GLP-1 / GIP dual, GLP-1 / GCG dual, and GLP-1 / GIP / GCG triple receptor agonists

Abstract

The present disclosure provides GLP-1 / GIP and GLP-1 / GCG dual receptor agonists comprising an insulinotropic analogue polypeptide and their use in the treatment or prevention of Type 2 diabetes (T2DM), hyperlipidemia / dyslipidemia, metabolic syndrome, metabolic dysfunction-associated fatty liver disease (MASLD), metabolic dysfunction-associated fatty hepatitis (MASH), neurodegenerative disorders, fibrosis, liver disease, liver disease, liver disease, liver disease, liver disease, liver disease, liver disease, liver disease, liver disease, liver disease, liver disease, liver disease, liver disease, liver disease, liver disease, liver disease, liver disease, liver disease, liver disease, liver disease, liver disease, liver disease, liver disease, the invention also relates to applications in cardiovascular risk and / or obesity.

Description

[0001] Cross-references to related applications

[0002] This application claims priority to Indian application No. 202321039646, filed on June 9, 2023, which is incorporated herein by reference in its entirety.

[0003] Reference to the electronic sequence list

[0004] This application includes a sequence list submitted electronically, which is incorporated herein by reference in its entirety. The sequence list was created on June 7, 2024, named "24-0805-WO_Sequence-Listing.xml", and is 110,592 bytes in size. Technical Field

[0005] This disclosure relates to glucagon-like peptide-1 (GLP-1) monoreceptor agonists, GLP-1 / glucose-dependent insulinotropic peptides or gastrointestinal peptide (GIP) dual-receptor agonists, and GLP-1 / glucagon (GCG) dual-receptor agonists. In particular, this disclosure relates to GLP-1 / GIP and GLP-1 / GCG dual-receptor agonists comprising incretin analog peptides. The peptides described herein have structural features providing balanced activity and prolonged duration of action on each of these receptors. The peptides according to the invention can be used to treat type 2 diabetes mellitus (T2DM), hyperlipidemia / dyslipidemia, metabolic syndrome, metabolic dysfunction-associated fatty liver disease (MASLD), metabolic dysfunction-associated steatohepatitis (MASH), neurodegenerative diseases, fibrosis, and / or obesity, as well as to reduce cardiovascular risk. Background Technology

[0006] Over the past few decades, the prevalence of diabetes has been steadily increasing. Type 2 diabetes mellitus (T2DM) is the most common form of diabetes, characterized by high blood sugar levels caused by insulin resistance. People with T2DM are more likely to develop comorbidities such as hyperlipidemia / dyslipidemia, metabolic syndrome, metabolic dysfunction-associated fatty liver disease (MASLD), metabolic dysfunction-associated steatohepatitis (MASH), neurodegenerative diseases, fibrosis, cardiovascular risk, and / or obesity.

[0007] Current treatments for type 2 diabetes mellitus (T2DM) include diet and exercise, as well as oral and injectable medications to lower blood sugar, including incretin-based therapies such as GLP-1 monoreceptor agonists and / or GLP-1 / GLP dual-receptor agonists. For example, Cotadutide (SEQ ID NO: 6), MK-1462 (SEQ ID NO: 7), and Mazdutide (SEQ ID NO: 8) are peptides that act as GLP-1 / GCG dual-receptor agonists.

[0008] WIPO publications WO2019 / 193576, WO2006 / 097537, and WO1998 / 008871 disclose GLP-1 single receptor agonist compounds. WIPO publications WO2022 / 079639, WO2021 / 260530, WO2017 / 74714A1, WO2020 / 23386, WO2020 / 023388, WO2015 / 067715, WO2016 / 111971, WO2014 / 192284, WO2011 / 119657, and WO2013 / 164483 disclose GLP-1 / GIP dual receptor agonist compounds. WIPO publications WO2011 / 075393, WO2012 / 177444, WO2014 / 091316 and WO2017 / 153575 disclose GLP-1 / GCG dual receptor agonist compounds.

[0009] Recent studies on GLP-1 / GCG dual and / or GLP-1 / GIP / GCG triple receptor agonists have also highlighted the importance of understanding the contribution and different effects of individual hormones by altering GLP-1:GCG activity and the ratio in GLP-1 / GCG dual receptor agonists and GLP-1 / GIP / GCG triple receptor agonists.

[0010] Glucagon receptor (GCGR) agonism is known to cause diabetes, increase heart rate and contractility, which can lead to adverse cardiovascular outcomes. Further chronic excess of glucagon also leads to the catabolism of amino acids and proteins, resulting in a loss of lean body mass. These side effects offset the weight loss benefits provided by GCG agonism.

[0011] A delicate balance needs to be struck between GLP-1 and GCG receptor agonism to achieve the best results with minimal side effects.

[0012] Compared to GLP-1R, the now-discontinued cotadiopeptide has shown higher efficacy against GCGR.

[0013] While the broad metabolic benefits of GLP-1 monoreceptor agonists or GLP-1 / GIP dual receptor agonist compounds have been established in therapeutic paradigms, there remains a need for treatments that can provide effective glucose control with weight loss benefits and a favorable profile of adverse effects, particularly for type 2 diabetes mellitus (T2DM). There is also a need for therapeutic agents that can be used with a sufficiently prolonged duration of action to allow for infrequent dosing, either twice weekly or once weekly. Summary of the Invention

[0014] In one aspect, this disclosure provides a polypeptide comprising the following amino acid sequence or a pharmaceutically acceptable salt thereof: X1-X2-X3-GTFTSD-X10-S-X12-X13-LD-X16-X17-X18-X19-X20-X21-F-X23-X 24-X25-L-X27-X28-X29-X30-X31-X32-X33-X34-X35-X36-X37-X38-X39 (SEQ ID NO: 1) in: X1 is H; X2 is D-Ser(OMe), Aib, or DS; X3 is Q; X10 is either K or Y; X12 is E, K, or I; X13 is Y, S(OMe), nor-V, nor-L, or αMe-L; X16 is S, E, or A; X17 is E, R, or K; X18 is R, K, or A; X19 is A; X20 is R, Q, or K; X21 is either D or E; X23 is either V or I; X24 is A, Q, or E; X25 is W; X27 is either E or L; X28 is A, D, or E; X29 is either G or T; X30 does not exist or is G; X31 does not exist or is P; X32 does not exist or is S; X33 does not exist or is S; X34 does not exist or is G; X35 does not exist; X36 does not exist; X37 does not exist; X38 does not exist; and X39 does not exist; The acid group of the C-terminal amino acid is either a free carboxylic acid group or is amidated to a C-terminal primary amide; and The condition is that at least one of X10 and X20 is K, and the further condition is that at least one of K contains a partially acylated side chain amino group selected from the following formula ( amino group:

[0015] In another respect, this disclosure provides a polypeptide comprising the following amino acid sequence or a pharmaceutically acceptable salt thereof: X1-X2-X3-GTFTSD-X10-S-X12-X13-LD-X16-X17-X18-X19-X20-X21-F-X23-X 24-X25-L-X27-X28-X29-X30-X31-X32-X33-X34-X35-X36-X37-X38-X39 (SEQ ID NO: 2) in: X1 is H; X2 is S, D-Ser(OMe), Aib, or DS; X3 is Q; X10 is either K or Y; X12 is E, K, or I; X13 is Y, S(OMe), nor-V, nor-L, or αMe-L; X16 is S, E, or A; X17 is E, R, or K; X18 is R, K, or A; X19 is A; X20 is R, Q, or K. X21 is either D or E; X23 is V; X24 is A, Q, or E; X25 is W; X27 is either E or L; X28 is A, D, or E; X29 is either G or T; X30 does not exist or is G; X31 does not exist or is P; X32 does not exist or is S; X33 does not exist or is S; X34 does not exist or is G; X35 does not exist; X36 does not exist; X37 does not exist; X38 does not exist; and X39 does not exist; The acid group of the C-terminal amino acid is either a free carboxylic acid group or is amidated to a C-terminal primary amide; and The condition is that at least one of X10 and X20 is K, and the further condition is that at least one of K contains a partially acylated side chain amino group selected from the following formula ( amino group:

[0016] In another respect, this disclosure provides a polypeptide comprising the following amino acid sequence or a pharmaceutically acceptable salt thereof: H-X2-QGTFTSD-X10-SEYLDSERARDFVAWLEAGG (SEQ IDNO: 3) in: X2 is S, DS(OMe), or Aib; and X10 is K; The acid group of the C-terminal amino acid is either a free carboxylic acid group or is amidated to a C-terminal primary amide. The side chain amino group of K at position X10 ( The amino group is partially acylated using the following formula:

[0017] The condition is that the polypeptide is not SEQ ID NO: 6.

[0018] In another respect, this disclosure provides a polypeptide comprising the following amino acid sequence or a pharmaceutically acceptable salt thereof: H-Aib-QGTFTSDYS-X12-X13-LDEKKA-X20-EFVEWLLEGPSSG (SEQ ID NO: 4) in: X12 is either K or I; X13 is either Y or nor-V; and X20 is K; The acid group of the C-terminal amino acid is either a free carboxylic acid group or is amidated to a C-terminal primary amide; and The side chain amino group of K at position X20 ( The amino group is partially acylated using the following formula:

[0019] In another respect, this disclosure provides a polypeptide comprising the following amino acid sequence or a pharmaceutically acceptable salt thereof: H-(DSer)-QGTFTSD-X10-SKYLDARAAQDFVQWLLDT (SEQID NO: 5) Where X10 is K; The acid group of the C-terminal amino acid is either a free carboxylic acid group or is amidated to a C-terminal primary amide; and The side chain amino group of K at position X10 ( The amino group is partially acylated using the following formula:

[0020] In one aspect, this disclosure provides a polypeptide selected from the group consisting of:

[0021] (SEQ ID NO: 18);

[0022] (SEQ ID NO: 21);

[0023] (SEQ ID NO: 22); and

[0024] (SEQ ID NO: 23).

[0025] On the other hand, this disclosure relates to an incretin analogue comprising: Having sequence X 1 The peptide residues of -Gln-Gly-Thr-Phe-Thr-Ser-Asp-Lys (SEQ ID NO: 29), wherein X 1 This indicates Aib or Ser(OMe), and wherein the lysine contains a fatty acid elongation group attached to the ε-nitrogen of the lysine; and Gly-Gly-OH peptide residues indirectly linked to the carboxyl group of the lysine.

[0026] On the other hand, this disclosure provides an incretin analog comprising: Having sequence X 1 The peptide residues of -Gln-Gly-Thr-Phe-Thr-Ser-Asp-Lys (SEQ ID NO: 29), wherein X 1 This represents Aib or Ser(OMe), and wherein the lysine contains a group of formula (I) linked to the ε-nitrogen of the lysine. (I) in: U does not exist or represents –C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-}, where} is the connection point with W; W indicates: –C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-], -C(O)-NH-(CH2)3-4-NH-], -C(O)-C(CH3)2-NH-] or

[0027] Where ] is the connection point with Y; Y does not exist or represents –C(O)-(CH2)2-CH(CO2H)NH-- or -C(O)CH((CH2) x CO2H)NH--, where x is 1, 2 or 3, and -- is the connection point with Z; and Z represents –C(O)-(CH2) n -COOH or –C(O)-(CH2) n -CH3, where n is an integer from 14 to 20; and Gly-Gly-OH peptide residues indirectly linked to the carboxyl group of the lysine.

[0028] On the other hand, this disclosure provides an incretin analog comprising: Peptide residues having the sequence Aib-Gln-Gly-Thr-Phe-Thr-Ser-Asp (SEQ ID NO: 30); Lysine residues indirectly linked to the carboxyl group of the residue Aib-Gln-Gly-Thr-Phe-Thr-Ser-Asp (SEQ ID NO: 30), wherein the lysine contains a group of formula (I) linked to the ε-nitrogen of lysine; (I) in: U does not exist or represents –C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-}, where} is the connection point with W; W indicates: –C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-]or -C(O)-NH-(CH2)3-4-NH-], -C(O)-C(CH3)2-NH-], Where ] is the connection point with Y; Y does not exist or represents –C(O)-(CH2)2-CH(CO2H)NH-- or -C(O)CH((CH2) x CO2H)NH--, where x is 1, 2 or 3, and -- is the connection point with Z; and Z represents –C(O)-(CH2) n -COOH or –C(O)-(CH2) n -CH3, where n is an integer from 14 to 20; and Gly-Gly-Pro-Ser-Ser-Gly-CONH2 peptide residues indirectly linked to the carboxyl group of the lysine.

[0029] In another respect, this disclosure relates to pharmaceutical compositions comprising a pharmaceutically acceptable carrier and an incretin analog or polypeptide as described herein.

[0030] In another aspect, this disclosure relates to a method for treating obesity, type 2 diabetes mellitus (T2DM), metabolic syndrome, metabolic dysfunction-associated fatty liver disease (MASLD), metabolic dysfunction-associated steatohepatitis (MASH), neurodegenerative diseases, fibrosis, cardiovascular risk, and / or hyperlipidemia / dyslipidemia, said method comprising administering an incretin analog or peptide as described herein to a patient in need of such treatment. Detailed Implementation

[0031] abbreviations

[0032] Aib: 2-Aminoisobutyric acid

[0033] DIPEA: N , N '-Diisopropylethylamine

[0034] HOBt: 1-Hydroxybenzotriazole

[0035] DIPC: N , N '-Diisopropylcarbodiimide

[0036] THF: Tetrahydrofuran

[0037] DCM: Dichloromethane

[0038] Fmoc: fluorenylmethoxycarbonyl

[0039] HOSu: N- Hydroxysuccinimide

[0040] DCC: Dicyclohexylcarbodiimide

[0041] DMAc: dimethylacetamide

[0042] IBCF: Isobutyl chloroformate

[0043] NMM: N -Methylmorpholine

[0044] DIC: Diisopropylcarbodiimide

[0045] definition

[0046] The term "pharmaceutically acceptable salt" according to this disclosure includes acid addition salts formed with organic or inorganic acids. Suitable pharmaceutically acceptable salts of the compounds of this disclosure include acid addition salts, which may be salts of inorganic acids such as hydrochloric acid, hydrobromic acid, phosphoric acid, etc., or salts of organic acids such as acetic acid, benzenesulfonic acid, methanesulfonic acid, benzoic acid, citric acid, lactic acid, fumaric acid, succinic acid, adipic acid, pimelic acid, octanoic acid, azelaic acid, malic acid, tartaric acid, amino acids (e.g., glutamic acid or aspartic acid), etc. Pharmaceutically acceptable acid addition salts of the compounds of this disclosure include salts formed by adding one or more equivalent amounts of acid, such as monohydrochloride or dihydrochloride. Salts can be prepared by any method within the knowledge of those skilled in the art. See , Berge et al. , J. Pharm. Sci. , 1977, 66, 1-19; and "Handbook ofPharmaceutical Salts: Properties, Selection, and Use," Stahl et al. edit, Verlag Helv. Chim. Acta Zurich, Switzerland, and Wiley-VCH , Weinheim, Germany, 2002.

[0047] As used in this article, terminology ""Effective dose" or "effective... dose" refers to an amount of compound that, when administered to a subject in a single or multiple doses, is sufficient to cure, alleviate, relieve, or partially resolve the clinical manifestations of a given disease or symptom and its complications, exceeding the clinical manifestations expected in the absence of such treatment. Therefore, the result may be a reduction and / or relief of signs, symptoms, or causes of the disease, or any other desired biological alteration. It should be understood that "therapeutic effective dose" can vary from subject to subject, depending on the subject's age, weight, general condition, the condition being treated, the severity of the condition being treated, and the prescribing physician's judgment.

[0048] As used in this article, the amino acid "Aib" can be represented by the following structure: It can also be defined by its chemical name, "2-aminoisobutyric acid".

[0049] As used in this article, the amino acid “S(OMe)” or “Ser(OMe)” can be represented by the following structure: It can also be defined by its chemical name, "serine methyl ether". The terms L-Ser(OMe) and D-Ser(OMe) refer to the "L" and "D" isomers of Ser(OMe), respectively.

[0050] As used herein, the amino acid “nor-V”, “nor-Val”, or “norvaline” can be represented by the following structure: It can also be defined by its chemical name, "2-aminovaleric acid". The terms L-valine and D-valine refer to the "L" and "D" isomers of valine, respectively.

[0051] As used in this article, the amino acid “nor-L”, “nor-Leu”, or “ortholeucine” can be represented by the following structure: It can also be defined by its chemical name, "2-aminohexanoic acid". The terms L-leucine and D-leucine refer to the "L" and "D" isomers of leucine, respectively.

[0052] As used in this article, the amino acid “αMe-L”, “αMe-Leu”, or “αMe-Leucine” can be represented by the following structures: It can also be defined by its chemical name, "2-amino-2,4-dimethylvaleric acid". The terms L-α-Me-leucine and D-α-Me-leucine refer to the "L" and "D" isomers of α-Me-leucine, respectively.

[0053] As described herein, this disclosure provides stable, long-acting GLP-1 monoreceptor, GLP-1 / GIP dual-receptor, and / or GLP-1 / GCG dual-receptor agonists that can be used to treat type 2 diabetes mellitus (T2DM), hyperlipidemia / dyslipidemia, metabolic syndrome, non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH), neurodegenerative diseases, fibrosis, and / or obesity, as well as to reduce cardiovascular risk.

[0054] In one aspect, this disclosure provides a polypeptide comprising the following amino acid sequence or a pharmaceutically acceptable salt thereof: X1-X2-X3-GTFTSD-X10-S-X12-X13-LD-X16-X17-X18-X19-X20-X21-F-X23-X 24-X25-L-X27-X28-X29-X30-X31-X32-X33-X34-X35-X36-X37-X38-X39 (SEQ ID NO: 1) in: X1 is H; X2 is D-Ser(OMe), Aib, or DS; X3 is Q; X10 is either K or Y; X12 is E, K, or I; X13 is Y, S(OMe), nor-V, nor-L, or αMe-L; X16 is S, E, or A; X17 is E, R, or K; X18 is R, K, or A; X19 is A; X20 is R, Q, or K; X21 is either D or E; X23 is either V or I; X24 is A, Q, or E; X25 is W; X27 is either E or L; X28 is A, D, or E; X29 is either G or T; X30 does not exist or is G; X31 does not exist or is P; X32 does not exist or is S; X33 does not exist or is S; X34 does not exist or is G; X35 does not exist; X36 does not exist; X37 does not exist; X38 does not exist; and X39 does not exist; The acid group of the C-terminal amino acid is either a free carboxylic acid group or is amidated to a C-terminal primary amide. The condition is that at least one of X10 and X20 is K, and the further condition is that at least one of K contains a partially acylated side chain amino group selected from the following formula ( amino group:

[0055] In one embodiment, the polypeptide of SEQ ID NO: 1 comprises the following sequence: X2 is D-Ser(OMe) or Aib; X10 is K; X12 is E; X13 is Y; X16 is S; X17 is E; X18 is R; X20 is R; X21 is D; X24 is A; X27 is E; X28 is A; X29 is G; X30 is G; and X31, X32, X33, and X34 do not exist; The side chain amino group of K at position X10 ( The amino group is partially acylated using the following formula:

[0056] In another respect, this disclosure provides a polypeptide comprising the following amino acid sequence or a pharmaceutically acceptable salt thereof: X1-X2-X3-GTFTSD-X10-S-X12-X13-LD-X16-X17-X18-X19-X20-X21-F-X23-X 24-X25-L-X27-X28-X29-X30-X31-X32-X33-X34-X35-X36-X37-X38-X39 (SEQ ID NO: 2) in: X1 is H; X2 is S, D-Ser(OMe), Aib, or DS; X3 is Q; X10 is either K or Y; X12 is E, K, or I; X13 is Y, S(OMe), nor-V, nor-L, or αMe-L; X16 is S, E, or A; X17 is E, R, or K; X18 is R, K, or A; X19 is A; X20 is R, Q, or K. X21 is either D or E; X23 is V; X24 is A, Q, or E; X25 is W; X27 is either E or L; X28 is A, D, or E; X29 is either G or T; X30 does not exist or is G; X31 does not exist or is P; X32 does not exist or is S; X33 does not exist or is S; X34 does not exist or is G; X35 does not exist; X36 does not exist; X37 does not exist; X38 does not exist; and X39 does not exist; The acid group of the C-terminal amino acid is either a free carboxylic acid group or is amidated to a C-terminal primary amide. The condition is that at least one of X10 and X20 is K, and the further condition is that at least one of K contains a partially acylated side chain amino group selected from the following formula ( amino group:

[0057] In one embodiment, the polypeptide of SEQ ID NO: 1 or SEQ ID NO: 2 comprises the following sequence: X2 is DS; X10 is K; X12 is K; X13 is Y; X16 is A; X17 is R; X18 is A; X20 is Q; X21 is D; X24 is Q; X27 is L; X28 is D; X29 is T; and X31, X32, X33, and X34 do not exist; The side chain amino group of K at position X10 ( The amino group is partially acylated using the following formula:

[0058] In another embodiment, the polypeptide of SEQ ID NO: 1 or SEQ ID NO: 2 comprises the following sequence: X2 is Aib; X10 is Y; X12 is either K or I; X13 is Y, nor-V, nor-L, or αMe-L; X16 is E; X17 is K; X18 is K; X20 is K; X21 is E; X24 is E; X27 is L; X28 is E; X29 is G; X30 is G X31 is P; X32 is S; X33 is S; and X34 is G; Among them, the side chain amino group of K at position X20 ( The amino group is partially acylated using the following formula:

[0059] In another respect, this disclosure provides a polypeptide comprising the following amino acid sequence or a pharmaceutically acceptable salt thereof: H-X2-QGTFTSD-X10-SEYLDSERARDFVAWLEAGG (SEQ IDNO: 3) in: X2 is S, DS(OMe), or Aib; and X10 is K; The acid group of the C-terminal amino acid is either a free carboxylic acid group or is amidated to a C-terminal primary amide; and The side chain amino group of K at position X10 ( The amino group is partially acylated using the following formula:

[0060] The condition is that the polypeptide is not SEQ ID NO: 6.

[0061] In one embodiment of the polypeptide of SEQ ID NO: 3, X2 is S; The side chain amino group of K at position X10 ( The amino group is partially acylated using the following formula:

[0062] In another embodiment of the polypeptide of SEQ ID NO: 3, X2 is D-Ser(OMe); The side chain amino group of K at position X10 ( The amino group is partially acylated by the following formula:

[0063] In another embodiment of the polypeptide of SEQ ID NO: 3, X2 is Aib; The side chain amino group of K at position X10 ( The amino group is partially acylated using the following formula:

[0064] In another respect, this disclosure provides a polypeptide comprising the following amino acid sequence or a pharmaceutically acceptable salt thereof: H-Aib-QGTFTSDYS-X12-X13-LDEKKA-X20-EFVEWLLEGPSSG (SEQ ID NO: 4) in: X12 is either K or I; X13 is either Y or nor-V; and X20 is K; The acid group of the C-terminal amino acid is either a free carboxylic acid group or is amidated to a C-terminal primary amide; and The side chain amino group of K at position X20 ( The amino group is partially acylated using the following formula:

[0065] In one embodiment of the polypeptide of SEQ ID NO: 4, X12 is K; and X13 is Y; The side chain amino group of K at position X20 ( The amino group is partially acylated using the following formula:

[0066] In another embodiment of the polypeptide of SEQ ID NO: 4, X12 is I; and X13 is NOR-V; The side chain amino group of K at position X20 ( The amino group is partially acylated by the following formula:

[0067] In another embodiment of the polypeptide of SEQ ID NO: 4, X12 is K; and X13 is NOR-V; The side chain amino group of K at position X20 ( The amino group is partially acylated by the following formula:

[0068] In another respect, this disclosure provides a polypeptide comprising the following amino acid sequence or a pharmaceutically acceptable salt thereof: H-(DSer)-QGTFTSD-X10-SKYLDARAAQDFVQWLLDT (SEQID NO: 5) Where X10 is K; The acid group of the C-terminal amino acid is either a free carboxylic acid group or is amidated to a C-terminal primary amide; and The side chain amino group of K at position X10 ( The amino group is partially acylated using the following formula:

[0069] On the other hand, this disclosure provides an incretin analog comprising: Having sequence X 1 The peptide residues of -Gln-Gly-Thr-Phe-Thr-Ser-Asp-Lys (SEQ ID NO: 29), wherein X1 This indicates Aib or Ser(OMe), and wherein the lysine contains a fatty acid elongation group attached to the ε-nitrogen of the lysine; and Gly-Gly-OH peptide residues indirectly linked to the carboxyl group of the lysine.

[0070] In one embodiment, the incretin analog has a lysine residue linked to a Gly-Gly-OH residue via a peptide residue comprising 18 amino acids.

[0071] On the other hand, this disclosure provides an incretin analog comprising: Having sequence X 1 The peptide residues of -Gln-Gly-Thr-Phe-Thr-Ser-Asp-Lys (SEQ ID NO: 29), wherein X 1 This represents Aib or Ser(OMe), and wherein the lysine contains a group of formula (I) linked to the ε-nitrogen of the lysine. (I) in: U does not exist or represents –C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-}, where} is the connection point with W; W indicates: –C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-], -C(O)-NH-(CH2)3-4-NH-], -C(O)-C(CH3)2-NH-] or , Where ] is the connection point with Y; Y does not exist or represents –C(O)-(CH2)2-CH(CO2H)NH-- or -C(O)CH((CH2) x CO2H)NH--, where x is 1, 2 or 3, and -- is the connection point with Z; and Z represents –C(O)-(CH2) n -COOH or –C(O)-(CH2) n -CH3, where n is an integer from 14 to 20; and Gly-Gly-OH peptide residues indirectly linked to the carboxyl group of the lysine.

[0072] In one embodiment, the incretin analog has a lysine residue linked to a Gly-Gly-OH residue via a peptide residue comprising 18 amino acids.

[0073] On the other hand, this disclosure provides an incretin analog comprising: Peptide residues having the sequence Aib-Gln-Gly-Thr-Phe-Thr-Ser-Asp (SEQ ID NO: 30); Lysine residues indirectly linked to the carboxyl group of the residue Aib-Gln-Gly-Thr-Phe-Thr-Ser-Asp (SEQ ID NO: 30), wherein the lysine contains a group of formula (I) linked to the ε-nitrogen of lysine; (I) in: U does not exist or represents –C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-}, where} is the connection point with W; W indicates –C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-]or -C(O)-NH-(CH2)3-4-NH-], -C(O)-C(CH3)2-NH-], Where ] is the connection point with Y; Y does not exist or represents –C(O)-(CH2)2-CH(CO2H)NH-- or -C(O)CH((CH2) x CO2H)NH--, where x is 1, 2 or 3, and -- is the connection point with Z; and Z represents –C(O)-(CH2) n -COOH or –C(O)-(CH2) n -CH3, where n is an integer from 14 to 20; and Gly-Gly-Pro-Ser-Ser-Gly-CONH2 peptide residues indirectly linked to the carboxyl group of the lysine.

[0074] In one embodiment, the incretin analog has a lysine residue linked to the Aib-Gln-Gly-Thr-Phe-Thr-Ser-Asp (SEQ ID NO: 30) residue via a peptide residue comprising 10 amino acids.

[0075] On the other hand, the peptides described herein do not include the peptides of SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8 and SEQ ID NO: 27.

[0076] On the other hand, this disclosure provides a polypeptide comprising an amino acid sequence selected from the group consisting of: i.) HSQGTFTSDK SEYLDSERARDFVAWLEAGG-OH (SEQ ID NO: 9); ii.) H-(DS(OMe))-QGTFTSDK SEYLDSERARDFVAWLEAGG-OH (SEQ ID NO: 10); iii.) H-Aib-SQGTFTSDK SEYLDSERARDFVAWLEAGG-OH (SEQ ID NO: 11); iv.) H-(DS)-QGTFTSDK SKYLDARAAQDFVQWLLDT-NH2 (SEQ ID NO: 12); v.) H-Aib-QGTFTSDYSKYLDEKKAK EFVEWLLEGPSSG-NH2 (SEQ ID NO: 13); vi.) H-Aib-QGTFTSDYSK-(nor-V)-LDEKKAK EFVEWLLEGGPSSG-NH2 (SEQ ID NO:14); and vii.) H-Aib-QGTFTSDYSI-(nor-V)-LDEKKAK EFVEWLLEGPSSG-NH2 (SEQ ID NO:15), Where K The side chain amino group ( The amino group is partially acylated using the following formula:

[0077] The polypeptides described herein are not selected from SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8 or SEQ ID NO: 27.

[0078] The sequences of the polypeptides described herein are represented by single-letter or three-letter codes of amino acids approved by the International Union of Pure and Applied Chemistry (IUPAC).

[0079] Unless otherwise stated, this disclosure is intended to cover L and D isomers of the amino acids in the sequences described herein. However, in some preferred embodiments, all amino acids are in the “L” configuration unless otherwise stated.

[0080] In another respect, this disclosure provides a polypeptide or a pharmaceutically acceptable salt thereof selected from one of the representative compounds in Table 1.

[0081] Table 1. Representative polypeptide compounds Unless otherwise stated, all amino acids mentioned in Table 1 above are in the “L” configuration.

[0082] Table 2. Structure of Parts A, B, C, D, E, F, G, H, and I

[0083] On the other hand, this disclosure provides a polypeptide selected from:

[0084] (SEQ ID NO: 18);

[0085] (SEQ ID NO: 21);

[0086] (SEQ ID NO: 22); and

[0087] (SEQ ID NO: 23).

[0088] On the other hand, this disclosure provides pharmaceutical compositions comprising a pharmaceutically acceptable carrier and an incretin analog or polypeptide as described herein.

[0089] In another aspect, this disclosure provides a method for treating obesity, type 2 diabetes mellitus (T2DM), metabolic syndrome, metabolic dysfunction-associated fatty liver disease (MASLD), metabolic dysfunction-associated steatohepatitis (MASH), fibrosis, cardiovascular disease, and / or hyperlipidemia / dyslipidemia, the method comprising administering an incretin analog or peptide as described herein to a patient in need of such treatment.

[0090] On the other hand, this disclosure provides a method for treating or preventing type 2 diabetes mellitus (T2DM).

[0091] On the other hand, this disclosure provides a method for treating or preventing hyperlipidemia / dyslipidemia.

[0092] On the other hand, this disclosure provides a method for treating or preventing obesity.

[0093] In another aspect, this disclosure provides a method for treating or preventing metabolic syndrome, non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH), neurodegenerative diseases, fibrosis, and / or cardiovascular risks.

[0094] In one implementation, the treatment method includes administering an effective amount of the polypeptide or a pharmaceutically acceptable salt thereof, as described herein, to a patient in need.

[0095] In another aspect, this disclosure provides a method for treating type 2 diabetes mellitus (T2DM), the method comprising administering to a patient in need of such treatment an effective amount of a polypeptide or a pharmaceutically acceptable salt thereof as described herein.

[0096] In another respect, this disclosure provides a method for treating obesity, the method comprising administering to a patient in need of such treatment an effective amount of a polypeptide or a pharmaceutically acceptable salt thereof as described herein.

[0097] In another respect, this disclosure provides a method for treating hyperlipidemia / dyslipidemia, the method comprising administering to a patient in need of such treatment an effective amount of a polypeptide or a pharmaceutically acceptable salt thereof as described herein.

[0098] In another aspect, this disclosure provides a pharmaceutical composition comprising one or more of a polypeptide or a pharmaceutically acceptable salt thereof as described herein, and a pharmaceutically acceptable carrier, diluent, or excipient.

[0099] The compounds of the present invention are preferably formulated into pharmaceutical compositions for administration via parenteral routes (e.g., subcutaneous, intravenous, intraperitoneal, intramuscular, or transdermal). Such pharmaceutical compositions and methods of their preparation are well known in the art. See , for example, "Remington: The Science and 50 Practice of Pharmacy, " DB Troy Edited, 21st Edition Lippincott, Williams & Wilkins , 2006.

[0100] On the other hand, this disclosure provides polypeptides as described herein or pharmaceutically acceptable salts thereof for use as medicines.

[0101] On the other hand, this disclosure provides polypeptides as described herein or pharmaceutically acceptable salts thereof for use in the treatment or prevention of type 2 diabetes mellitus (T2DM).

[0102] On the other hand, this disclosure provides polypeptides or pharmaceutically acceptable salts thereof, as described herein, for use in the treatment or prevention of hyperlipidemia / dyslipidemia.

[0103] On the other hand, this disclosure provides polypeptides or pharmaceutically acceptable salts thereof, as described herein, for use in the treatment or prevention of obesity.

[0104] On the other hand, this disclosure provides for the use of polypeptides or pharmaceutically acceptable salts thereof described herein for the treatment or prevention of diseases selected from the group consisting of: metabolic syndrome, non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH), neurodegenerative diseases, fibrosis, and cardiovascular risks.

[0105] On the other hand, the polypeptides or pharmaceutically acceptable salts thereof described herein may be administered simultaneously, separately or sequentially in combination with effective amounts of one or more other therapeutic agents.

[0106] On the other hand, the pharmaceutical composition according to this disclosure comprises a polypeptide or a pharmaceutically acceptable salt thereof as described herein, used as a medicament.

[0107] On the other hand, the pharmaceutical compositions according to this disclosure comprise a polypeptide or a pharmaceutically acceptable salt thereof as described herein for the treatment or prevention of type 2 diabetes mellitus (T2DM).

[0108] On the other hand, the pharmaceutical compositions according to this disclosure comprise a polypeptide or a pharmaceutically acceptable salt thereof as described herein, used for the treatment or prevention of hyperlipidemia / dyslipidemia.

[0109] On the other hand, the pharmaceutical compositions according to this disclosure comprise a polypeptide or a pharmaceutically acceptable salt thereof, as described herein, used for the treatment or prevention of obesity.

[0110] On the other hand, the pharmaceutical compositions according to this disclosure comprise a polypeptide or a pharmaceutically acceptable salt thereof as described herein for the treatment or prevention of diseases selected from the group consisting of: metabolic syndrome, non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH), neurodegenerative diseases, fibrosis, and cardiovascular risks.

[0111] On the other hand, the pharmaceutical composition according to this disclosure comprises a polypeptide or a pharmaceutically acceptable salt thereof, as described herein, provided simultaneously, separately or sequentially with an effective amount of one or more other therapeutic agents.

[0112] This disclosure may relate to one or more embodiments as described herein. It should be understood that the embodiments described herein are illustrative of this disclosure and are not intended to limit the claims to the specific embodiments exemplified. It should also be understood that the embodiments as defined herein may be used independently or in combination with any definition or any other embodiment as defined herein. Therefore, this disclosure contemplates all possible combinations and permutations of the various independently described embodiments.

[0113] Other features of this disclosure will become apparent to those skilled in the art based on the following embodiments. In general, this disclosure can be extended to any novel features as described herein, including the appended claims and drawings. Therefore, features, integers, properties, compounds, or chemical portions described in conjunction with a particular aspect, embodiment, or example of this disclosure should be understood to be applicable to any other aspect, embodiment, or example as described herein, unless incompatible therewith.

[0114] Furthermore, unless otherwise stated, any feature disclosed herein may be replaced by an alternative feature serving the same or similar purpose.

[0115] Example

[0116] Instruments and analytical methods. The instruments used for characterizing and analyzing the compounds described herein include a high-performance liquid chromatograph (HPLC) (Waters e2695 Alliance; detector: Waters (2489 UV / visible)).

[0117] Mass spectrometry instrument: HPLC: Waters e2695 Alliance; and detector: Acquity-QDa.

[0118] The compounds described herein were purified using the preparative HPLC procedure described below.

[0119] Preparative 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 µm, flow rate: 75 mL / min

[0120] Mobile phase:

[0121] The purity of the compounds described herein was analyzed using one of the RP-HPLC methods described below.

[0122] HPLC Method A

[0123] Column: Xbridge peptide BEH C18 (4.6 mm x 250 mm, 3.5 μm)

[0124] Elution buffer: Mobile phase A: Buffer: Acetonitrile = 900:100 Mobile phase B: Buffer: Acetonitrile = 300:700 Buffer solution: aqueous solution of potassium dihydrogen phosphate, adjusted to pH 3.0 ± 0.1 with orthophosphate. Flow rate: 0.8 mL / min Detection: UV detection at 210 nm Column temperature: 65°C Sample tray temperature: 5°C Running time: 90 minutes

[0125] HPLC Method B

[0126] Column: YMC Pack Pro C18 (4.6 mm x 250 mm, 3.0μ)

[0127] Elution buffer: Mobile phase A: Buffer: Acetonitrile = 900:100 Mobile phase B: Buffer: Acetonitrile = 300:700 Buffer solution: aqueous solution of potassium dihydrogen phosphate, adjusted to pH 3.0 ± 0.1 with orthophosphate. Flow rate: 1.0 mL / min Detection: UV detection at 210 nm Column temperature: 50°C Sample tray temperature: 5°C Running time: 38 minutes

[0128] HPLC method C

[0129] Column: X-Select CSH C18, 130 A°, 2.5 µm, (4.6 X 150) mm

[0130] Elution buffer: Mobile phase A: Buffer: Acetonitrile = 900:100 Mobile phase B: Buffer: Acetonitrile = 300:700 Buffer solution: potassium dihydrogen phosphate aqueous solution; trimethylamine added; pH adjusted to 2.5 ± 0.1 with orthophosphate. Flow rate: 0.5 mL / min Detection: UV detection at 214 nm Column temperature: 60°C Sample tray temperature: 5°C Running time: 90 minutes

[0131] HPLC method D

[0132] Column: X-Select CSH C18, 130 A°, 2.5 µm, (4.6 X 150) mm

[0133] Elution buffer: Mobile phase A: Buffer: Acetonitrile = 900:100 Mobile phase B: Buffer: Acetonitrile = 300:700 Buffer solution: potassium dihydrogen phosphate aqueous solution; trimethylamine added; pH adjusted to 2.5 ± 0.1 with orthophosphate. Flow rate: 0.8 mL / min Detection: UV detection at 210 nm Column temperature: 60°C Sample tray temperature: 5°C Running time: 33 minutes

[0134] Preparation method

[0135] Example A: Part A-II Uncle Preparation of butyl ester

[0137] Partial A-II was prepared using solid-phase synthesis. UncleButyl ester. In the presence of DIPEA, 2-[2-(2-Fmoc-aminoethoxy)ethoxy]acetic acid is linked to 2-chlorotriphenylmethyl chloro resin to produce 2-[2-(2-Fmoc-aminoethoxy)ethoxy]acetic acid-2-chlorotriphenylmethyl-resin. The Fmoc protecting group is removed by selective deblocking with piperidine, followed by coupling with Fmoc-Aib-OH in THF using DIPC and HOBt to produce 2-[2-[2-[(2-Fmoc-amino-2-methyl-propionyl)amino]ethoxy]ethoxy]acetic acid-2-chlorotriphenylmethyl-resin. The Fmoc group is removed by selective deblocking with piperidine, and the free amino group is coupled with Fmoc-Glu-OtBu using HOBt and DIPC to produce 2-[2-[2-[[2-[[(4)] S )-4-Fmoc-amino-5- Uncle Butoxy-5-oxo-pentanoyl]amino]-2-methyl-propionyl]amino]ethoxy]ethoxy]acetic acid-2-chlorotriphenylmethyl-resin. The Fmoc group of the resulting compound was selectively deblocked using piperidine, and then the free amino group was reacted with octadecanoic acid mono-resin. Uncle Butyl ester coupling yields 2-[2-[2-[[2-[[(4)] S )-5- Uncle Butoxy-4-[(18- Uncle Butoxy-18-oxo-octadecanoyl)amino]-5-oxo-pentanoyl]amino]-2-methyl-propionyl]-amino]ethoxy]ethoxy]acetic acid-2-chlorotriphenylmethyl-resin. The intermediate was then cleaved from the 2-chlorotriphenylmethyl-resin using trifluoroethanol:DCM (1:1) to give 2-[2-[2-[[2-[[(4) S )-5- Uncle Butoxy-4-[(18- Uncle [Butoxy-18-oxo-octadecanoyl]amino]-5-oxo-pentanoyl]amino]-2-methyl-propionyl]amino]ethoxy]ethoxy]acetic acid (partial A-di) Uncle Butyl ester). LCMS = m / z: 786.39 (M+H) + ).

[0138] Preparation of part of A-OSu

[0139] Then, in the presence of dicyclohexylcarbodiimide (DCC), the resulting A-di Uncle Butyl ester reacts with HOSu to produce a succinimide-protected intermediate, which is then deprotected with trifluoroacetic acid to produce the title compound A-OSu.

[0140] Example B: Part B-II Uncle Preparation of butyl ester

[0141] Part B-II was prepared using a method similar to that given in Example A. Uncle Butyl ester, wherein 20-( Uncle (Butoxy)-20-oxoeicosanoic acid replacing octadecanoic acid monobutylene oxide Uncle Butyl ester, yielding 2-[2-[2-[[2-[[(4) S )-5- Uncle Butoxy-4-[(20- Uncle Butoxy-20-oxo-eicosanoyl)amino]-5-oxo-pentanoyl]amino]-2-methyl-propionyl]amino]ethoxy]ethoxy]acetic acid-2-chlorotriphenylmethyl-resin. This intermediate is then cleaved from the 2-chlorotriphenylmethyl-resin using trifluoroethanol:DCM (1:1) to give 2-[2-[2-[[2-[[(4)] S )-5- Uncle Butoxy-4-[(20- Uncle [Butoxy-20-oxo-eicosanoyl]amino]-5-oxo-pentanoyl]amino]-2-methyl-propionyl]amino]ethoxy]ethoxy]acetic acid (partially β-di) Uncle Butyl ester). LCMS = m / z: 814.10 (M+H) + ).

[0142] Preparation of some B-OSu

[0143] Then, in the presence of dicyclohexylcarbodiimide (DCC), the resulting partial B-di Uncle Butyl ester reacts with HOSu to produce a succinimide-protected intermediate, which is then deprotected with trifluoroacetic acid to produce the title compound, part B-OSu.

[0144] Example C: Part C-II Uncle Preparation of butyl ester

[0145] Partial C-II was prepared using solid-phase synthesis. Uncle Butyl ester. In the presence of DIPEA, 2-[2-(2-Fmoc-aminoethoxy)ethoxy]acetic acid is linked to a 2-chlorotriphenylmethyl chlororesin to produce 2-[2-(2-Fmoc-aminoethoxy)ethoxy]acetic acid-2-chlorotriphenylmethyl-resin. The Fmoc protecting group is removed by selectively deblocking the amino group using piperidine, and then... rightNitrophenyl chloroformate activates the free amino group in THF and DIPEA, followed by reaction with Fmoc-aminobutyramine hydrochloride in THF:DMAc and DIPEA to produce 2-[2-[2-(4-Fmoc-aminobutyrcarbamoylamino)ethoxy]ethoxy]acetic acid-2-chlorotriphenylmethyl-resin. The Fmoc group is removed by selective deblocking with piperidine, and then the free amino group is coupled to Fmoc-Glu-OtBu using HOBt and DIPC to produce 2-[2-[2-[4-[[(4-aminobutyrcarbamoylamino)ethoxy]ethoxy]acetic acid-2-chlorotriphenylmethyl-resin. S )-4-Fmoc-amino-5- Uncle Butoxy-5-oxo-pentanoyl]amino]butylcarbamoylamino]ethoxy]ethoxy]acetic acid-2-chlorotriphenylmethyl-resin. The resulting 2-[2-[2-[4-[[(4)] resin was selectively prepared using piperidine. S )-4-Fmoc-amino-5- Uncle Butoxy-5-oxo-pentanoyl]amino]-butylcarbamoylamino]ethoxy]ethoxy]acetic acid-2-chlorotriphenylmethyl- resin deblocking, then with octadecanoic acid mono- Uncle Butyl ester coupling yields intermediate 2-[2-[2-[4-[[(4)] S )-5- Uncle Butoxy-4-[(18-tert-butoxy-18-oxo-octadecanoyl)amino]-5-oxo-pentanoyl]amino]butylcarbamoylamino]ethoxy]ethoxy]acetic acid-2-chlorotriphenylmethyl-resin. The intermediate was then cleaved from the 2-chlorotriphenylmethyl-resin using trifluoroethanol:DCM (1:1) to give 2-[2-[2-[4-[[(4 S )-5- Uncle Butoxy-4-[(18- Uncle [Butoxy-18-oxo-octadecanoyl]amino]-5-oxo-pentanoyl]amino]butylcarbamoylamino]ethoxy]ethoxy]acetic acid (partial C-di) Uncle Butyl ester). LCMS = m / z: 814.56 (M+H) + ).

[0146] Preparation of some C-OSu

[0147] Some C-OSu

[0148] Then, in the presence of dicyclohexylcarbodiimide (DCC), the resulting C-di... Uncle Butyl ester reacts with HOSu to produce a succinimide-protected intermediate, which is then deprotected with trifluoroacetic acid to produce the title compound moiety C-OSu.

[0149] Example D: Part D-II Uncle Preparation of butyl ester

[0150] Part D2 Uncle Butyl acetate

[0151] Part B-II was prepared using a method similar to that given in Example C. Uncle Butyl ester, wherein 20-( Uncle (Butoxy)-20-oxoeicosanoic acid replacing octadecanoic acid monobutylene oxide Uncle Butyl ester, yielding intermediate 2-[2-[2-[4-[[5- Uncle Butoxy-4-[(20- Uncle [Butoxy-20-oxo-eicosanoyl]amino]-5-oxo-pentanoyl]amino]butylcarbamoylamino]ethoxy]ethoxy]acetic acid-2-chlorotriphenylmethyl-resin. The intermediate was then cleaved from the 2-chlorotriphenylmethyl-resin using trifluoroethanol:DCM (1:1) to give 2-[2-[2-[4-[[5- Uncle Butoxy-4-[(20- Uncle [Butoxy-20-oxo-eicosanoyl]amino]-5-oxo-pentanoyl]amino]butylcarbamoylamino]ethoxy]ethoxy]acetic acid (partial D-di) Uncle Butyl ester). LCMS = m / z: 843.14 (M+H) + ).

[0152] Preparation of part of D-OSu

[0153] Some D-OSu

[0154] Then, in the presence of dicyclohexylcarbodiimide (DCC), the resulting partial D-di... Uncle Butyl ester reacts with HOSu to produce a succinimide-protected intermediate, which is then deprotected with trifluoroacetic acid to produce the title compound, part D-OSu.

[0155] Example E: Preparation of a portion of E-OSu

[0156] Part E-OSu

[0157] L-glutamic acid α- Uncle Butyl ester (H-Glu-OtBu) reacts with palmitic acid in the presence of IBCF and NMM to produce CH3-(CH2). 14-C(O)-Glu-OtBu, then reacted with HOSu in the presence of IBCF and NMM to produce CH3-(CH2). 14 -C(O)-Glu(OSu)-O t Bu, then deprotected with trifluoroacetic acid to produce partial E-OSu.

[0158] Example F: Part F-II Uncle Preparation of butyl ester

[0159] Partial F-2 Uncle Butyl acetate

[0160] Partial F-II was prepared using solid-phase synthesis. Uncle Butyl ester. In the presence of DIPEA, 2-[2-(2-Fmoc-aminoethoxy)ethoxy]acetic acid is linked to a 2-chlorotriphenylmethyl chloro resin to produce 2-[2-(2-Fmoc-aminoethoxy)ethoxy]acetic acid-2-chlorotriphenylmethyl resin. The Fmoc protecting group is removed by selectively deblocking the amino group with piperidine, followed by coupling with 2-[2-(2-Fmoc-aminoethoxy)ethoxy]acetic acid in THF using DIPC and HOBt to produce {(Fmoc-aminoethoxy)-ethoxy}-acetyl-{(-aminoethoxy)-ethoxy}-acetic acid-2-chlorotriphenylmethyl resin. By selectively deblocking the Fmoc group with piperidine and coupling the free amino group with Fmoc-Glu-OtBu using HOBt and DIPC, Fmoc-Glu({(amino-ethoxy)-ethoxy}-acetyl-{(-amino-ethoxy)-ethoxy}-acetic acid-2-chlorotriphenylmethyl-resin)-OtBu is generated. The Fmoc group of the resulting compound is then selectively deblocked with piperidine, and the free amino group is then coupled with octadecanoic acid monoester. Uncle Butyl ester coupling yields 2-[2-[2-[[2-[2-[2-[5-] Uncle Butoxy-4-[(18- Uncle Butoxy-18-oxo-octadecanoyl)amino]-5-oxo-pentanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetic acid-2-chlorotriphenylmethyl-resin. The intermediate was then cleaved from the 2-chlorotriphenylmethyl-resin using trifluoroethanol:DCM (1:1) to give 2-[2-[2-[[2-[2-[2-[5- Uncle Butoxy-4-[(18- Uncle [Butoxy-18-oxo-octadecanoyl]amino]-5-oxo-pentanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetic acid (partial F-di) UncleButyl ester). LCMS = m / z: 846.10 (M+H) + ).

[0161] Preparation of part of F-OSu

[0162] Some F-OSu

[0163] Then, in the presence of dicyclohexylcarbodiimide (DCC), the resulting partial F-di... Uncle Butyl ester reacts with HOSu to produce a succinimide-protected intermediate, which is then deprotected with trifluoroacetic acid to produce the title compound, part F-OSu.

[0164] Example G: Preparation Part G

[0165] Part G-2 Uncle Butyl acetate

[0166] Partial G-II was prepared using a method similar to that given in Example F. Uncle Butyl ester, wherein 20-( Uncle (Butoxy)-20-oxoeicosanoic acid replacing octadecanoic acid monobutylene oxide Uncle Butyl ester was used to give the intermediate 2-[2-[2-[[2-[2-[2-[[5-tert-butoxy-4-[(20-tert-butoxy-20-oxo-eicosanoyl)amino]-5-oxo-pentanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetic acid-2-chlorotriphenylmethyl-resin. The intermediate was then cleaved from the 2-chlorotriphenylmethyl-resin using trifluoroethanol:DCM (1:1) to give 2-[2-[2-[[2-[2-[2-[[5-tert-butoxy-4-[(20-tert-butoxy-20-oxo-eicosanoyl)amino]-5-oxo-pentanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetic acid (partially G-di- Uncle Butyl ester). LCMS = m / z: 874.15 (M+H) + ).

[0167] Preparation of part of G-OSu

[0168] Some G-OSu

[0169] Then, in the presence of dicyclohexylcarbodiimide (DCC), the resulting G-di... Uncle Butyl ester reacts with HOSu to produce a succinimide-protected intermediate, which is then deprotected with trifluoroacetic acid to produce the title compound, G-OSu.

[0170] Example H: Preparation of partial H-di Uncle Butyl acetate

[0171] Partial H-2 Uncle Butyl acetate

[0172] Partial H-dihydrochloride was prepared using solid-phase synthesis with 2-chlorotriphenylmethyl chloride resin. Uncle Butyl ester. In the presence of DIPEA, 2-[2-(2-Fmoc-aminoethoxy)ethoxy]acetic acid is linked to a 2-chlorotriphenylmethyl chloro resin to produce 2-[2-(2-Fmoc-aminoethoxy)ethoxy]acetic acid-2-chlorotriphenylmethyl-resin. The Fmoc protecting group is removed by selectively deblocking the amino group with piperidine, followed by coupling with Fmoc-Glu-OtBu using HOBt and DIPC to produce 2-[2-[2-[[(4S)-5- Uncle Butoxy-4-(9 H [fluorene-9-ylmethoxycarbonylamino]-5-oxo-pentanoyl]amino]ethoxy]ethoxy]acetic acid-2-chlorotriphenylmethyl-resin. The Fmoc group of the resulting compound was selectively deblocked using piperidine, and then the free amino group was reacted with octadecanoic acid mono- Uncle Butyl ester coupling yields 2-[2-[2-[[(4S)-5- Uncle Butoxy-4-[(18-tert-butoxy-18-oxo-octadecanoyl)amino]-5-oxo-pentanoyl]amino]ethoxy]ethoxy]acetic acid 2-chlorotriphenylmethyl-resin. The intermediate was then cleaved from the 2-chlorotriphenylmethyl-resin using trifluoroethanol:DCM (1:1) to give 2-[2-[2-[[(4S)-5-tert-butoxy-4-[(18-tert-butoxy-18-oxo-octadecanoyl)amino]-5-oxo-pentanoyl]amino]ethoxy]ethoxy]acetic acid (partially H-diethyl) Uncle Butyl ester). LCMS = m / z: 700.94 (M+H) + ).

[0173] Preparation of part of H-OSu

[0174] Partial H-OSu

[0175] Then, in the presence of dicyclohexylcarbodiimide (DCC), the resulting H-di... Uncle Butyl ester reacts with HOSu to produce a succinimide-protected intermediate, which is then deprotected with trifluoroacetic acid to produce the title compound, part H-OSu.

[0176] Example I: Preparation Part I

[0177] Part I-II Uncle Butyl acetate

[0178] Part I-II was prepared using a method similar to that given in Example H. Uncle Butyl ester, wherein 20-( Uncle (Butoxy)-20-oxoeicosanoic acid replacing octadecanoic acid monobutylene oxide Uncle Butyl ester, yielding intermediate 2-[2-[2-[[(4S)-5-tert-butoxy-4-[(20- Uncle Butoxy-20-oxo-eicosanoyl)amino]-5-oxo-pentanoyl]amino]ethoxy]ethoxy]acetic acid 2-chlorotriphenylmethyl-resin. The intermediate was then cleaved from the 2-chlorotriphenylmethyl-resin using trifluoroethanol:DCM (1:1) to give 2-[2-[2-[[(4S)-5-tert-butoxy-4-[(20- Uncle [Butoxy-20-oxo-eicosanoyl]amino]-5-oxo-pentanoyl]amino]ethoxy]ethoxy]acetic acid (partial I-di) Uncle Butyl ester). LCMS = m / z: 728.99 (M+H) + ).

[0179] Preparation of part of I-OSu

[0180] Part I-OSu

[0181] Then, in the presence of dicyclohexylcarbodiimide (DCC), the resulting partial I-di Uncle Butyl ester reacts with HOSu to produce a succinimide-protected intermediate, which is then deprotected with trifluoroacetic acid to produce the title compound, part I-OSu.

[0182] Example 1: Synthesis of Compound 1

[0183] Part A: Synthesis of the linear peptide backbone

[0184] The peptide backbone was synthesized via a solid-phase method. Wang resin was used as the starting resin. Fmoc-protected Gly-OH groups were used for coupling with the Wang resin. The synthesis was carried out using diisopropylcarbodiimide in the presence of 4-dimethylaminopyridine (DMAP). N1,3-hydroxybenzotriazole (DIC-HOBt) was used as a coupling agent to generate Fmoc-Gly-Wang resin. Acetic anhydride and diisopropylethylamine / pyridine were used to terminate uncoupled amino groups at each amino acid coupling site. Piperidine was used to selectively de-block the amino groups of the Fmoc-Gly-Wang resin, followed by coupling with HOBt and DIPC to Fmoc-Gly-OH to generate Fmoc-Gly-Gly-Wang resin. This completed one cycle.

[0185] For the remaining 28 amino acid residues, repeat the above three steps: selective end-capping, selective deblocking of the Fmoc protection of the resin-linked amino acid, and sequential coupling of the next amino acid residue with the Fmoc-protected amino group. Selective deblocking (deprotection of the Fmoc group) is performed using piperidine, and coupling with the next Fmoc-protected amino acid is performed using HOBt / DIPC. The side chains of the Fmoc-protected amino acids are orthogonally protected (the hydroxyl groups of serine, tyrosine, or threonine are protected with tert-butyl (-) groups). t The amino and guanidinyl groups of lysine and arginine are protected with tert-butoxycarbonyl (-Boc) and 2,2,4,6,7-pentamethyldihydrobenzofuran-5-sulfonyl (-Pbf) groups, respectively; the imidazole group of histidine is protected with tert-butoxycarbonyl (-Boc); the carboxylic acid group of aspartic acid or glutamic acid is protected with (-tBu) group; and the amide group of glutamine is protected with triphenylmethyl (-Trt) group. Perform the above two steps: selectively deblock, and then couple with the next Fmoc-protected amino acid to obtain Fmoc-His(Boc)-Ser(tBu)-Gln(Trt)-Gly-Thr(tBu)-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Lys(Boc)-Ser(tBu)-Glu(OtBu)-Tyr(tBu)-Leu-Asp(OtBu)-Ser(tBu)-Glu(OtBu)-Arg(Pbf)-Ala-Arg(Pbf)-Asp(OtBu)-Phe-Val-Ala-Trp-Leu-Glu(OtBu)-Ala-Gly-Gly- resin.

[0186] Piperidine was used to treat Fmoc-His(Boc)-Ser(tBu)-Gln(Trt)-Gly-Thr(tBu)-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Lys(Boc)-Ser(tBu)-Glu(OtBu)-Tyr(tBu)-Leu-Asp(OtBu)-Ser(tBu)-Glu(OtBu)-Arg(Pb) f)-Ala-Arg(Pbf)-Asp(OtBu)-Phe-Val-Ala-Trp-Leu-Glu(OtBu)-Ala-Gly-Gly-resin was deblocked, followed by cleavage and deprotection with trifluoroacetic acid, ethane-1,2-dithiol, and triisopropylsilane to obtain the crude product H-His-Ser-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Lys( -NH2)-Ser-Glu-Tyr-Leu-Asp-Ser-Glu-Arg-Ala-Arg-Asp-Phe-Val-Ala-Trp-Leu-Glu-Ala-Gly-Gly-OH was purified by preparative HPLC.

[0187] Part B: Grafting of activated fatty acid chains onto lining peptides

[0188] The activated fatty acid chain moiety C-Osu was grafted onto the purified linear peptide obtained from moiety A in water:acetonitrile at approximately pH 11 to produce crude title peptide compound 1, which was purified by preparative HPLC.

[0189] Mass spectrometry (LCMS): m / z = 1012.36 (MH4 4+); calculated mass = 4045.40; and HPLC purity (Method B): 94.48%.

[0190] Example 2: Synthesis of Compound 2

[0191] Part A: Synthesis of the linear peptide backbone

[0192] The linear peptide backbone of compound 2 was prepared by a solid-phase method according to a similar method given in Part A of Example 1, wherein Fmoc-D-Ser(OMe)-OH was used instead of Fmoc-Ser(tBu)-OH at position 2.

[0193] Part B: Grafting of activated fatty acid chains onto lining peptides

[0194] The activated fatty acid chain moiety E-OSu was grafted onto the purified linear peptide obtained in moiety A in a water:acetonitrile solution at approximately pH 11: H-His-D-Ser(OMe)-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Lys( -NH2)-Ser-Glu-Tyr-Leu-Asp-Ser-Glu-Arg-Ala-Arg-Asp-Phe-Val-Ala-Trp-Leu-Glu-Ala-Gly-Gly-OH, yielding crude title peptide compound 2, which was purified by preparative HPLC.

[0195] Mass spectrometry (LCMS): m / z = 936.10 (MH4 4+); calculated mass = 3739.96; and HPLC purity (Method B): 95.05%.

[0196] Example 3: Synthesis of Compound 3

[0197] Part A: Synthesis of the linear peptide backbone

[0198] The linear peptide backbone of compound 3 was prepared by solid-phase method according to a similar method given in Part A of Example 1, wherein Fmoc-Aib-OH was used instead of Fmoc-Ser(tBu)-OH at position 2.

[0199] Part B: Grafting of activated fatty acid chains onto lining peptides

[0200] The activated fatty acid chain moiety E-OSu was grafted onto the purified linear peptide obtained from moiety A in a water:acetonitrile solution at approximately pH 11: H-His-Aib-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Lys( -NH2)-Ser-Glu-Tyr-Leu-Asp-Ser-Glu-Arg-Ala-Arg-Asp-Phe-Val-Ala-Trp-Leu-Glu-Ala-Gly-Gly-OH, yielding crude title peptide compound 3, which was purified by preparative HPLC.

[0201] Mass spectrometry (LCMS): m / z = 932.13 (MH4 4+); calculated mass = 3724.48; and HPLC purity (Method B): 96.1%.

[0202] Example 4: Synthesis of Compound 4

[0203] Part A: Synthesis of the linear peptide backbone

[0204] The linear peptide backbone of compound 4 was prepared by a solid-phase method, following a similar method to that given in Part A of Example 3.

[0205] Part B: Grafting of activated fatty acid chains onto lining peptides

[0206] The activated fatty acid chain moiety A-OSu was grafted onto a purified linear peptide obtained from moiety A in a water:acetonitrile solution at approximately pH 11: H-His-Aib-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Lys( -NH2)-Ser-Glu-Tyr-Leu-Asp-Ser-Glu-Arg-Ala-Arg-Asp-Phe-Val-Ala-Trp-Leu-Glu-Ala-Gly-Gly-OH, yielding crude title peptide compound 4, which was purified by preparative HPLC.

[0207] Mass spectrometry (LCMS): m / z = 1004.34 (MH4 4+); calculated mass = 4013.33; and HPLC purity (method D): 96.93%.

[0208] Example 5: Synthesis of Compound 5

[0209] Part A: Synthesis of the linear peptide backbone

[0210] The linear peptide backbone of compound 4 was prepared by solid-phase method following a similar method given in Part A of Example 1.

[0211] Part B: Grafting of activated fatty acid chains onto lining peptides

[0212] The activated fatty acid chain moiety A-OSu was grafted onto the purified linear peptide obtained from moiety A in a water:acetonitrile solution at approximately pH 11: H-His-Ser-Gln-Gly-Thr-Phe-Thr-Ser-Asp-Lys( -NH2)-Ser-Glu-Tyr-Leu-Asp-Ser-Glu-Arg-Ala-Arg-Asp-Phe-Val-Ala-Trp-Leu-Glu-Ala-Gly-Gly-OH, yielding crude title peptide compound 5, which was purified by preparative HPLC.

[0213] Mass spectrometry (LCMS): m / z = 1004.96 (MH4 4+); calculated mass = 4015.81; and HPLC purity (Method B): 97.33%.

[0214] Example 6: Synthesis of Compound 6

[0215] Part A: Synthesis of the linear peptide backbone

[0216] Compound 6 was synthesized via a solid-state method. The starting resin used in the synthesis was a Fmoc-Rink amide resin. Piperidine was used to selectively deblock the Fmoc-protected amino groups of the Rink amide resin, followed by coupling of Fmoc-Gly-OH with the Rink amide resin. This was achieved by using diisopropylcarbodiimide... N 1-Hydroxybenzotriazole (DIPC-HOBt) was used as a coupling agent to generate Fmoc-Gly-Rink amide resin, completing the first cycle. Acetic anhydride and diisopropylethylamine were used to terminate / cap uncoupled amino groups at each amino acid coupling point. Piperidine was used to selectively de-block the amino groups of the Fmoc-Gly-Rink amide resin, and then HOBt and DIPC were used to couple Fmoc-Ser(tBu)-OH to generate Fmoc-Ser(tBu)-Gly-Rink amide resin, completing the second cycle.

[0217] For the remaining 32 amino acid residues, repeat the above three steps: end capping, selectively decapping the Fmoc protection of the amino acid linked to the resin, and sequentially coupling the next amino acid residue to the Fmoc-protected amino group. The side chains of the Fmoc-protected amino acids are orthogonally protected (the hydroxyl groups of serine, tyrosine, or threonine are protected using...). Uncle Butyl (-tBu) group protection, lysine amino group protected by... Uncle The butoxycarbonyl (-Boc) and (4,4-dimethyl-2,6-dioxocyclohexyl-1-ylidene)-3-methylbutyl (IVDde) groups are protected, and the carboxylic acid group of aspartic acid or glutamic acid is protected with (- t The amide group of glutamine is protected by a triphenylmethyl (-Trt) group. Perform the above three steps: selective end-capping, de-blocking, and then coupling with the next Fmoc-protected amino acid to obtain Fmoc-His(Boc)-Aib-Gln(Trt)-Gly-Thr(tBu)-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Tyr(tBu)-Ser(tBu)-Lys(Boc)-Tyr(tBu)-Leu-Asp(OtBu)-Glu(OtBu)-Lys(Boc)-Lys(Boc)-Ala-Lys(IVDde)-Glu(OtBu)-Phe-Val-Glu(OtBu)-Trp-Leu-Leu-Glu(OtBu)-Gly-Gly-Pro-Ser(tBu)-Ser(tBu)-Gly-resin.

[0218] Piperidine was used to deblock Fmoc-His(Boc)-Aib-Gln(Trt)-Gly-Thr(tBu)-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Tyr(tBu)-Ser(tBu)-Lys(Boc)-Tyr(tBu)-Leu-Asp(OtBu)-Glu(OtBu)-Lys(Boc)-Lys(Boc)-Ala-Lys(IVDde)-Glu(OtBu)-Phe-Val-Glu(OtBu)-Trp-Leu-Leu-Glu(OtBu)-Gly-Gly-Pro-Ser(tBu)-Ser(tBu)-Gly- resin. The obtained peptide-resin was protected with Boc anhydride to produce Boc-His(Boc)-Aib-Gln(Trt)-Gly-Thr(tBu)-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Tyr(tBu)-Ser(tBu)-Lys(Boc)-Tyr(tBu)-Leu-Asp(OtBu)-Glu(OtBu)-Lys(Boc)-Lys(Boc)-Ala-Lys(IVDde)-Glu(OtBu)-Phe-Val-Glu(OtBu)-Trp-Leu-Leu-Glu(OtBu)-Gly-Gly-Pro-Ser(tBu)-Ser(tBu)-Gly-resin. The IVDde groups of the obtained peptide-resin were deprotected using hydrazine hydrate to produce a linear peptide-resin: Boc-His(Boc)-Aib-Gln(Trt)-Gly-Thr(tBu)-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Tyr(tBu)-Ser(tBu)-Lys(Boc)-Tyr(tBu)-Leu-Asp(OtBu)-Glu(OtBu)-Lys(Boc)-Lys(Boc)-Ala-Lys( -NH2)-Glu(OtBu)-Phe-Val-Glu(OtBu)-Trp-Leu-Leu-Glu(OtBu)-Gly-Gly-Pro-Ser(tBu)-Ser(tBu)-Gly-resin.

[0219] Part B: Grafting of activated fatty acid chains onto lining peptides

[0220] Using diisopropylcarbodiimide, N -Hydroxybenzotriazole (DIPC-HOBt) was used as a coupling agent to partially bind β-dihydroxybenzotriazole. UncleButyl ester was coupled onto a linear peptide-resin obtained in part A to produce compound 6-resin. The resin was cleaved and deprotected using trifluoroacetic acid with ethane-1,2-dithiol and triisopropylsilane, followed by preparative HPLC purification to obtain pure compound 6.

[0221] Mass spectrometry (LCMS): m / z = 1126.36 (MH4 4+); calculated mass = 4501.41; and HPLC purity (Method A): 97.16%.

[0222] Example 7: Synthesis of Compound 7

[0223] Compound 7 was prepared by a solid-state method following a similar approach to that given in Example 6, wherein after deprotection of IVDde, it is partially A-di Uncle Coupling of butyl ester, rather than partial β-diethyl ester. Uncle Butyl ester coupling.

[0224] Mass spectrometry (LCMS): m / z = 1120.09 (MH4 4+); calculated mass = 4476.33; and HPLC purity (Method A): 98.95%.

[0225] Example 8: Synthesis of Compound 8

[0226] Compound 8 was prepared by a solid-state method following a similar approach to that given in Example 6, wherein after deprotection of IVDde, it is partially D-dioxanone. Uncle Coupling of butyl ester, rather than partial β-diethyl ester. Uncle Butyl ester coupling.

[0227] Mass spectrometry (LCMS): m / z = 1134.05 (MH4 4+) and calculated mass = 4532.17.

[0228] Example 9: Synthesis of Compound 9

[0229] Compound 9 was prepared by a solid-state method following a similar approach to that given in Example 6, wherein after deprotection of IVDde, it is partially C-di Uncle Coupling of butyl ester, rather than partial β-diethyl ester. Uncle Butyl ester coupling.

[0230] Mass spectrometry (LCMS): m / z = 1127.22 (MH4 4+) and calculated mass = 4504.85.

[0231] Example 10: Synthesis of Compound 10

[0232] Compound 9 was prepared by solid-phase method following a similar method as given in Example 6, wherein Fmoc-n-valine-OH was used instead of Fmoc-Tyr(tBu)-OH at position 13, and after IVDde deprotection, it was partially A-di Uncle Coupling of butyl ester, rather than partial β-diethyl ester. Uncle Butyl ester coupling.

[0233] Mass spectrometry (LCMS): m / z = 1103.56 (MH4 4+) and calculated mass = 4410.21.

[0234] Example 11: Synthesis of Compound 11

[0235] Compound 11 was prepared by solid-state method according to a similar method given in Example 6, wherein (i) Fmoc-Ile-OH was used instead of Fmoc-Lys(Boc)-OH at position 12, (ii) Fmoc-n-valine-OH was used instead of Fmoc-Tyr(tBu)-OH at position 13, and (iii) after IVDde deprotection, it is partially A-di Uncle Coupling of butyl ester, rather than partial β-diethyl ester. Uncle Butyl ester coupling.

[0236] Mass spectrometry (LCMS): m / z = 1100.11 (MH4 4+) and calculated mass = 4396.41.

[0237] Example 25: Synthesis of Compound 25

[0238] Compound 25 was prepared by a solid-state method following a similar approach to that given in Example 6, wherein after deprotection of IVDde, it is partially F-di Uncle Coupling of butyl ester, rather than partial β-diethyl ester. Uncle Butyl ester coupling.

[0239] Mass spectrometry (LCMS): m / z = 1135.11 (MH4 4+); calculated mass = 4536.41; HPLC purity (Method A): 98.69%.

[0240] Biological research

[0241] Example 1: Oral glucose tolerance test (OGTT) in rats; single injection; 1 mg / kg dose

[0242] Study 1: Animals were divided into four groups (n=4 / group): a normal control group, cotadiopeptide (1 mg / kg), compound 1 (1 mg / kg), and smegglutide (1 mg / kg). Animals were fasted for 12 hours prior to the start of the oral glucose tolerance test (OGTT). Blood glucose levels were measured using a glucometer at 22 and 166 hours after subcutaneous injection of the test compound, cotadiopeptide, and smegglutide (measurement at time 0). All animals were orally administered 2 g / kg of glucose solution. Blood glucose levels were measured at 20, 40, 60, 90, and 120 minutes after the glucose challenge. Body weight and food intake were recorded at 48 and 154 hours.

[0243] Study 2: Animals were divided into four groups (n=4 / group): a normal control group, compound 2 (1 mg / kg), compound 3 (1 mg / kg), and semaglutide (1 mg / kg). Animals were fasted for 12 hours prior to the start of the oral glucose tolerance test (OGTT). Blood glucose levels were measured using a glucometer at 22 and 166 hours after subcutaneous injection of the test compound and semaglutide (measurement at time 0). All animals were orally administered 2 g / kg of glucose solution. Blood glucose levels were measured at 20, 40, 60, 90, and 120 minutes after the glucose challenge. Body weight and food intake were recorded at 48 and 154 hours.

[0244] Table 3. Blood glucose AUC 22 hours after subcutaneous injection (0-120分钟) Changes

[0245] Compared to the normal control p<0.05, p<0.01, p < 0.001; one-way ANOVA was followed by Bonferroni post-test.

[0246] Table 4. Blood glucose AUC 166 hours after subcutaneous injection (0-120分钟) Changes

[0247] Compared to the normal control p<0.05, p<0.01, p < 0.001, and relative to smegglutinin, #p < 0.05, ##p < 0.01 and ###p < 0.001; one-way ANOVA was followed by Bonferroni post-test.

[0248] Table 5. Effects on body weight

[0249] Compared to the normal control p<0.05, p<0.01, p < 0.001, and relative to smegglutinin, #p < 0.05, ##p < 0.01 and ###p < 0.001; one-way ANOVA was followed by Bonferroni post-test.

[0250] Table 6. Effects on food intake

[0251] Compared to the normal control p<0.05, p<0.01, p < 0.001, and relative to smegglutinin, #p < 0.05, ##p < 0.01, and ###p < 0.001; one-way ANOVA was followed by Bonferroni post-test.

[0252] Example 2: Efficacy study in db / db mice at a dose of 12 nM / kg

[0253] The effects of the compounds described herein on blood glucose, food intake, and body weight were investigated in mice. This study was conducted in a type 2 diabetic mouse (db / db) model. Animals were randomly assigned to five treatment groups (n=4 / group): diabetic control, cotadiopeptide (12 nM / kg), compound 3 (12 nM / kg), compound 4 (12 nM / kg), and smegglutinin (12 nM / kg). Baseline blood glucose was measured in all animals. The test compounds were administered subcutaneously to all animals. Blood glucose was measured at 4, 8, 24, 48, 72, and 96 hours post-treatment. Delta blood glucose (mM) was calculated.

[0254] Table 7. Effects on blood glucose

[0255] Compared to the diabetic control group p<0.05, p<0.01, p<0.001, and a two-way ANOVA was subsequently performed using the Bonferroni post-test.

[0256] Example 3: Efficacy study in db / db mice at a dose of 10 nM / kg

[0257] The effects of the compound described herein on blood glucose, food intake, and body weight were investigated in mice. This study was conducted in a type 2 diabetic mouse (db / db) model. Animals were randomly assigned to two treatment groups (n=5 / group): a diabetic control group and compound 5 (10 nM / kg). Baseline blood glucose was measured in all animals. The test compound was administered subcutaneously to all animals. Blood glucose was measured at 4, 8, 12, 24, 48, 72, and 96 hours post-treatment. Delta blood glucose (mM) was calculated. Body weight change and cumulative food intake were measured at 48 and 96 hours post-treatment.

[0258] Table 8. Effects on blood glucose

[0259] Compared to the diabetic control group p<0.05, p<0.01, p<0.001, and a two-way ANOVA was subsequently performed using the Bonferroni post-test.

[0260] Table 9. Effects on body weight

[0261] Compared to the diabetic control group p<0.05, p<0.01, p < 0.001; one-way ANOVA was followed by Dunnett post-test.

[0262] Table 10. Effects on food intake

[0263] Compared to the diabetic control group p<0.05, p<0.01, p < 0.001; one-way ANOVA was followed by Dunnett post-test.

[0264] Example 4: Efficacy study in db / db mice at a dose of 10 nM / kg

[0265] The effects of the compounds described herein on blood glucose, food intake, and body weight were investigated in mice. This study was conducted in a type 2 diabetic mouse (db / db) model. Animals were randomly assigned to 12 treatment groups (n=4 / group): diabetic control, mascara (10 nM / kg), compound 6 (10 nM / kg), compound 7 (10 nM / kg), compound 25 (10 nM / kg), and tirzepatide (10 nM / kg). Baseline blood glucose was measured in all animals. The test compounds were administered subcutaneously to all animals. Blood glucose was measured at 4, 8, 12, 24, 48, 72, and 96 hours post-treatment. Delta blood glucose (mM) was calculated. Body weight change and cumulative food intake were measured at 48 and 96 hours post-treatment.

[0266] Table 11. Effects on blood glucose

[0267] Compared to the diabetic control group p<0.05, p<0.01, p<0.001, and a two-way ANOVA was subsequently performed using the Bonferroni post-test.

[0268] Table 12. Effects on body weight

[0269] Compared to the diabetic control group p<0.05, p<0.01, p < 0.001; one-way ANOVA was followed by Dunnett post-test.

[0270] Table 13. Effects on food intake

[0271] Compared to the diabetic control group p<0.05, p<0.01, p < 0.001; one-way ANOVA was followed by Dunnett post-test.

[0272] Example 5: in vitro Measurement

[0273] Cell lines stably expressing GLP-1R, GIPR, or GCGR were used to assay compounds as described herein. in vitroEfficacy. Signaling in GLP-1R, GIP-R, and GCG-R involves the activation of adenylate cyclase and the production of cAMP. The Hit Hunter® cAMP assay uses a technique called Enzyme Fragment Complementation (EFC) developed by DiscoverX, with β-galactosidase (β-Gal) as the functional reporter, to monitor the activation of GLP-1R, GIPR, or GCGR via Gi and Gs secondary messenger signaling. The enzyme is split into two complementary parts: EA stands for enzyme acceptor, and ED stands for enzyme donor. ED is fused to cAMP and competes with cell-produced cAMP for binding to a cAMP-specific antibody during the assay. Active β-Gal is formed by the complementation of exogenous EA with any unbound ED cAMP. The active enzyme can then convert a chemiluminescent substrate, producing an output signal detectable on a standard microplate reader.

[0274] Three different assays were performed using cells expressing any of the three receptors. The cAMP Hunter cell line was expanded from frozen stock according to standard procedures. 20 μL of cells were seeded into white-walled 384-well microplates and incubated at 37°C for an appropriate time prior to assay. The culture medium was aspirated, and the cells were treated with 15 μL of cAMP-conjugated antibody and 5 μL of the assay compound. After incubation with the appropriate compound, an assay signal was generated by incubation with 20 μL of cAMP-ED cell lysis mixture for 1 hour, followed by incubation with 20 μL of cAMP-EA reagent for 3 hours at room temperature. Free cAMP-ED available in the system is complementary to free cAMP-EA to form active β-Gal, which reacts with the substrate to generate a chemiluminescent signal. The microplate was read using a PerkinElmer Envision™ instrument after signal generation for chemiluminescent signal detection. The amount of signal was proportional to the concentration of cAMP generated as a result of the response. Different concentrations of samples (different for different compounds) were used to generate log concentration versus % effect curves. Four-parameter logistic curves were generated, and EC50 was determined. Use the appropriate assay reference for each assay (exotropic peptide-4 for GLP-1R, GIP for GIPR, glucagon for GCGR).

[0275] Cellular cAMP assays were performed for mascara, compound 6, compound 7 and compound 8, and the half-maximal effective concentrations for GLP-1R-expressing cells and GIPR-expressing cells are shown in Table 14.

[0276] Table 14.

Claims

1. A polypeptide or a pharmaceutically acceptable salt thereof, comprising an amino acid sequence: X1-X2-X3-GTFTSD-X10-S-X12-X13-LD-X16-X17-X18-X19-X20-X21-F-X23-X 24-X25-L-X27-X28-X29-X30-X31-X32-X33-X34-X35-X36-X37-X38-X39 (SEQ ID NO: 1) in: X1 is H; X2 is D-Ser(OMe), Aib, or DS; X3 is Q; X10 is either K or Y; X12 is E, K, or I; X13 is Y, S(OMe), nor-V, nor-L, or αMe-L; X16 is S, E, or A; X17 is E, R, or K; X18 is R, K, or A; X19 is A; X20 is R, Q, or K; X21 is either D or E; X23 is either V or I; X24 is A, Q, or E; X25 is W; X27 is either E or L; X28 is A, D, or E; X29 is either G or T; X30 does not exist or is G; X31 does not exist or is P; X32 does not exist or is S; X33 does not exist or is S; X34 does not exist or is G; X35 does not exist; X36 does not exist; X37 does not exist; X38 does not exist; and X39 does not exist; The acid group of the C-terminal amino acid is either a free carboxylic acid group or is amidated to a C-terminal primary amide; and The condition is that at least one of X10 and X20 is K, and the further condition is that at least one of K contains a partially acylated side chain amino group selected from the following formula ( amino group: 。 2. A polypeptide or a pharmaceutically acceptable salt thereof, comprising an amino acid sequence: X1-X2-X3-GTFTSD-X10-S-X12-X13-LD-X16-X17-X18-X19-X20-X21-F-X23-X 24-X25-L-X27-X28-X29-X30-X31-X32-X33-X34-X35-X36-X37-X38-X39 (SEQ ID NO: 2) in: X1 is H; X2 is S, D-Ser(OMe), Aib, or DS; X3 is Q; X10 is either K or Y; X12 is E, K, or I; X13 is Y, S(OMe), nor-V, nor-L, or αMe-L; X16 is S, E, or A; X17 is E, R, or K; X18 is R, K, or A; X19 is A; X20 is R, Q, or K. X21 is either D or E; X23 is V; X24 is A, Q, or E; X25 is W; X27 is either E or L; X28 is A, D, or E; X29 is either G or T; X30 does not exist or is G; X31 does not exist or is P; X32 does not exist or is S; X33 does not exist or is S; X34 does not exist or is G; X35 does not exist; X36 does not exist; X37 does not exist; X38 does not exist; and X39 does not exist; The acid group of the C-terminal amino acid is either a free carboxylic acid group or is amidated to a C-terminal primary amide; and The condition is that at least one of X10 and X20 is K, and the further condition is that at least one of K contains a partially acylated side chain amino group selected from the following formula ( amino group: 。 3. The polypeptide according to claim 1, wherein: X2 It is either D-Ser(OMe) or Aib; X10 is K; X12 is E; X13 is Y; X16 is S; X17 is E; X18 is R; X20 is R; X21 is D; X24 is A; X27 is E; X28 is A; X29 is G; X30 is G; and X31, X32, X33, and X34 do not exist; The side chain amino group of K at position X10 ( The amino group is partially acylated using the following formula: 。 4. The polypeptide according to claim 1 or 2, wherein: X2 is DS; X10 is K; X12 is K; X13 is Y; X16 is A; X17 is R; X18 is A; X20 is Q; X21 is D; X24 is Q; X27 is L; X28 is D; X29 is T; and X31, X32, X33, and X34 do not exist; The side chain amino group of K at position X10 ( The amino group is partially acylated using the following formula: 。 5. The polypeptide according to claim 1 or 2, wherein: X2 is Aib; X10 is Y; X12 is either K or I; X13 is Y or nor-V, nor-L or αMe-L; X16 is E; X17 is K; X18 is K; X20 is K; X21 is E; X24 is E; X27 is L; X28 is E; X29 is G; X30 is G X31 is P; X32 is S; X33 is S; and X34 is G; The side chain amino group of K at position X20 ( The amino group is partially acylated using the following formula: 。 6. A polypeptide or a pharmaceutically acceptable salt thereof, comprising an amino acid sequence: H-X2-QGTFTSD-X10-SEYLDSERARDFVAWLEAGG (SEQ ID NO:3) in: X2 is S, DS(OMe), or Aib; and X10 is K; The acid group of the C-terminal amino acid is either a free carboxylic acid group or is amidated to a C-terminal primary amide; and The side chain amino group of K at position X10 ( The amino group is partially acylated using the following formula: The condition is that the polypeptide is not SEQ ID NO:

6.

7. The polypeptide according to claim 6, wherein: X2 is S; The side chain amino group of K at position X10 ( The amino group is partially acylated using the following formula: 。 8. The polypeptide according to claim 6, wherein: X2 is D-Ser(OMe); The side chain amino group of K at position X10 ( The amino group is partially acylated by the following formula: 。 9. The polypeptide according to claim 6, wherein: X2 is Aib; The side chain amino group of K at position X10 ( The amino group is partially acylated using the following formula: 。 10. A polypeptide or a pharmaceutically acceptable salt thereof, comprising an amino acid sequence: H-Aib-QGTFTSDYS-X12-X13-LDEKKA-X20-EFVEWLLEGPSSG (SEQ ID NO: 4) in: X12 is either K or I; X13 is either Y or nor-V; and X20 is K; The acid group of the C-terminal amino acid is either a free carboxylic acid group or is amidated to a C-terminal primary amide. The side chain amino group of K at position X20 ( The amino group is partially acylated using the following formula: 。 11. The polypeptide according to claim 10, wherein: X12 is K; and X13 is Y; The side chain amino group of K at position X20 ( The amino group is partially acylated using the following formula: 。 12. The polypeptide according to claim 10, wherein: X12 is I; and X13 is NOR-V; The side chain amino group of K at position X20 ( The amino group is partially acylated by the following formula: 。 13. The polypeptide according to claim 10, wherein: X12 is K; and X13 is NOR-V; The side chain amino group of K at position X20 ( The amino group is partially acylated by the following formula: 。 14. A polypeptide or a pharmaceutically acceptable salt thereof, comprising an amino acid sequence: H-(DSer)-QGTFTSD-X10-SKYLDARAAQDFVQWLLDT (SEQ IDNO: 5) Where X10 is K; The acid group of the C-terminal amino acid is either a free carboxylic acid group or is amidated to a C-terminal primary amide; and The side chain amino group of K at position X10 ( The amino group is partially acylated using the following formula: 。 15. A polypeptide selected from: (SEQ ID NO: 18); (SEQ ID NO: 21); (SEQ ID NO: 22); and (SEQ ID NO: 23).

16. An incretin analogue comprising: Having sequence X 1 The peptide residues of -Gln-Gly-Thr-Phe-Thr-Ser-Asp-Lys (SEQ ID NO: 29), wherein X 1 This indicates Aib or Ser(OMe), and wherein the lysine contains a fatty acid elongation group attached to the ε-nitrogen of the lysine; and Gly-Gly-OH peptide residues indirectly linked to the carboxyl group of the lysine.

17. An incretin analog comprising: Having sequence X 1 The peptide residues of -Gln-Gly-Thr-Phe-Thr-Ser-Asp-Lys (SEQ ID NO: 29), wherein X 1 This represents Aib or Ser(OMe), and wherein the lysine contains a group of formula (I) linked to the ε-nitrogen of the lysine. (I) in: U does not exist or represents –C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-}, where} is the connection point with W; W indicates: –C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-], -C(O)-NH-(CH2)3-4-NH-], -C(O)-C(CH3)2-NH-] or , Where ] is the connection point with Y; Y does not exist or represents –C(O)-(CH2)2-CH(CO2H)NH-- or -C(O)CH((CH2) x CO2H)NH--, where x is 1, 2 or 3, and -- is the connection point with Z; and Z represents –C(O)-(CH2) n -COOH or –C(O)-(CH2) n -CH3, where n is an integer from 14 to 20; and Gly-Gly-OH peptide residues indirectly linked to the carboxyl group of the lysine.

18. The incretin analogue of claim 16 or claim 17, wherein the lysine is linked to the Gly-Gly-OH residue via a peptide residue comprising 18 amino acids.

19. An incretin analogue comprising: Peptide residues having the sequence Aib-Gln-Gly-Thr-Phe-Thr-Ser-Asp (SEQ ID NO: 30); Lysine residues indirectly linked to the carboxyl group of the residue Aib-Gln-Gly-Thr-Phe-Thr-Ser-Asp (SEQ ID NO: 30), wherein the lysine contains a group of formula (I) linked to the ε-nitrogen of lysine; (I) in: U does not exist or represents –C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-}, where} is the connection point with W; W indicates –C(O)-CH2-O-(CH2)2-O-(CH2)2-NH-]or -C(O)-NH-(CH2)3-4-NH-], -C(O)-C(CH3)2-NH-], Where ] is the connection point with Y; Y does not exist or represents –C(O)-(CH2)2-CH(CO2H)NH-- or -C(O)CH((CH2) x CO2H)NH--, where x is 1, 2 or 3, and -- is the connection point with Z; and Z represents –C(O)-(CH2) n -COOH or –C(O)-(CH2) n -CH3, where n is an integer from 14 to 20; and Gly-Gly-Pro-Ser-Ser-Gly-CONH2 peptide residues indirectly linked to the carboxyl group of the lysine.

20. The incretin analog of any one of claims 19, wherein the lysine is linked to the Aib-Gln-Gly-Thr-Phe-Thr-Ser-Asp (SEQ ID NO: 30) residue via a peptide residue comprising 10 amino acids.

21. A pharmaceutical composition comprising a pharmaceutically acceptable carrier and an incretin analogue as claimed in any one of claims 1 to 20.

22. A method of treating obesity, comprising administering an incretin analogue as described in any one of claims 1 to 20 to a patient in need of such treatment.

23. A method of treating type 2 diabetes mellitus (T2DM) comprising administering an incretin analogue as described in any one of claims 1 to 20 to a patient in need of such treatment.

24. A method of treating metabolic syndrome, comprising administering an incretin analogue as described in any one of claims 1 to 20 to a patient in need of such treatment.

25. A method of treating metabolic dysfunction-associated fatty liver disease (MASLD), comprising administering an incretin analogue as described in any one of claims 1 to 20 to a patient in need of such treatment.

26. A method of treating metabolic dysfunction-associated steatohepatitis (MASH), comprising administering an incretin analogue as described in any one of claims 1 to 20 to a patient in need of such treatment.

27. A method of treating a neurodegenerative disease, comprising administering an incretin analogue as described in any one of claims 1 to 20 to a patient in need of such treatment.

28. A method of treating fibrosis, comprising administering an incretin analogue as described in any one of claims 1 to 20 to a patient in need of such treatment.

29. A method for reducing cardiovascular risk, comprising administering an incretin analogue as described in any one of claims 1 to 20 to a patient in need of such treatment.

30. A method of treating hyperlipidemia / dyslipidemia, comprising administering an incretin analog as described in any one of claims 1 to 20 to a patient in need of such treatment.

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