] [Table 9]

[0201] [Table 10]

[0202] The results show that compounds 1 and 2 demonstrated a statistically significant reduction in blood glucose up to 96 hours post-treatment. The effect of the compounds on blood glucose reduction was superior to that of tilzepatide when tested at the same concentrations.

[0203] Compounds 1 and 2 also showed statistically significant weight loss comparable to that of tirzepatide. Compound 1 showed a significant reduction in food consumption comparable to that of tirzepatide. No significant reduction in food consumption was observed with compound 2 compared to diabetic controls.

[0204] Similarly, the results show that compounds 3, 4, 5, 6, and 7 of the present invention demonstrated a statistically significant reduction in blood glucose up to 96 hours post-treatment. Furthermore, statistically significant reductions in food intake and body weight were also observed with these compounds compared to diabetic controls.

[0205] Example 13: Efficacy study in db / db mice at doses of 3 and 20 nM / kg The effects of the compounds of the present invention on blood glucose levels, food intake, and body weight were studied in mice. This study was conducted using a type 2 diabetic mouse (db / db) model. The animals were divided into five treatment groups (n=8 per group): a diabetic control group, compound 1 (3 nM / kg and 20 nM / kg), and tilzepatide (3 nM / kg and 20 nM / kg). Baseline blood glucose levels were measured in all animals. All animals were administered the test compound subcutaneously. Blood glucose levels were measured at 4, 24, 48, and 72 hours post-treatment. Delta blood glucose levels (mM) were calculated. Body weight change and cumulative food consumption were measured at 72 hours post-treatment. The results for delta blood glucose are provided in Table 6. Similarly, the results for body weight change are shown in Table 7, and food consumption is presented in Table 8.

[0206] [Table 11]

[0207] [Table 12]

[0208] [Table 13]

[0209] The results show that compound 1 at 3 nM / kg and 20 nM / kg demonstrated a dose-dependent improvement in glucose-lowering effect up to 72 hours. The effect was superior to that of tilzepatide at similar doses.

[0210] The effects of compound 1 on food intake and body weight at doses of 3 and 20 nM / kg were significant, dose-dependent, and comparable to those of tilzepatide.

[0211] The above results indicate that the compounds of the present invention are potent inhibitors of GLP-1 and GIP receptors and may be effective in treating type 2 diabetes, diabetes with obesity, obesity, and hyperlipidemia.

[0212] Example 16: Cellular cAMP assay In vitro efficacy measurements were performed using a cAMP assay. Activation of G protein-coupled receptors (GPCRs) following ligand binding initiates a second messenger cascade that leads to a cellular response. Signaling by GLP-1R and GIP-R involves activation of adenylyl cyclase and cAMP production. Cellular cAMP production was determined using the cAMP Hunter® eXpress GPCR assay (Eurofins DiscoveRx).

[0213] Cellular cAMP assays were performed for tilzepatide, compound 1, compound 3, compound 6, compound 10, and compound 11, and the semi-effective concentrations on GLP-1R-expressing cells and GIPR-expressing cells are shown in Table 9 below.

[0214] [Table 14]

Claims

1. It contains the following amino acid sequence, Y - In the formula, X1 is Aib and X2 is (L)-norvaline; or both X1 and X2 are (L)-norvaline; or X1 is Aib and X2 is Leu. X4 is Ile, X3 is Lys, and in the formula, the amino (ε-amino) group of the Lys side chain is acylated at the following point: 【Chemistry 1】 In the formula, U is -C(O)-CH ２ -O-(CH ２ ) ２ -O-(CH ２ ) ２ } is -NH-, where} is the attachment point with the base W, W is -C(O)-C(CH ３ ) ２ -NH-], where ] is the attachment point with base Y, Y is -C(O)-(CH ２ ) ２ -CH(COOH)NH--, where -- is the attachment point to group Z Z is -C(O)-(CH ２ ) ｎ -COOH, where n is an integer between 14 and 20. Furthermore, in the formula, the acid group of the C-terminal amino acid is either a free carboxylic acid group or is amidated as a C-terminal primary amide. A polypeptide or a pharmaceutically acceptable salt thereof.

2. X1 is Aib, X2 is (L)-norvaline, W is -C(O)-C(CH ３ ) ２ -NH-] and Z is -C(O)-(CH ２ ) ｎ A polypeptide according to claim 1, wherein n is 18, and a pharmaceutically acceptable salt thereof.

3. Z is -C(O)-(CH ２ ) ｎ A polypeptide according to claim 1, wherein n is -COOH and n is 16 or 18, or a pharmaceutically acceptable salt thereof.

4. A polypeptide according to claim 1, comprising an amino acid sequence selected from the group consisting of the following, or a pharmaceutically acceptable salt thereof. i.) Tyr Aib Glu Gly Thr Phe Thr Ser Asp Tyr Ser Ile L-norvaline Leu Asp Lys Ile Ala Gln Lys Ala Phe Val Gln Trp Leu Ile Ala Gly Gly Pro Ser Ser Gly Ala Pro Pro Pro Ser-NH ２ (SEQ ID NO: 04) ii.) Tyr Aib Glu Gly Thr Phe Thr Ser Asp Tyr Ser Ile Leu Leu Asp Lys Ile Ala Gln Lys Ala Phe Val Gln Trp Leu Ile Ala Gly Gly Pro Ser Ser Gly Ala Pro Pro Pro Ser-NH ２ (SEQ ID NO: 06) iii.) Tyr L-norvaline Glu Gly Thr Phe Thr Ser Asp Tyr Ser Ile L-norvaline Leu Asp Lys Ile Ala Gln Lys Ala Phe Val Gln Trp Leu Ile Ala Gly Gly Pro Ser Ser Gly Ala Pro Pro Pro Ser-NH ２ (Sequence ID 07)

5. A polypeptide according to any one of claims 1 to 4, wherein -U-W-Y-Z is selected from the group consisting of the following: a pharmaceutically acceptable salt thereof. 【Chemistry 2】

6. X1 is Aib, X2 is (L)-norvaline, -U-W-Y-Z is part A below, 【Transformation 3】 A polypeptide according to any one of claims 1 to 5 or a pharmaceutically acceptable salt thereof.

7. The polypeptide according to any one of claims 1 to 6, wherein the C-terminal amino acid is amidated as a C-terminal primary amide, or a pharmaceutically acceptable salt thereof.

8. The polypeptide according to any one of claims 1 to 6, wherein the C-terminal amino acid is a free carboxylic acid, or a pharmaceutically acceptable salt thereof.

9. A polypeptide or a pharmaceutically acceptable salt thereof, selected from the group consisting of the following: Compound 1: 【Chemistry 4】 ;Compound 3: 【Transformation 5】 ； Compound 4: 【Transformation 6】 ; and Compound 7: 【Transformation 7】 ； In the formula, part A is, 【Transformation 8】 And; and Part B is, 【Chemistry 9】 That is the case.

10. A pharmaceutical composition comprising a polypeptide according to any one of claims 1 to 9 or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable excipients.

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