GLP-1r / GIPR / GCGR triple agonist and use thereof

Abstract

Provided are a GLP-1R / GIPR / GCGR triple agonist and the use thereof. Specifically disclosed is a peptide compound comprising an amino acid sequence represented by general formula (I), and described are the use of the GLP-1R / GIPR / GCGR triple agonist and the peptide compound in a medicament for treating and / or preventing metabolic disorder-related diseases, bone-related diseases, cardiovascular diseases, and neurodegenerative diseases.

Description

A GLP-1R / GIPR / GCGR triple agonist and its uses

[0001] This disclosure claims priority to Chinese Patent Application No. 2024117812039, filed on December 5, 2024, entitled “A GLP-1R / GIPR / GCGR Triple Agonist and Its Use Thereof”, the entire contents of which are incorporated herein by reference. Technical Field

[0002] This invention relates to novel compounds and their uses as agonists of glucagon-like peptide-1 receptor (GLP-1R), glucose-dependent insulinotropic peptide receptor (GIPR), and glucagon receptor (GCGR). Background Technology

[0003] Currently, one-third of the world's population is overweight or obese, a number projected to reach 500 million by 2025. Diabetes and obesity are health concerns worldwide and are linked to a variety of diseases, particularly cardiovascular disease, obstructive sleep apnea, stroke, peripheral artery disease, microvascular complications, and osteoarthritis. Type 2 diabetes mellitus (T2DM) is a chronic metabolic disorder characterized by increased hepatic glucose output, pancreatic beta-cell dysfunction, insufficient insulin secretion, and insulin resistance, ultimately leading to persistent hyperglycemia. T2DM is also a leading cause of kidney failure, blindness, and amputation, and is closely associated with a high risk of death from cardiovascular diseases. Furthermore, the rapid increase in obesity will exacerbate the prevalence of T2DM, a phenomenon particularly pronounced in developing countries. While many medications for treating diabetes have been approved by the FDA (Food and Drug Administration), new treatments are still needed to better control blood sugar and weight. Currently, the most effective treatment for type 2 diabetes is insulin injection, but its side effect is the risk of hypoglycemia. Other commonly used small-molecule drugs for diabetes, such as sulfonylureas and meglitinides, can also easily cause hypoglycemia. Metformin affects vitamin absorption, and DPP-4 inhibitors may have some drawbacks such as increased blood pressure and triggering immune responses.

[0004] Incretins are a group of gastrointestinal hormones involved in various physiological functions, including glucose homeostasis, insulin secretion, gastric emptying, intestinal growth, and regulation of food intake. Proglucagon is a 158-amino acid precursor peptide that is processed in different tissues to form many different peptides. Incretins include various proglucagon-derived peptides, including glucagon (GCG), glucagon-like peptide-1 (GLP-1, amino acids 7-36 and 7-35), glucagon-like peptide-2 (GLP-2), and gastrin (OXM).

[0005] Glucagon-like peptide-1 (GLP-1) is a glucose-dependent incretin. After binding to the GLP-1 receptor (GLP-1R) on pancreatic β-cells, GLP-1 activates the intracellular cyclic adenosine monophosphate (cAMP) and mitogen-activated protein kinase (MAPK) pathways, synergistically stimulating insulin synthesis and secretion with glucose. In addition, GLP-1 also possesses pharmacological functions such as protecting and promoting pancreatic β-cell proliferation, improving insulin sensitivity, inhibiting glucagon secretion, inhibiting gastric emptying, reducing appetite, suppressing food intake, and controlling weight. Because natural human GLP-1 has a very short half-life and lacks drug-like properties, structural optimization and modification of natural GLP-1 are necessary to improve the biological half-life of such drugs. These GLP-1 derivatives are structurally and functionally similar to GLP-1, and can bind to and activate GLP-1R; therefore, they are called GLP-1 analogs or GLP-1R agonists. In the rodent model of NASH, GLP-1 analogues can reduce liver enzyme levels and oxidative stress, improve hepatic lipid metabolism disorders, inhibit lipid oxidation, and improve the degree of liver histological damage (Trevaskis JL, Griffin PS, Carrie W, et al. Ajp Gastrointestinal & Liver Physiology, 2012, 302(8): G762-72.).

[0006] Glucagon is a hormone secreted by pancreatic α cells, consisting of a single-chain polypeptide of 29 amino acids. Glucagon exerts its physiological effects by specifically binding to glucagon receptors (GCGRs) on the surface of target cells in the liver and kidneys, activating intracellular adenylate cyclase, increasing intracellular cAMP levels. Glucagon is a catabolism-promoting hormone; short-term injections of glucagon can promote glycogenolysis and gluconeogenesis, leading to elevated blood glucose levels. Glucagon and insulin are an opposite pair of hormones, forming a negative feedback regulatory loop in maintaining blood glucose homeostasis. More importantly, animal and human studies have shown that long-term glucagon injection to activate GCGRs can reduce appetite, stimulate fatty acid breakdown, and significantly increase energy expenditure in adipose tissue (Campbell JE, Drucker DJ. Nature Reviews Endocrinology, 2015, 11(6):329–338.).

[0007] Glucose-dependent insulinotropic peptide (GIP) is a member of the secretin hormone family. GIP is derived from the 153-amino acid proprotein encoded by the GIP gene and circulates as a biologically active 42-amino acid peptide. The GIP gene is expressed in the small intestine and salivary glands and acts as a weak inhibitor of gastric acid secretion. In addition to its inhibitory effect in the stomach, GIP can enhance insulin release from pancreatic β-islet cells when administered in physiological doses in the presence of glucose. GIP is considered an intestinal factor that stimulates pancreatic insulin release and may play a physiological role in maintaining glucose homeostasis.

[0008] Currently, next-generation drug development primarily focuses on concentrating these agonist activities into a single molecule, such as peptide compounds with GLP-1R / GIPR / GCGR triple agonist activity. Several publications (WO 2019 / 125929, WO 2019 / 125938, and WO 2015 / 067716) have reported on GLP-1 / GIP / GCG receptor triple agonists and their potential pharmaceutical applications. However, there remains room for further optimization and improvement of the triple agonist molecule. Summary of the Invention

[0009] To address the aforementioned deficiencies in the prior art, this invention provides a novel polypeptide compound with triple agonist activity of GLP-1 / GIP / GCG receptors, which can be used in the treatment of metabolic diseases such as type II diabetes, obesity, dyslipidemia, and non-alcoholic fatty liver disease / non-alcoholic steatohepatitis.

[0010] To achieve the above and other related objectives, the present invention adopts the following technical solution:

[0011] This invention provides a polypeptide compound of formula (I) or a pharmaceutically acceptable salt, ester, solvate, optical isomer, tautomer, isotope label, or prodrug thereof: Y-Xaa2-QGTFTSDY-Xaa 11 -I-Xaa 13 -LDK-Xaa 17 -Xaa 18 -Q-Xaa 20 -Xaa 21 -FIEYLLE-Xaa 29 -Xaa 30 -PSS-Xaa 34 -Xaa 35 -PPPS-R1 Formula (I);

[0012] in,

[0013] Xaa2 is either Aib or Ac4c;

[0014] Xaa 11 For S or Dap;

[0015] Xaa 13 For L or 2-MeL;

[0016] Xaa 17 K is either unmodified or modified, wherein the modification is: ([2-(2-amino-ethoxy)-ethoxy]-acetyl) a -(γGlu) b -CO-(CH2) c -COOH is conjugated to the ε-amino group of the K side chain, wherein a is 1 or 2, b is 1 or 2, and c is 16 or 18;

[0017] Xaa 18 It can be A or Y;

[0018] Xaa 20 For Aib, H, or Q;

[0019] Xaa 21 The answer is A, E, or D;

[0020] Xaa 29 For G, Aib, or Q;

[0021] Xaa 30 For G, Aib, or K;

[0022] Xaa 34 For G or Aib;

[0023] Xaa 35 It can be A or L;

[0024] R1 is NH2 or OH, or a pharmaceutically acceptable salt and / or ester thereof;

[0025] Furthermore, the compound of formula (I) is not one of the following compounds:

[0026] The effects of the invention

[0027] The compounds of this invention exhibit agonistic activity against GLP-1, GIP, and GCG receptors, with significant agonistic effects. Compared to wild-type peptides GLP-1 / GIP / GCG and existing tri-agonist molecules, the peptide compounds provided by this invention show enhanced activity against ECs in various tissues and cells. 50Even better. Pharmacokinetic studies have shown that the polypeptide compound of this invention has a long half-life and high exposure. In vivo pharmacodynamic studies in a DIO mouse obesity model have shown that the polypeptide compound of this invention resulted in a significantly higher reduction in mouse body weight at the experimental endpoint compared to the positive control group. The GLP-1R / GIPR / GCGR triple receptor agonist provided by this invention exhibits good stability, a sufficiently long duration of action, and effectively reduces body weight and inhibits food intake.

[0028] The term "agonistic activity" refers to the compound's ability to stimulate specific receptor cells to produce cAMP. The cells used can be host cells, pancreatic islet cells, adipocytes, hepatocytes, etc., that overexpress GLP-1, GIP, or GCG receptors, as constructed by those skilled in the art. The receptor agonistic activity can be achieved by stimulating ECMO (electrode-associated cells) in receptor cells to produce cAMP. 50 Value as a numerical measure. EC 50 The value refers to the drug concentration required to achieve half (50%) of the compound's maximum activity in a specific assay system. Attached Figure Description

[0029] Figure 1 shows the mass spectrum of compound 1.

[0030] Figure 2 shows the mass spectrum of compound 2.

[0031] Figure 3 shows the mass spectrum of compound 3.

[0032] Figure 4 shows the mass spectrum of compound 4.

[0033] Figure 5 shows the mass spectrum of compound 7.

[0034] Figure 6 shows the curves of mouse body weight change over time after administration of the drug in Test Example 5 of this invention.

[0035] Figure 7 shows the relative changes in mouse body weight over time after administration of the drug in Test Example 5 of this invention.

[0036] Figure 8 shows the curves of food intake in mice after administration of the drug over time in Test Example 5 of the present invention. Detailed Implementation

[0037] To facilitate a clearer understanding of this disclosure, certain terms are first defined. As used herein, unless otherwise expressly specified herein, each of the following terms shall have the meaning given below. Other definitions are set forth throughout the application.

[0038] Definitions and General Terms

[0039] As used herein, the terms “comprising” or “including” are open-ended expressions, meaning they include the contents specified in this invention but do not exclude other aspects.

[0040] As used herein, the terms “optionally,” “optionally,” or “optionally” generally refer to an event or condition that may, but may not, occur, and the description includes both cases in which the event or condition occurs and cases in which the event or condition does not occur.

[0041] As used in this article, the singular forms of “a,” “an,” and “the” include plural references, and vice versa, unless the context clearly indicates otherwise.

[0042] As used herein, “about” means within a statistically significant range of one or more values, such as, for example, a specified concentration, length, molecular weight, pH, sequence identity, duration of time, temperature, volume, etc. Such values ​​or ranges may be on the order of typically 20%, more typically 10%, or even more typically 5% of a given value or range. The permissible deviation covered by “about” will depend on the specific system under study and may be readily apparent to those skilled in the art.

[0043] As used herein, the term “compound” is also intended to cover its pharmaceutically relevant form, namely, the compound as defined herein or its pharmaceutically acceptable salt, ester, solvate, optical isomer, tautomer, isotopic label, or prodrug.

[0044] As used herein, the term "peptide" refers to a sequence of two or more amino acids. "Peptide" may also include amino acid elongations at the N-terminus and / or C-terminus, and / or truncations at the N-terminus and / or C-terminus. Typically, amino acid residues are represented by their full name, their single-letter code, and / or their three-letter code. These three methods are completely equivalent.

[0045] As used in the context of this disclosure, the term "pharmaceutically acceptable salt" refers to a salt prepared from a compound of this disclosure with a relatively non-toxic acid or base. When a compound of this disclosure contains a relatively acidic functional group (e.g., a carboxyl or sulfonic acid group), a base addition salt can be obtained by contacting it with a sufficient amount of base in a pure solution or a suitable inert solvent in its free form. Non-limiting examples of pharmaceutically acceptable base addition salts include, but are not limited to, sodium, potassium, ammonium, calcium, magnesium, organic amine, or similar salts. When a compound of this disclosure contains a relatively basic functional group (e.g., an amino or guanidinyl group), an acid addition salt can be obtained by contacting it with a sufficient amount of acid in a pure solution or a suitable inert solvent in its free form. Non-limiting examples of pharmaceutically acceptable acid addition salts include, but are not limited to, inorganic acid salts (e.g., hydrochloride, hydrobromide, hydroiodide, nitrate, carbonate, bicarbonate, phosphate, monohydrogen phosphate, dihydrogen phosphate, phosphite, sulfate, hydrogen sulfate, etc.), organic acid salts (e.g., acetate, propionate, isobutyrate, malonate, succinate, octanoate, maleate, fumarate, citrate, tartrate, lactate, mandelate, benzoate, phthalate, methanesulfonate, benzenesulfonate, p-toluenesulfonate, glucuronic acid, etc.), and amino acid salts (e.g., arginine salts). For specific forms of pharmaceutically acceptable salts, see Berge et al., “Pharmaceutical Salts”, Journal of Pharmaceutical Science, 1977, 66:1-19.

[0046] GLP-1 / GIP / GCG receptor triple agonist

[0047] This invention relates to compounds that act as agonists of the GLP-1 receptor, GIP receptor, and GCG receptor. Each of these agonists may also be referred to as a “GLP-1 / GIP / glucagon receptor triple agonist” or simply a “triple agonist.” Receptor agonists as described herein are compounds that bind to a receptor and elicit a response from a natural ligand (see, for example, “Principles of Biochemistry,” AL Lehninger, DL Nelson, MM Cox, 2nd ed., Worth Publishers, 1993, p. 763).

[0048] In some implementations, the GLP-1 / GIP / GCG triple agonist is a compound that binds to each of the three receptors, GLP-1R, GIPR, and GCGR, and elicits a response at each receptor, i.e., it can activate the GLP-1R, GIPR, and GCGR receptors.

[0049] polypeptide compounds

[0050] This invention provides a polypeptide compound of formula (I) or a pharmaceutically acceptable salt, ester, solvate, optical isomer, tautomer, isotope label, or prodrug thereof: Y-Xaa2-QGTFTSDY-Xaa 11 -I-Xaa 13 -LDK-Xaa 17 -Xaa 18 -Q-Xaa 20 -Xaa 21 -FIEYLLE-Xaa 29 -Xaa 30 -PSS-Xaa 34 -Xaa 35 -PPPS-R1 Formula (I);

[0051] in,

[0052] Xaa2 is either Aib or Ac4c;

[0053] Xaa 11 For S or Dap;

[0054] Xaa 13 For L or 2-MeL;

[0055] Xaa 17 K is either unmodified or modified, wherein the modification is: ([2-(2-amino-ethoxy)-ethoxy]-acetyl) a -(γGlu) b -CO-(CH2) c -COOH is conjugated to the ε-amino group of the K side chain, wherein a is 1 or 2, b is 1 or 2, and c is 16 or 18;

[0056] Xaa 18 It can be A or Y;

[0057] Xaa 20 For Aib, H, or Q;

[0058] Xaa 21 The answer is A, E, or D;

[0059] Xaa 29 For G, Aib, or Q;

[0060] Xaa 30 For G, Aib, or K;

[0061] Xaa 34 For G or Aib;

[0062] Xaa 35 It can be A or L;

[0063] R1 is NH2 or OH, or a pharmaceutically acceptable salt and / or ester thereof;

[0064] Furthermore, the compound of formula (I) is not one of the following compounds:

[0065] In a preferred embodiment, the polypeptide compound represented by formula (I) above, Xaa 13 It is 2-MeL.

[0066] In some embodiments, the amino acid sequence of the polypeptide compound represented by formula (I) is as follows: Y-Xaa2-QGTFTSDY-Xaa 11 -I-Xaa 13 -LDK-Xaa 17 -Xaa 18 -Q-Xaa 20 -Xaa 21 -FIEYLLE-Xaa 29 -Xaa 30 -PSS-Xaa 34 -Xaa 35 -PPPS-R1 Formula (I);

[0067] in,

[0068] Xaa2 is either Aib or Ac4c;

[0069] Xaa 11 For DAP;

[0070] Xaa 13 It is 2-MeL;

[0071] Xaa 17 K is either unmodified or modified, wherein the modification is: ([2-(2-amino-ethoxy)-ethoxy]-acetyl) a -(γGlu) b -CO-(CH2) c -COOH is conjugated to the ε-amino group of the K side chain, wherein a is 1 or 2, b is 1 or 2, and c is 16 or 18;

[0072] Xaa 18 It can be A or Y;

[0073] Xaa 20 For Aib;

[0074] Xaa 21 The answer is A, E, or D;

[0075] Xaa 29 For G or Aib;

[0076] Xaa 30 For G, Aib, or K;

[0077] Xaa 34 For G or Aib;

[0078] Xaa 35 It can be A or L.

[0079] In some implementations, Xaa 29 It is Aib.

[0080] In some implementations, Xaa 29 For Aib, Xaa 21 The answer is A.

[0081] In some implementations, Xaa2 is Aib, Xaa 21 for A, Xaa 29 It is Aib.

[0082] In some implementations, Xaa2 is Aib, Xaa 18 for A, Xaa 21 for A, Xaa 29 It is Aib.

[0083] In some implementations, Xaa2 is Aib or Ac4c, Xaa 21 It can be D or E.

[0084] In some implementations, Xaa2 is Aib or Ac4c, Xaa 21 For D, Xaa 29 It is Aib.

[0085] In some implementations, Xaa2 is Ac4c, Xaa 21 For D, Xaa 29 It is Aib.

[0086] In some embodiments, the amino acid sequence of the polypeptide compound represented by formula (I) is as follows: Y-Xaa2-QGTFTSDY-Xaa 11 -I-Xaa 13 -LDK-Xaa 17 -Xaa 18 -Q-Xaa 20 -Xaa 21 -FIEYLLE-Xaa 29 -Xaa 30 -PSS-Xaa 34 -Xaa 35 -PPPS-R1 Formula (I);

[0087] Xaa2 is Aib;

[0088] Xaa 11 S;

[0089] Xaa 13 For L or 2-MeL;

[0090] Xaa 17 K is either unmodified or modified, wherein the modification is: ([2-(2-amino-ethoxy)-ethoxy]-acetyl) a -(γGlu) b -CO-(CH2) c -COOH is conjugated to the ε-amino group of the K side chain, wherein a is 1 or 2, b is 1 or 2, and c is 16 or 18;

[0091] Xaa 18 A;

[0092] Xaa 20 For Aib, H, or Q;

[0093] Xaa 21 E;

[0094] Xaa 29 For G, Aib, or Q;

[0095] Xaa 30 G;

[0096] Xaa 34 G;

[0097] Xaa 35 The answer is A.

[0098] In some implementations, Xaa 13 It is 2-MeL.

[0099] In some implementations, Xaa 13 For 2-MeL, Xaa 29 For G or Aib.

[0100] In some implementations, Xaa 13 For 2-MeL, Xaa 20 For Aib or H, Xaa 29 It is Aib.

[0101] In some implementations, Xaa 17 Let K be the modified form, where a is 1, b is 1, and c is 18.

[0102] In some implementations, the modified structure of K is shown below:

[0103] In some embodiments, R1 is NH2. In this case, -S-NH2 is serineamide, with the following structure:

[0104] In some implementations, R1 is OH. In this case, -S-OH is serine, with the following structure:

[0105] In some embodiments, the polypeptide compound is selected from any of the following:

[0106] In some embodiments, the polypeptide compound is selected from any of the following:

[0107] Amino acids are molecules containing an amino group and a carboxylic acid group, and optionally one or more additional groups, often referred to as side chains. The amino acids in the compound sequences described in this invention are derived from natural amino acids or related amino acid variants and / or derivatives. The abbreviations and codes of the natural amino acids follow generally accepted rules well known to those skilled in the art. Amino acids are typically described using standard single-letter codes (e.g., L = leucine) and α-methyl substituted residues of natural amino acids (e.g., α-methylleucine, or αMeL) and certain other non-natural amino acids (such as "Aib", "Dap", "Ac4c", etc.). Unless explicitly stated otherwise, all amino acid residues in this invention are preferably in the L-configuration, e.g., dS represents D-type Ser.

[0108] Amino acids, their abbreviations, and English abbreviations are shown in Table 1:

[0109] Table 1

[0110] The structures of non-natural amino acids and other abbreviations are shown below:

[0111] In this article, the term "2-MeL" or "αMeL" refers to 2-methyl-leucine, whose structural formula is as follows:

[0112] In this article, the term "Aib" refers to α-aminoisobutyric acid, whose structural formula is as follows:

[0113] In this article, the term "Ac4c" refers to 1-amino-cyclobutane carboxylic acid, whose structural formula is:

[0114] In this article, the term "Dap" refers to diaminopropionic acid, whose structural formula is:

[0115] In this article, when referring to peptides, "amino acid" and "amino acid residue" have the same meaning, referring to the amino acid residues that remain after some groups are lost due to their participation in the formation of the linking bond when amino acids are linked by chemical bonds.

[0116] As used herein, the term “pharmaceutical acceptable” means that such compounds, materials, compositions, and / or dosage forms are suitable for contact with patient tissues without excessive toxicity, irritation, allergic reactions, or other problems or complications, within reasonable medical judgment, have a reasonable benefit / risk ratio, and are effective for their intended use.

[0117] As used herein, the term "pharmaceutically acceptable salt" means a salt obtained by reacting a compound of the present invention with a free acid or a free base, wherein the free acid is obtained by reacting with a non-toxic inorganic or organic base, and the free base is obtained by reacting with a non-toxic inorganic or organic acid.

[0118] As used herein, the term "solvent" refers to the physical bond between a compound of the present invention and one or more, preferably one to three, solvent molecules, whether organic or inorganic. This physical bond includes hydrogen bonds. In some cases, such as when one or more, preferably one to three, solvent molecules are incorporated into the lattice of a crystalline solid, the solvate will be separated. Exemplary solvates include, but are not limited to, hydrates, ethanolates, methanolates, and isopropanolates. Solvation methods are well known in the art.

[0119] Functional properties

[0120] The triple agonists described herein are agonists targeting all receptors of GLP-1, GIP, and GCG, as tested by the in vitro potency described in the examples herein. Therefore, the triple agonists described herein can activate each of GLP-1R, GIPR, and GCGR.

[0121] Bioactivity – In vitro efficacy

[0122] In this article, the term “potency” may be used interchangeably with “bioactivity” or “activity”.

[0123] The triple agonists or peptide compounds described herein are effective against GLP-1R, GIPR, and GCGR. In some embodiments, the triple agonists described herein have effective in vitro activation of hGLP-1R, hGIPR, and hGCGR.

[0124] In some implementations, potency is determined by in vitro potency, i.e., the performance of the triple agonist (or peptide described herein) against each of GLP-1R, GIPR, and GCGR in a functional receptor assay.

[0125] As used in this article, the half-maximum effective concentration (also known as "EC50") is... 50 ") refers to the concentration at which a response is induced at half the baseline and maximum value, based on a reference dose-response curve (e.g., cAMP). EC 50 It is used to measure the potency of a compound and represents the concentration at which 50% of its maximum effect is observed.

[0126] Therefore, the in vitro potency of the triple agonist as described herein can be determined by measuring ECGs as follows. 50 To determine. EC 50 The lower the value, the better the efficacy.

[0127] Pharmacokinetic profile – half-life in mice

[0128] Triple agonists, as described herein, possess improved pharmacokinetic properties and, compared to natural hormones such as GLP-1, GIP, and GCG, have a prolonged terminal half-life. A prolonged terminal half-life means that the compounds in question are cleared from the body more slowly. For the triple agonists described herein, this results in an extended duration of pharmacological action.

[0129] The pharmacokinetic properties of the triple agonists described herein can be appropriately determined in vivo in pharmacokinetic (PK) studies. Such studies are conducted to evaluate how the drug compound is absorbed, distributed, and eliminated in vivo over time, and how these processes affect the concentration of the triple agonist in vivo.

[0130] In the discovery and preclinical stages of drug development, animal models such as mice, rats, monkeys, dogs, or pigs can be used for pharmacokinetic (PK) studies. Any such model can be used to test the PK properties of the triple agonists described in this article.

[0131] In these types of studies, a single dose of the drug is typically administered to animals intravenously (iv), subcutaneously (sc), or orally (po) in the form of the relevant formulation. Blood samples are collected at predetermined time points after administration, and the drug concentration in the samples is analyzed by relevant quantitative assays. Based on these measurements, a time-plasma concentration curve of the compound under study is plotted, and the data are subjected to so-called non-compartmental pharmacokinetic analysis.

[0132] Terminal half-life is an important parameter because a long half-life indicates that it is possible to administer the compound at a lower frequency.

[0133] Preparation methods of peptide compounds

[0134] In a second aspect, the present invention provides a method for preparing the polypeptide compound described in any of the above claims or its pharmaceutically acceptable salt, ester, solvate, optical isomer, tautomer, isotope label, or prodrug, wherein the method is to synthesize the polypeptide compound using a chemical synthesis method.

[0135] In some embodiments, the polypeptide compounds of the present invention can be prepared by standard peptide synthesis methods, such as by standard solid-phase or liquid-phase methods, stepwise or by fragment assembly, followed by separation and purification of the final peptide compound product, or by any combination of recombinant and synthetic methods. Optionally, the polypeptide compounds of the present invention can be synthesized by solid-phase or liquid-phase peptide synthesis methods.

[0136] In some embodiments, methods for preparing triple agonists are described herein. In some embodiments, the methods for preparing the triple agonists described herein include a solid-phase peptide synthesis step.

[0137] Pharmaceutical Composition

[0138] Thirdly, the present invention provides a pharmaceutical composition comprising the polypeptide compound described in any one of the preceding claims or a pharmaceutically acceptable salt, ester, solvate, optical isomer, tautomer, isotope label, or prodrug thereof, and optionally, at least one pharmaceutically acceptable carrier.

[0139] As used herein, the term "pharmaceutical composition" refers to a composition that is pharmaceutically usable and comprises one or more compounds as shown in Formula I or in a pharmaceutically acceptable form thereof (e.g., salt, hydrate, solvate, stereoisomer, tautomer, metabolite, prodrug, etc.), and other components (e.g., pharmaceutically acceptable excipients).

[0140] As used herein, the term "pharmaceuticalally acceptable carrier" refers to excipients (excipients) widely used in the pharmaceutical manufacturing industry. The primary purpose of using excipients is to provide a pharmaceutical composition that is safe to use, stable in nature, and / or has specific functionalities, and also to provide a method for the active ingredient to dissolve at a desired rate or to promote the effective absorption of the active ingredient in a subject after administration of the drug. Pharmaceutically acceptable excipients can be inert fillers or functional components that provide a function to the pharmaceutical composition (e.g., stabilizing the overall pH of the composition or preventing degradation of the active ingredient in the composition). Non-limiting examples of pharmaceutically acceptable excipients include, but are not limited to, binders, suspending agents, emulsifiers, diluents (or fillers), granulating agents, adhesives, disintegrants, lubricants, anti-adhesion agents, flow aids, wetting agents, gelling agents, absorption delay agents, dissolution inhibitors, enhancers, adsorbents, buffers, chelating agents, preservatives, colorants, flavoring agents, sweeteners, etc.

[0141] The pharmaceutical compositions disclosed herein can be prepared using any method known to those skilled in the art. For example, conventional mixing, dissolving, granulation, emulsification, grinding, encapsulation, embedding, and / or lyophilization processes.

[0142] In this disclosure, the purpose of using the pharmaceutical composition is to facilitate administration to a living organism, thereby promoting the absorption of the active ingredient and exerting its biological activity. The pharmaceutical compositions of this disclosure can be administered in any form, including injection (intra-arterial, intravenous, intramuscular, intraperitoneal, subcutaneous), mucosal, oral (oral solid dosage forms, oral liquid dosage forms), rectal, inhalation, implantation, and topical (e.g., ocular) administration. Non-limiting examples of oral solid dosage forms include, but are not limited to, powders, capsules, lozenges, granules, tablets, etc. Non-limiting examples of oral or mucosal liquid dosage forms include, but are not limited to, suspensions, tinctures, elixirs, solutions, etc. Non-limiting examples of topical dosage forms include, but are not limited to, emulsions, gels, ointments, creams, patches, pastes, foams, lotions, drops, or serum preparations. Non-limiting examples of parenteral dosage forms include, but are not limited to, solutions for injection, dry powders for injection, suspensions for injection, emulsions for injection, etc. The pharmaceutical compositions of this disclosure can also be formulated into controlled-release or delayed-release dosage forms (e.g., liposomes or microspheres).

[0143] In this disclosure, the application method can be varied or modified in any applicable manner to meet the needs of the properties of the drug, the convenience of patients and medical personnel, and other relevant factors.

[0144] Fourthly, the present invention provides the polypeptide compound described in any of the above claims or its pharmaceutically acceptable salt, ester, solvate, optical isomer, tautomer, isotope label or prodrug or its salt or solvate and / or the pharmaceutical composition described above as an agonist of GLP-1, GCG, GIP receptors.

[0145] Fifthly, the present invention provides the use of any of the above-described polypeptide compounds or their pharmaceutically acceptable salts, esters, solvates, optical isomers, tautomers, isotope markers or prodrugs and / or the above-described pharmaceutical compositions in the preparation of medicaments for the treatment and / or prevention of metabolic disorder-related diseases, bone-related diseases, cardiovascular diseases and neurodegenerative diseases.

[0146] As used herein, “metabolic disorder” can refer to disorders of glucose metabolism such as diabetes, diabetes complications, obesity, and obesity-related complications. Since the link between obesity, diabetes, and glucose metabolism is well-known, these conditions may, but do not necessarily, be separate or mutually exclusive. In some embodiments, diabetes or diabetes complications include insulin resistance, impaired glucose tolerance, elevated fasting blood glucose, prediabetes, type 1 diabetes, type 2 diabetes, gestational diabetes mellitus, hypertension, dyslipidemia, or a combination thereof. In some embodiments, obesity complications include obesity-related inflammation, obesity-related gallbladder disease, or obesity-induced sleep apnea, or may be selected from the following related conditions: atherosclerotic dyslipidemia, dyslipidemia, elevated blood pressure, hypertension, prethrombotic states, and pro-inflammatory states, or a combination thereof.

[0147] In some implementations, the metabolic disorder-related diseases include obesity, diabetes, dyslipidemia-related diseases, fatty liver disease, metabolic syndrome, non-alcoholic steatohepatitis, and non-alcoholic fatty liver disease.

[0148] In some implementations, the diabetes includes type 2 diabetes.

[0149] In some implementations, the neurodegenerative diseases include Alzheimer's disease and Parkinson's disease.

[0150] In a sixth aspect, the present invention provides a method for treating metabolic disorder-related diseases or neurodegenerative diseases, the method comprising: administering to a subject an effective dose of any of the above-described polypeptide compounds or pharmaceutically acceptable salts, esters, solvates, optical isomers, tautomers, isotope labels, or prodrugs, or combinations thereof.

[0151] In some implementations, the metabolic disorder-related diseases include obesity, diabetes, dyslipidemia-related diseases, fatty liver disease, metabolic syndrome, non-alcoholic steatohepatitis, and non-alcoholic fatty liver disease.

[0152] In some implementations, the diabetes includes type 2 diabetes.

[0153] In some embodiments, the neurodegenerative diseases include Alzheimer's disease and Parkinson's disease. The effective amount of the polypeptide compound or its pharmaceutically acceptable salt, ester, solvate, optical isomer, tautomer, isotope label, prodrug, or pharmaceutical composition described in this invention may vary depending on the administration method and the severity of the disease to be treated. A preferred effective amount can be determined by those skilled in the art based on various factors (e.g., through clinical trials). These factors include, but are not limited to, the pharmacokinetic parameters of the active ingredient, such as bioavailability, metabolism, and half-life; the severity of the disease to be treated, the patient's weight, the patient's immune status, and the route of administration. However, in addition to the route of administration and treatment frequency of the pharmaceutical composition, various factors including the patient's age, weight, health status, sex, disease severity, diet, and excretion rate are considered in determining the effective dose of the peptide compound. In this respect, those skilled in the art can readily determine the effective dose suitable for the specific use of the composition of this invention. The compositions according to the invention are not particularly limited in formulation and route of administration or method, as long as they exhibit the effects of the invention.

[0154] The polypeptide compounds or their salts or solvates, or pharmaceutical compositions described in this invention, can be incorporated into medicaments suitable for parenteral administration (e.g., intravenous, subcutaneous, intraperitoneal, intramuscular). These medicaments can be prepared in various forms, such as liquids, semi-solids, and solid dosage forms, including but not limited to liquid solutions (e.g., injection solutions and infusion solutions) or lyophilized powders. Typical medicaments are in the form of injection solutions or infusion solutions. The aforementioned polypeptides or their derivatives, or pharmaceutical compositions, can be administered by intravenous infusion or injection, or by intramuscular or subcutaneous injection.

[0155] The following specific examples illustrate the implementation of the present invention. Those skilled in the art can easily understand other advantages and effects of the present invention from the content disclosed in this specification. The present invention can also be implemented or applied through other different specific embodiments, and various details in this specification can also be modified or changed based on different viewpoints and applications without departing from the spirit of the present invention.

[0156] Before further describing specific embodiments of the present invention, it should be understood that the scope of protection of the present invention is not limited to the specific embodiments described below; it should also be understood that the terminology used in the embodiments of the present invention is for describing specific embodiments and not for limiting the scope of protection of the present invention.

[0157] Unless otherwise stated, the experimental methods, detection methods, and preparation methods disclosed in this invention all employ conventional techniques in molecular biology, biochemistry, chromatin structure and analysis, analytical chemistry, cell culture, recombinant DNA technology, and related fields. These techniques have been well described in existing literature; see Sam Brook et al., *MOLECULAR CLONING: A LABORATORY MANUAL*, Second edition, Cold Spring Harbor Laboratory Press, 1989 and Third edition, 2001; Ausubel et al., *CURRENT PROTOCOLS IN MOLECULAR BIOLOGY*, John Wiley & Sons, New York, 1987 and periodic updates; the series *METHODS IN ENZYMOLOGY*, Academic Press, San Diego; Wolffe, *CHROMATIN STRUCTURE AND FUNCTION*, Third edition, Academic Press, San Diego, 1998; *METHODS IN ENZYMOLOGY*, Vol. 304, Chromatin (PM Wassarman and AP Wolffe, eds.), Academic Press, San Diego, 1999; and *METHODS IN MOLECULAR*. BIOLOGY, Vol. 119, Chromatin Protocols (PB Becker, ed.) Humana Press, Totowa, 1999, etc.

[0158] In addition to the specific methods, equipment, and materials used in the embodiments, based on the knowledge of those skilled in the art and the description of this invention, any prior art methods, equipment, and materials similar to or equivalent to those described, used, and materials in the embodiments of this invention can be used to implement this invention. Unless otherwise stated, parts and percentages are parts by weight and weight percentages.

[0159] The raw materials and reagents used in this invention are all common reagents in the art, and unless otherwise specified, they can all be purchased commercially available. Of course, it is not excluded that they can be synthesized according to the methods disclosed in the prior art.

[0160] In this invention, some commonly used abbreviations have the following meanings:

[0161] Fmoc refers to fluorenylmethoxycarbonyl;

[0162] HBTU refers to 2-(1H-benzotriazol-1-yl)-1,1,3,3,-tetramethylurea hexafluorophosphate;

[0163] DCM refers to dichloromethane;

[0164] cAMP: Cyclic adenosine monophosphate;

[0165] Boc: tert-butyloxycarbonyl;

[0166] OtBu: Oxy-tert-butyl;

[0167] tBu: tert-butyl;

[0168] Dde: 1-(4,4-dimethyl-2,6-dioxocyclohexylene)ethyl

[0169] Trt: Triphenylmethyl;

[0170] Boc-Tyr(tBu)-Aib-OH: Tyrosine-2-methylalanine protected dipeptide fragment;

[0171] AEEA: Diethylene glycol (i.e., 2-(2-(2-aminoethoxy)ethoxy)acetic acid);

[0172] tBuO-Ara(OH): Tetrabutyl eicosanoate monotert-butyl ester;

[0173] DMF: Dimethylformamide;

[0174] TFA: Trifluoroacetic acid;

[0175] TIS: Triisopropylsilane;

[0176] TBME: tert-butyl methyl ether;

[0177] HOBt: 1-hydroxybenzotriazole;

[0178] DIC: N,N-diisopropylcarbodiimide (DIC).

[0179] Example 1

[0180] The raw materials and reagents used in this invention are all common reagents in the field, and unless otherwise specified, they are all purchased from commercial products.

[0181] The target compound was synthesized using the conventional Fmoc solid-phase synthesis method. Fmoc-Rink MBHA Amide resin was used, and Fmoc was removed with 20% (v / v) piperidine / DMF. HOBT / DIC was used as the coupling reagent, and DMF was used as the reaction solvent. The reaction was monitored using ninhydrin detection. The amino acids in the main chain were coupled sequentially. After the main chain was completed, the -Dde side-chain protecting group of lysine was removed with hydrazine hydrate / DMF solution. The side-chain fragments were then sequentially linked. After solid-phase synthesis, the compound was cleaved with 95% TFA / 2.5% water / 2.5% TIS (v / v), precipitated with cold TBME, purified by HPLC, and lyophilized to obtain the target compound.

[0182] This embodiment uses the synthesis of a positive control (compound 1, retaloupeptide, structural formula shown below) as an example to illustrate the synthesis and preparation method of the compounds of the present invention.

[0183] The protected amino acids used in the entire synthesis process are as follows: Fmoc-Ser(tBu)-OH, Fmoc-Pro-OH, Fmoc-Pro-OH, Fmoc-Pro-OH, Fmoc-Ala-OH, Fmoc-Gly-OH, Fmoc-Ser(tBu)-OH, Fmoc-Ser(tBu)-OH, Fmoc-Pro-OH, Fmoc-Gly-OH, Fmoc-Gly-OH, Fmoc-Glu(OtBu)-OH, Fmoc-Leu-OH, Fmoc-Leu-OH, Fmoc-Tyr(tBu)-OH, Fmoc-Glu(OtBu)-OH, Fmoc-Ile-OH, Fmoc-Phe-OH, Fmoc-Ala-OH, Fmoc-Aib- OH, Fmoc-Gln(Trt)-OH, Fmoc-Ala-OH, Fmoc-Lys(Dde)-OH, Fmoc-Lys(Boc)-OH, Fmoc-Asp(OtBu)-OH, Fmoc-Leu-OH, Fmoc-α-Me-Leu-OH, Fmoc-Ile-OH, Fmoc-Ser(tBu)-OH, Fm oc-Tyr(tBu)-OH, Fmoc-Asp(OtBu)OH, Fmoc-Ser(tBu)-OH, Fmoc-Thr(tBu)-OH, Fmoc-Phe-OH, Fmoc-Thr(tBu)-OH, Fmoc-Gly-OH, Fmoc-Gln(Trt)-OH, Boc-Tyr(tBu)-Aib-OH.

[0184] (1) The protective amino acids were sequentially attached to the Rink MBHA Amide resin;

[0185] (2) Removal of Lys side chain protecting group: Add 2% hydrazine hydrate / DMF solution to remove -Dde protecting group, wash with DMF, ninhydrin test positive, then condense Fmoc-AEEA, Fmoc-Glu-OtBu and eicosanoic acid monotert-butyl ester in sequence to obtain the fully protected resin of compound 1.

[0186] (3) Then, the product was precipitated with 95% TFA / 2.5% water / 2.5% TIS, followed by precipitation with ice-cold TBME and washing. The crude product was purified by reversed-phase HPLC and freeze-dried to obtain target compound 1.

[0187] The mass spectrum of compound 1 prepared in this embodiment is shown in Figure 1, and its information is shown in Table 2.

[0188] Example 2

[0189] Compound 2 was prepared according to the amino acid sequence of compound 2, following the preparation method of Example 1, except that Fmoc-Ser(tBu)-OH at position 11 was replaced with Fmoc-Dap(Boc)-OH. The mass spectrometry data of compound 2 in this example are as follows: molecular weight calculated to be 4730.44; [M-3H] was detected by MS. 3- 1575.83, [M-4H] 4- 1181.72, The mass spectrum of compound 2 prepared in this embodiment is shown in Figure 2, and its information is shown in Table 2.

[0190] Example 3

[0191] Compound 3 was prepared according to the preparation method of Example 1, following the amino acid sequence of compound 3, except that Fmoc-Ala-OH at position 21 was replaced with Fmoc-Glu(OtBu)-OH. The mass spectrometry data of the compound in this example are as follows: molecular weight calculated to be 4789.46, and [M-3H] was detected by MS. 3- 1595.57, [M-4H] 4- 1196.47. The mass spectrum of compound 3 prepared in this embodiment is shown in Figure 3, and its information is shown in Table 2.

[0192] Example 4

[0193] Compound 4 was prepared according to the amino acid sequence of compound 4, following the preparation method of Example 1, except that Fmoc-Ser(tBu)-OH at position 11 was replaced with Fmoc-Dap(Boc)-OH, and Fmoc-Ala-OH at position 21 was replaced with Fmoc-Glu(OtBu)-OH. The mass spectrometry data of the compound in this example are as follows: molecular weight calculated to be 4788.47, and [M-3H] was found by MS. 3- 1595.17, [M-4H]4- 1196.08, The mass spectrum of compound 4 prepared in this embodiment is shown in Figure 4, and its information is shown in Table 2.

[0194] Example 5

[0195] Compound 5 was prepared according to the amino acid sequence of compound 5, following the preparation method of Example 1, except that Fmoc-Ala-OH at position 21 was replaced with Fmoc-Glu(OtBu)-OH, and Fmoc-Gly-OH at position 29 was replaced with Fmoc-Aib-OH. The mass spectrometry data of the compound in this example are as follows: molecular weight calculated to be 4817.51, and [M-3H] was detected by MS. 3- 1605.63, [M-4H] 4- 1204.03, [M-5H] 5- 962.52, the information of which is shown in Table 2.

[0196] Example 6

[0197] Compound 6 was prepared according to the preparation method of Example 1, following the amino acid sequence of compound 6. The difference was that Fmoc-Aib-OH at position 20 was replaced with Fmoc-His-OH, Fmoc-Ala-OH at position 21 was replaced with Fmoc-Glu(OtBu)-OH, and Fmoc-Gly-OH at position 29 was replaced with Fmoc-Aib-OH. The mass spectrometry data of the compound in this example are as follows: molecular weight calculated to be 4869.55, and [M-3H] was found by MS. 3- 1623.10, [M-4H] 4- 1216.51, [M-5H] 5- 973.20, its information is shown in Table 2.

[0198] Example 7

[0199] Compound 7 was prepared according to the preparation method of Example 1, the difference being that the order of synthesis of the amino acid raw materials to be replaced was different. The preparation method is as follows:

[0200] The conventional Fmoc solid-phase synthesis was performed using Fmoc-Rink MBHA Amide resin. Fmoc was removed using 20% ​​piperidine / DMF. HOBT / DIC was used as the coupling agent, and DMF was used as the reaction solvent. The reaction was monitored using ninhydrin detection. The amino acids in the main chain were coupled sequentially: Fmoc-Ser(tBu)-OH, Fmoc-Pro-OH, Fmoc-Pro-OH, Fmoc-Ala-OH, Fmoc-Gly-OH, Fmoc-Ser(tBu)-OH, Fmoc-Ser(tBu)-OH, and Fmoc... -Pro-OH, Fmoc-Gly-OH, Fmoc-Aib-OH, Fmoc-Glu(OtBu)-OH, Fmoc-Leu-OH, Fmoc-Leu-OH, Fmoc-Tyr(tBu)-OH, Fmoc -Glu(OtBu)-OH, Fmoc-Ile-OH, Fmoc-Phe-OH, Fmoc-Ala-OH, Fmoc-Aib-OH, Fmoc-Gln(Trt)-OH, Fmoc-Ala-OH, Fmoc- Lys(Dde)-OH, Fmoc-Lys(Boc)-OH, Fmoc-Asp(OtBu)-OH, Fmoc-Leu-OH, Fmoc-α-Me-Leu-OH, Fmoc-Ile-OH, Fmoc-Da p(Boc)-OH, Fmoc-Tyr(tBu)-OH, Fmoc-Asp(OtBu)OH, Fmoc-Ser(tBu)-OH, Fmoc-Thr(tBu)-OH, Fmoc-Phe-OH, Fmoc- After the main chain was completed (Th(tBu)-OH, Fmoc-Gly-OH, Fmoc-Gln(Trt)-OH, Boc-Tyr(tBu)-Aib-OH), the lysine side-chain protecting group -Dde was removed using hydrazine hydrate / DMF solution. Then, side-chain fragments Fmoc-AEEA, Fmoc-Glu-OtBu, and eicosanoic acid monotert-butyl ester were sequentially linked to obtain the fully protected resin of compound 7. The resin was then cleaved using 95% TFA / 2.5% water / 2.5% TIS, followed by precipitation with ice-cold TBME and washing. The crude product was purified by reversed-phase HPLC and freeze-dried to obtain the pure product. The mass spectrometry data of the compound in this example are as follows: molecular weight calculated as 4758.49, [M+3H] was detected by MS. 3+ 1586.91, [M+4H] 4+ 1190.23, [M+5H] 5+ 952.77. The mass spectrum of compound 7 prepared in this embodiment is shown in Figure 5, and its information is shown in Table 2.

[0201] Example 8

[0202] This invention follows the preparation method of Example 1, preparing compound 8 according to its amino acid sequence, with the difference being that Fmoc-Ser(tBu)-OH at position 11 is replaced with Fmoc-Dap(Boc)-OH, Fmoc-Ala-OH at position 21 is replaced with Fmoc-Asp(OtBu)-OH, and Fmoc-Gly-OH at position 29 is replaced with Fmoc-Aib-OH. The mass spectrometry data of this compound are as follows: calculated molecular weight: 4802.50, MS detected [M+3H]. 3+ 1601.96, [M+4H] 4+ 1201.54, [M+5H] 5+ 961.74, the information is shown in Table 2.

[0203] Example 9

[0204] This invention prepares compound 9 according to the preparation method of Example 1, following the amino acid sequence of compound 9. The difference lies in replacing Fmoc-Aib-OH at position 2 with Fmoc-Ac4c-OH, replacing Fmoc-Ser(tBu)-OH at position 11 with Fmoc-Dap(Boc)-OH, replacing Fmoc-Ala-OH at position 21 with Fmoc-Glu(OtBu)-OH, and replacing Fmoc-Gly-OH at position 29 with Fmoc-Aib-OH. The mass spectrometry data of the compound in this example are as follows: calculated molecular weight: 4828.54, MS detected [M+3H]. 3+ 1610.70, [M+4H] 4+ 1208.25, [M+5H] 5+ 966.89, the information is shown in Table 2.

[0205] Example 10

[0206] This invention follows the preparation method of Example 1, preparing compound 10 according to its amino acid sequence, with the difference being that Fmoc-Ser(tBu)-OH at position 11 is replaced with Fmoc-Dap(Boc)-OH, Fmoc-Ala-OH at position 18 is replaced with Fmoc-Tyr(tBu)-OH, Fmoc-Ala-OH at position 21 is replaced with Fmoc-Glu(OtBu)-OH, and Fmoc-Gly-OH at position 29 is replaced with Fmoc-Aib-OH. The mass spectrometry data of this compound are as follows: calculated molecular weight: 4908.62, MS detected [M+3H]. 3+ 1637.53, [M+4H] 4+ 1228.53, [M+5H] 5+982.88, its information is shown in Table 2.

[0207] Example 11

[0208] This invention prepares compound 11 according to the preparation method of Example 1, following the amino acid sequence of compound 11. The difference lies in replacing Fmoc-Ser(tBu)-OH at position 11 with Fmoc-Dap(Boc)-OH, Fmoc-Ala-OH at position 21 with Fmoc-Glu(OtBu)-OH, and Fmoc-Gly-OH at position 29 with Fmoc-Aib-OH. The mass spectrometry data of the compound in this example are as follows: calculated molecular weight: 4816.53, MS detected [M-3H]. 3- 1604.93, [M-4H] 4- 1203.41, [M-5H] 5- 962.28, its information is shown in Table 2.

[0209] Example 12

[0210] The present invention prepared compound 12 according to the preparation method of Example 1, based on the amino acid sequence of compound 12. The difference is that Fmoc-Aib-OH at position 2 was replaced with Fmoc-Ac4c-OH, Fmoc-Ser(tBu)-OH at position 11 was replaced with Fmoc-Dap(Boc)-OH, and Fmoc-Gly-OH at position 29 was replaced with Fmoc-Aib-OH. Its purity and molecular weight are shown in Table 2.

[0211] Example 13

[0212] The present invention prepared compound 13 according to the preparation method of Example 1, following the amino acid sequence of compound 13. The difference is that Fmoc-Ser(tBu)-OH at position 11 was replaced with Fmoc-Dap(Boc)-OH, Fmoc-Ala-OH at position 18 was replaced with Fmoc-Tyr(tBu)-OH, and Fmoc-Gly-OH at position 29 was replaced with Fmoc-Aib-OH. The purity and molecular weight are shown in Table 2.

[0213] Example 14

[0214] The present invention prepared compound 14 according to the preparation method of Example 1, following the amino acid sequence of compound 14. The difference is that Fmoc-Aib-OH at position 2 was replaced with Fmoc-Ac4c-OH, Fmoc-Ser(tBu)-OH at position 11 was replaced with Fmoc-Dap(Boc)-OH, Fmoc-Ala-OH at position 18 was replaced with Fmoc-Tyr(tBu)-OH, and Fmoc-Gly-OH at position 29 was replaced with Fmoc-Aib-OH. Its purity and molecular weight are shown in Table 2.

[0215] Example 15

[0216] The present invention prepared compound 15 according to the preparation method of Example 1, following the amino acid sequence of compound 15. The difference is that Fmoc-Aib-OH at position 2 was replaced with Fmoc-Ac4c-OH, Fmoc-Ser(tBu)-OH at position 11 was replaced with Fmoc-Dap(Boc)-OH, Fmoc-Ala-OH at position 21 was replaced with Fmoc-Asp(OtBu)-OH, and Fmoc-Gly-OH at position 29 was replaced with Fmoc-Aib-OH. The purity and molecular weight are shown in Table 2.

[0217] Example 16

[0218] The present invention prepared compound 16 according to the preparation method of Example 1, following the amino acid sequence of compound 16. The difference is that Fmoc-Ser(tBu)-OH at position 11 was replaced with Fmoc-Dap(Boc)-OH, Fmoc-Gly-OH at position 29 was replaced with Fmoc-Aib-OH, and Fmoc-Gly-OH at position 34 was replaced with Fmoc-Aib-OH. The purity and molecular weight are shown in Table 2.

[0219] Example 17

[0220] The present invention prepared compound 17 according to the preparation method of Example 1, following the amino acid sequence of compound 17. The difference is that Fmoc-Aib-OH at position 2 was replaced with Fmoc-Ac4c-OH, Fmoc-Ser(tBu)-OH at position 11 was replaced with Fmoc-Dap(Boc)-OH, Fmoc-Gly-OH at position 29 was replaced with Fmoc-Aib-OH, and Fmoc-Gly-OH at position 34 was replaced with Fmoc-Aib-OH. Its purity and molecular weight are shown in Table 2.

[0221] Example 18

[0222] The present invention prepared compound 18 according to the preparation method of Example 1, following the amino acid sequence of compound 18. The difference is that Fmoc-Ser(tBu)-OH at position 11 was replaced with Fmoc-Dap(Boc)-OH, Fmoc-Gly-OH at position 29 was replaced with Fmoc-Aib-OH, and Fmoc-Gly-OH at position 30 was replaced with Fmoc-Aib-OH. The purity and molecular weight are shown in Table 2.

[0223] Example 19

[0224] Compound 19 was prepared according to the amino acid sequence of compound 19 using the preparation method of Example 1. The difference was that Fmoc-Ser(tBu)-OH at position 11 was replaced with Fmoc-Dap(Boc)-OH, Fmoc-Gly-OH at position 29 was replaced with Fmoc-Aib-OH, and Fmoc-Gly-OH at position 30 was replaced with Fmoc-Lys(Boc)-OH.

[0225] Example 20

[0226] Compound 20 was prepared according to the preparation method of Example 1, and compound 20 was prepared according to its amino acid sequence. The difference was that Fmoc-Ser(tBu)-OH at position 11 was replaced with Fmoc-Dap(Boc)-OH, Fmoc-Gly-OH at position 29 was replaced with Fmoc-Aib-OH, and Fmoc-Ala-OH at position 35 was replaced with Fmoc-Leu-OH. Its purity and molecular weight are shown in Table 2.

[0227] Table 2

[0228] Test Example 1: Activity test of the peptide compounds of the present invention on human GLP-1 / GIP / GCG receptor stable cells

[0229] The reagents and equipment used in this test example are shown below:

[0230] cAMP detection kit, from Cisbio; 1M HEPES, from Invitrogen; 1 ╳HBSS, from Invitrogen; Casein, from Sigma; IBMX, from Sigma; Glacagon, from HaoYuan; GIP, from Tocris; GLP-1(7-37), from HaoYuan; OptiPlate-384plate, from PerkinElmer; 384 echo plate, from Labcyte; EnVision, from PerkinElmer; honeycomb counter, from Beckman.

[0231] The cell lines used in this test case were all derived from WuXi AppTec, and detailed information is shown in Table 3:

[0232] Table 3

[0233] The specific steps are as follows:

[0234] I) Preparation of compound plates:

[0235] (1) Dissolve and dilute all compounds to the same working concentration as the plate plot, and then perform a 4-fold serial dilution in a Bravo 384-well qualified plate for a total of 10 dilutions. After dilution, centrifuge at 1000 rpm for 1 minute.

[0236] (2) Using Bravo, transfer 5 μL of the compound to OptiPlate-384 and centrifuge at 1000 rpm for 5 seconds.

[0237] II) Preparation of cell suspension:

[0238] (1) Quickly place a tube of cells into a 37°C water bath to thaw;

[0239] (2) Transfer the cell suspension to a 15mL centrifuge tube and gently rinse with 10mL HBSS.

[0240] (3) Centrifuge the cells at 1000 rpm for 5 minutes at room temperature to precipitate the cells.

[0241] (4) Carefully aspirate the supernatant, being careful not to aspirate the cells.

[0242] (5) Gently tap the cell clumps to loosen them, then gently rinse the cell clumps with 10 mL of HBSS. Use a sterile pipette to stir up and down to separate the clumps, and centrifuge again at 1000 rpm for 5 minutes.

[0243] (6) Gently shake the cell clumps to loosen the cells, then resuspend the cell clumps in 5 mL of detection buffer and measure cell density and activity using Vi-cell.

[0244] (7) The cells were suspended in the reagent solution at a concentration of 2.0*E5 / mL (GIPR 4.0×E5 / mL);

[0245] (8) The mixed and prepared cells were transferred separately into OptiPlate-384 containing the compound.

[0246] III) HTRF agonist cAMP assay:

[0247] (1) After incubating at room temperature for 30 minutes, add cAMP detection solution;

[0248] (2) Add 10 μL of cAMP detection solution using a multichannel pipette;

[0249] (3) Cover the OptiPlate-384plate with TopSeal-A film and incubate at room temperature for 60 minutes;

[0250] (4) Remove TopSeal-A and read the value in EnVision.

[0251] Data on the stable cell activity of each compound against human GLP-1 / GIP / GCG receptors are shown in Tables 4 to 8:

[0252] Table 4

[0253] Table 5

[0254] Table 6

[0255] Table 7

[0256] Table 8

[0257] As shown in Tables 4-8, all compounds exhibited effective functional activation in human GLP-1 / GIP / GCG receptor-stabilized cells. Compound 1 was the control group, retaglutide. Compounds 2-20 in this patent all showed agonistic activity against GLP-1 / GIP / GCG. In particular, compound 7 showed approximately 5 times greater agonistic activity against human GCGR than the positive control group, and approximately 2 times greater agonistic activity against human GLP-1R than the positive control group.

[0258] Test Example 2: Sample Detection in Competitive Affinity Assay for Human GLP-1 / GIP / GCG Receptors

[0259] The main objective of this experiment is to investigate the affinity of the peptide compounds of this invention for human GLP-1 / GIP / GCG receptors, and their inhibitory activity at IC50 levels, using isotope affinity assays.50 The determination is performed to provide reference information for the biological characteristics of the test sample.

[0260] The reagents and equipment used in this test example are shown below:

[0261] [ 125 I]GLP-1, [ 125 I]GCGR、[ 125 [I] GIP sourced from Revvity; HEPES from Gibco; EGTA, BSA, and MgCl2 from Aladdin; CaCl2, Casein, PEI, Tween 20, and DMSO from Sigma; Tris-HCl from BBI; Microscint-O from PerkinElmer; GLP-1 (7-37), Glucagon, and GIP from HaoYuan; sealing film and 96GF / C filter plates from Revvity; 96-well polypropylene plates from Agilent; plate reader and plate washer from PerkinElmer.

[0262] The reagents used in this test example were prepared as follows:

[0263] Detection buffer (GLP-1R): 50 mM HEPES, 5 mM EGTA, 5 mM MgCl2, 0.005% Tween-20, store at 4°C.

[0264] Detection buffer (GIPR): 50 mM HEPES pH 7.4, 5 mM MgCl2, 1 mM CaCl2, 0.1% Casein, store at 4°C.

[0265] Detection buffer (GCGR): 50 mM HEPES, 5 mM EGTA, 5 mM MgCl2, 0.005% Tween-20, 0.2% Casein, pH 7.4, store at 4°C.

[0266] Plate washing buffer (GLP-1R): 50 mM HEPES, 5 mM EGTA, 5 mM MgCl2, 0.005% Tween-20, pH 7.4, store at 4°C.

[0267] Wash plate buffer (GIPR): 50 mM Tris-HCl pH 7.4, 125 mM NaCl, 0.05% BSA, store at 4°C.

[0268] Wash buffer (GCGR): 50 mM HEPES, 500 mM NaCl, 0.1% BSA, pH 7.4, store at 4°C.

[0269] Plate soaking buffer (GLP-1R): 50 mM HEPES, 0.5% BSA, pH 7.4, store at 4°C.

[0270] Infiltration buffer (GIPR): 0.3% PEI, store at 4°C.

[0271] Dip buffer (GCGR): 0.3% PEI, store at 4°C.

[0272] The cell lines used in this test were all derived from WuXi AppTec, and detailed information is shown in Table 3.

[0273] The procedure for the receptor affinity test in this example is as follows:

[0274] (1) Prepare the sample as follows:

[0275] a) Weigh the positive references GLP-1(7-37), GIP, and Glucagon, then dissolve them in ultrapure water and DMSO, respectively, and dilute them with detection buffer to working concentrations (2000 nM, 400 nM, and 40000 nM, respectively). Use a liquid workstation to perform 4-fold serial dilutions of the positive references, 10 concentration points, double replicates; the initial and final concentrations of the positive references GLP-1(7-37), GIP, and Glucagon for GLP-1, GIPR, and GCGR were 500 nM, 100 nM, and 10000 nM, respectively. LC represents the highest concentration of the positive reference, and HC represents the detection buffer.

[0276] b) Prepare a 200 μM stock solution of the above test samples using PBS. For human GLP-1R, GIPR, and GCGR target testing, dilute the sample stock solution with detection buffer to working concentrations of 500 nM, 100 nM, and 10000 nM, respectively. Perform gradient dilution of the test samples using detection buffer according to Tables 9, 10, and 11, with a 4-fold gradient, 10 concentration points, and double-duplicate wells. The initial and final concentrations of the compounds are 500 nM, 100 nM, and 10000 nM, respectively.

[0277] Table 9. Gradient dilution protocol for the test compounds (GLP-1R)

[0278] Table 10 Gradient dilution protocol (GIPR) for the test compounds

[0279] Table 11 Gradient dilution scheme for the test compounds (GCGR)

[0280] c) Conduct three formal experiments according to the detection method.

[0281] (2) Experimental operation steps:

[0282] 1) Perform serial dilution of the sample to be tested and transfer 25 μL to the detection plate.

[0283] 2) Prepare the previously prepared GLP-1, GIP, and GCG receptor cell membranes to the specified concentration using detection buffer, and add them to the detection plate (96-well conical polypropylene plate), mix them with the cell membranes, 50 μL per well.

[0284] 3) [ 125 I]-GLP-1, [ 125 I]-GIP, [ 125 I]-Glucagon was prepared into 600pM, 160pM, and 240pM working solutions using detection buffer and added to the detection plate, 25μL per well.

[0285] 4) Seal the 96-well plate with sealing film and incubate it on a shaker at room temperature for 1 hour. At the same time, soak the GF / C filter plate (Unifilter-96GF / C filter plate) in soaking buffer for 0.5 hours.

[0286] 5) After incubation, collect the reaction solution onto a GF / C filter plate using a cell collector, wash 6 times with washing buffer, and dry in a 50°C oven for 1 hour.

[0287] 6) Seal the bottom of the dried GF / C filter plate with a membrane, add 50 μL of scintillation fluid to each well, and seal it.

[0288] 7) Using Microbeta 2 Take the readings.

[0289] 8) Results Analysis:

[0290] Using the raw data directly, we fitted the IC to the "log(antagonist) vs. response - Variable slope" model in GraphPad Prism 5 and calculated it. 50 The results are shown in Table 12, where IC 50 It is expressed as geometric mean ± standard error (SEM), with the number of repetitions (n) shown in parentheses. NA indicates that it is not applicable.

[0291] Table 12

[0292] This experiment tested the affinity of human GLP-1, GIP, and GCG receptors. The control GLP-1 (7-37), GIP, Glucagon, sample compound 7, and compound 1 were tested. Under the conditions of this test, compound 7 showed a much stronger affinity for human GCG receptors than compound 1.

[0293] Test Example 3: In vitro activity test

[0294] The main objective of this study was to investigate the EC50 of the agonistic activity of samples in human glucagon-like peptide-1 receptor (GLP-1R), gastric inhibitory peptide receptor (GIPR), and glucagon receptor (GCGR) cell function experiments using the cAMP assay method, thereby providing reference information for the biological characteristics of the test samples.

[0295] The reagents and equipment used in this test example are shown below:

[0296] cAMP detection kit was purchased from CisBio, containing 1M HEPES, 1 ╳ HBSS was purchased from Invitrogen, IBMX and Casein solution from bovine milk were purchased from Sigma, PP-384 well plate was purchased from Greiner, OptiPlate-384 and Plate sealing film were purchased from PerkinElmer, EnVision (Envision2105) was supplied by PerkinElmer, cell counter (Rigel S2) was supplied by Countstar, Bravo (Bravo V11) and Agilent Incubator (371) were supplied by Thermo, and Centrifuge (Allegra-X15R) was supplied by Beckman.

[0297] The cell lines used in this test case were all derived from WuXi AppTec, and detailed information is shown in Table 3. The culture medium for the cell lines was:

[0298] The culture medium for Human GLP-1R / HEK293 consisted of the following components: 10% FBS, 600 μg / mL G418, 6 μg / mL blasticidin, and 1% P / S in DMEM medium.

[0299] The culture medium for Human GIPR / CHO-K1 consisted of the following components: 10% FBS, 600 μg / mL G418, and 1% P / S in F12 medium;

[0300] The culture medium for Human GCGR / HEK293 consisted of the following components: 10% FBS, 300 μg / mL G418, 2 μg / mL blasticidin, 5 μg / mL HB, and 1% P / S in DMEM medium.

[0301] The experimental procedure for this test case is as follows:

[0302] (1) Preparation of detection buffer: HBSS, 20mM HEPES, 500μM IBMX, 0.1% Casein, pH 7.4.

[0303] (2) Compound dilution, the steps of which include:

[0304] Weigh the positive reference GLP-1(7-37), GIP, and Glucagon to prepare a 1 mM stock solution. Dilute the stock solution with the detection buffer to the working concentrations shown in Table 13. Use a liquid workstation to perform a 4-fold serial dilution of the positive reference, resulting in 10 concentration points in duplicate wells. The initial concentrations were 20 nM, 100 nM, and 10 nM. HC represents the corresponding positive reference at the initial concentration, and LC represents the detection buffer.

[0305] The above-mentioned test samples were prepared into a 200 μM stock solution using PBS. For the human GLP-1R / GIPR / GCGR target assay, the sample stock solution was diluted with detection buffer to working concentrations of 40 nM, 200 nM, and 20 nM, respectively. The working concentrations were then used to perform gradient dilutions of the test samples according to Tables 14, 15, and 16, with a 4-fold gradient, 10 concentration points, and double replicates. The initial compound concentrations were 20 nM, 100 nM, and 10 nM, respectively.

[0306] Table 13 Positive Reference Gradient Dilution Protocol

[0307] Table 14 Gradient Dilution Protocol for Test Compounds (GLP-1R)

[0308] Table 15 Gradient dilution scheme (GIPR) for the test compounds

[0309] Table 16 Gradient Dilution Protocol for Test Compounds (GCGR)

[0310] (3) Cell resuscitation: Resuscitate the stable cell line required in a sterile water bath at 37°C, and gently shake until the ice is completely melted.

[0311] (4) Removal of DMSO: Transfer the revived cells to a 15 mL sterile centrifuge tube, add about 10 mL of HBSS preheated to 37 °C, mix gently, centrifuge at 1000 rpm for 5 minutes, and discard the supernatant. Add about 10 mL of HBSS again, mix gently, centrifuge at 1000 rpm for 5 minutes, and discard the supernatant.

[0312] (5) Cell counting: Resuspend the cells in approximately 1 mL of detection buffer, count them using a cell counter, and dilute the hGLP-1R / hGCGR cells to 2.0 × 10⁻⁶ cells with detection buffer. 5 hGIPR cells were diluted to 4.0 × 10⁶ / mL. 5 / mL.

[0313] (6) Sample addition: According to the dilution scheme and arrangement of the compounds, add 5 μL of each prepared and diluted test compound and positive reference solution to the 384-well reaction plate using Bravo.

[0314] (7) Sample incubation: Transfer cells to reaction plates with 1000, 2000 and 1000 human GLP-1R / GIPR / GCGR cells per well, respectively, and load 5 μL. Seal with transparent sealing film and incubate at 23°C for 30 minutes.

[0315] (8) cAMP incubation: Add 10 μL of cAMP detection working solution to the corresponding well of the reaction plate using an electroporator. Cover the plate and incubate at room temperature in the dark for 1 hour.

[0316] (9) Reading: After incubation for 1 hour, read the value in an EnVision microplate reader. The final value is the ratio of the emitted light at 665 nm to that at 615 nm.

[0317] (10) Results Analysis:

[0318] The EC was calculated using the "log(antagonist) vs. response -- variable slope" model from GraphPad Prism. 50 .

[0319] Response (% activity) = 100 × (original sample value - Low Control average) / (High Control average - Low Control average);

[0320] The signal ratio of the reference standard or sample = the average of the raw data at the maximum dose / the average of the raw data at the minimum dose.

[0321] Analysis revealed the EC effects of each compound on each target site. 50The values ​​and maximum activity are shown in Table 17, where EC 50 The value represents the geometric mean ± SEM, and n represents the number of repeated measurements; maximum activity: arithmetic mean ± SEM.

[0322] Table 17

[0323] The agonist activities of compound 7 and compound 1 against human GLP-1R, GIPR, and GCGR targets were evaluated using cAMP assays. In three independent replicate experiments, all EC50 values ​​were [data missing]. 50 The values ​​were all within three times the average value, which meets the acceptable standard for in vitro experiments, indicating that the test system is stable and reliable.

[0324] Compound 7 has a similar activation effect on GLP-1R and GIPR as compound 1, and a stronger activation effect on GCGR than compound 1.

[0325] Test Example 4: Pharmacokinetic Study

[0326] Three male SD rats in each group were administered a single subcutaneous injection of the compound at 0.5 mg / kg or 1 mg / kg. Plasma samples were collected from the animals at 15 min, 30 min, 1 h, 2 h, 4 h, 8 h, 24 h, 36 h, 48 h, 72 h, and 96 h post-administration. The concentration of the corresponding parent compound in the plasma samples was determined using LC-MS / MS. The plasma drug concentration data of the compound were processed using WinNonlin pharmacokinetic software with a non-compartmental model. Relevant pharmacokinetic parameters were calculated using the linear logarithmic trapezoidal method, as shown in Tables 18 and 19.

[0327] Table 18

[0328] Table 19

[0329] As shown in Tables 18-19, the polypeptide compounds provided in this paper have a long half-life and a high exposure level. The pharmacokinetic study methods for other compounds in this invention are basically the same as those for the test examples, and they also have a long half-life and a high exposure level.

[0330] Test Case 5: In vivo efficacy evaluation of the DIO mouse obesity model

[0331] The efficacy of compounds 1, 2, and 7 was evaluated using a mouse model of disproportionately large (DIO) mice induced by a 60% high-fat diet (animal information is shown in Table 20). Animals were screened based on body weight, and suitable DIO mice were selected and randomly divided into four groups for drug administration. The animal administration regimen is shown in Table 21. Drug administration began on the day of grouping and continued every three days for 28 days; the route of administration was subcutaneous injection (sc) in the back.

[0332] Table 20

[0333] The efficacy of compound 1 (group D), compound 2 (group B), and compound 7 (group C) in a DIO mouse obesity model was evaluated. The grouping information and implementation plan for each group are shown in Table 21.

[0334] Table 21

[0335] Table 22 shows the changes in body weight of the three compounds DIO in a mouse obesity model after 28 days of administration.

[0336] Table 22

[0337] The in vivo efficacy data above show that compounds 2 and 7 achieved weight loss rates of 45% and 46% respectively at the experimental endpoint, significantly better than the control group compound 1 (37%). Compounds 2 and 7 provided by this invention exhibit stable efficacy in mice. Figure 6 shows the weight change curves of mice over time after administration, and Figure 7 shows the relative change curves.

[0338] Furthermore, the effects of compounds 1, 2, and 7 on food intake in mice were evaluated using the aforementioned mouse obesity (DIO) model. The experimental protocols for each group and the changes in food intake in the animals over 28 days of administration are shown in Table 23. The curves showing the changes in food intake in mice over time after administration are shown in Figure 8.

[0339] Table 23

[0340] As can be seen from Table 23 and Figure 8, during the entire experiment, compound 7 had a more sustained and stable effect of inhibiting food intake compared to compound 1 (positive control group).

[0341] The efficacy evaluation methods for other compounds of this invention in the DIO mouse obesity model and the experimental methods for their effects on the food intake of mice are basically the same as those in test example 5. The other compounds showed a higher rate of weight loss in mice at the experimental endpoint and stable efficacy; they also had a relatively long-lasting and stable effect of inhibiting food intake in mice.

[0342] It should be understood that the above embodiments are exemplary and are not intended to encompass all possible implementations included in the claims. Various modifications and changes can be made to the above embodiments without departing from the scope of this disclosure. Similarly, the various technical features of the above embodiments can be arbitrarily combined to form other embodiments of the present invention that may not be explicitly described. Therefore, the above embodiments only illustrate several implementations of the present invention and do not limit the scope of protection of this patent.

Claims

1. A polypeptide compound of formula (I) or a pharmaceutically acceptable salt, ester, solvate, optical isomer, tautomer, isotope label, or prodrug thereof: Y-Xaa2-QGTFTSDY-Xaa 11 -I-Xaa 13 -LDK-Xaa 17 -Xaa 18 -Q-Xaa 20 -Xaa 21 -FIEYLLE-Xaa 29 -Xaa 30 -PSS-Xaa 34 -Xaa 35 -PPPS-R1 formula (I); in, Xaa2 is either Aib or Ac4c; Xaa 11 For S or Dap; Xaa 13 For L or 2-MeL; Xaa 17 K is either unmodified or modified, wherein the modification is: ([2-(2-amino-ethoxy)-ethoxy]-acetyl) a -(γGlu) b -CO-(CH2) c -COOH is conjugated to the ε-amino group of the K side chain, wherein a is 1 or 2, b is 1 or 2, and c is 16 or 18; Xaa 18 It can be A or Y; Xaa 20 For Aib, H, or Q; Xaa 21 The answer is A, E, or D; Xaa 29 For G, Aib, or Q; Xaa 30 For G, Aib, or K; Xaa 34 For G or Aib; Xaa 35 It can be A or L; R1 is NH2 or OH, or a pharmaceutically acceptable salt and / or ester thereof; Furthermore, the compound of formula (I) is not one of the following compounds:

2. The polypeptide compound according to claim 1, or its pharmaceutically acceptable salt, ester, solvate, optical isomer, tautomer, isotope label, or prodrug, characterized in that, Xaa 11 For DAP; Xaa 13 It is 2-MeL; Xaa 20 For Aib; Xaa 29 For G or Aib.

3. The polypeptide compound according to claim 2, or its pharmaceutically acceptable salt, ester, solvate, optical isomer, tautomer, isotope label, or prodrug, characterized in that, Xaa 29 It is Aib.

4. The polypeptide compound according to claim 3, or its pharmaceutically acceptable salt, ester, solvate, optical isomer, tautomer, isotope label, or prodrug, characterized in that, Xaa 21 For A; preferably, Xaa2 is Aib, Xaa 21 Let A be the highest value; further preferably, Xaa2 is Aib, X 18 for A, Xaa 21 The answer is A.

5. The polypeptide compound according to claim 3, or its pharmaceutically acceptable salt, ester, solvate, optical isomer, tautomer, isotope label, or prodrug, characterized in that, Xaa2 is either Aib or Ac4c, Xaa 21 The preferred value is D; preferably, Xaa2 is Ac4c, Xaa 21 The answer is D.

6. The polypeptide compound according to claim 1, or its pharmaceutically acceptable salt, ester, solvate, optical isomer, tautomer, isotope label, or prodrug, characterized in that, Xaa2 is Aib; Xaa 11 S; Xaa 18 is A; Xaa 21 is E; Xaa 30 G; Xaa 34 G; Xaa 35 is A.

7. The polypeptide compound according to claim 6, or its pharmaceutically acceptable salt, ester, solvate, optical isomer, tautomer, isotope label, or prodrug, characterized in that, Xaa 13 For 2-MeL, Xaa 29 For G or Aib.

8. The polypeptide compound according to claim 7, or a pharmaceutically acceptable salt, ester, solvate, optical isomer, tautomer, isotope label, or prodrug thereof, characterized in that, Xaa 20 For Aib or H; Xaa 29 It is Aib.

9. The polypeptide compound according to any one of claims 1 to 8, or a pharmaceutically acceptable salt, ester, solvate, optical isomer, tautomer, isotope label, or prodrug thereof, characterized in that, Xaa 17 Let K be the modified form. The structural formula of the modified K is shown below: Preferably, R1 is NH2.

10. A polypeptide compound as described in claim 1, or a pharmaceutically acceptable salt, ester, solvate, optical isomer, tautomer, isotope label, or prodrug thereof, characterized in that, The polypeptide compound is selected from any of the following:

11. The polypeptide compound according to claim 10, or a pharmaceutically acceptable salt, ester, solvate, optical isomer, tautomer, isotope label, or prodrug thereof, characterized in that, The polypeptide compound is selected from any of the following:

12. A pharmaceutical composition comprising the polypeptide compound of any one of claims 1 to 11 or a pharmaceutically acceptable salt, ester, solvate, optical isomer, tautomer, isotope label, or prodrug thereof, and optionally, at least one pharmaceutically acceptable carrier.

13. The polypeptide compound of any one of claims 1 to 11 or its pharmaceutically acceptable salt, ester, solvate, optical isomer, tautomer, isotope label or prodrug and / or the pharmaceutical composition of claim 12 as an agonist of GLP-1, GCG, or GIP receptors.

14. Use of the polypeptide compound of any one of claims 1 to 11 and / or the pharmaceutical composition of claim 12 in the preparation of medicaments for the treatment and / or prevention of metabolic disorders, bone-related diseases, cardiovascular diseases and neurodegenerative diseases.

15. The use according to claim 14, characterized in that, The metabolic disorders mentioned include obesity, diabetes, dyslipidemia, fatty liver disease, metabolic syndrome, non-alcoholic steatohepatitis, and non-alcoholic fatty liver disease. Preferably, the diabetes includes type II diabetes; Preferably, the neurodegenerative diseases include Alzheimer's disease and Parkinson's disease.

16. A method for treating metabolic disorder-related diseases or neurodegenerative diseases, characterized in that, The method comprises administering to a subject an effective dose of the polypeptide compound of any one of claims 1 to 11 or a pharmaceutically acceptable salt, ester, solvate, optical isomer, tautomer, isotope label, or prodrug and / or the pharmaceutical composition of claim 12; Preferably, the metabolic disorder-related diseases include obesity, diabetes, dyslipidemia-related diseases, fatty liver disease, metabolic syndrome, non-alcoholic steatohepatitis, and non-alcoholic fatty liver disease; Preferably, the diabetes includes type II diabetes; Preferably, the neurodegenerative diseases include Alzheimer's disease and Parkinson's disease.

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