Corticotropin releasing factor 2 receptor agonist and use thereof

Abstract

Provided are a corticotropin releasing factor 2 receptor agonist and use thereof. The corticotropin releasing factor 2 receptor agonist can be used for preventing and / or treating heart failure, hypertension, obesity, sarcopenia, diabetes, chronic kidney disease, and related diseases.

Description

Adrenocorticotropic hormone-releasing factor 2 receptor agonists and their applications Technical Field

[0001] This disclosure pertains to the field of biomedicine and relates to corticotropin-releasing factor 2 receptor (CRF2R) agonists and their applications. Background Technology

[0002] The continued rise in the global obesity population has driven the rapid development of the weight loss product market, with GLP-1 and GIP-targeted drugs exhibiting fierce competition. However, studies have shown that nearly 40% of the weight loss in these drug users is muscle, and muscle loss increases the risk of cardiovascular disease, osteoporosis, and other diseases. Therefore, muscle loss remains an unmet clinical need for addressing metabolic diseases, and the value of GLP-1, a "next-generation" target for improving muscle mass, is gradually gaining attention.

[0003] The corticotropin-releasing factor (CRF) family comprises four members (CRF, Urocortin1, Urocortin2, and Urocortin3), which function physiologically through two distinct G protein-coupled receptors (CRHR1 / CRF1R / CRFR1 and CRHR2 / CRF2R / CRFR2). Urocortin2 (UCN2) is a selective agonist of CRF2R and is highly expressed in skeletal muscle. UCN2 has been shown to activate cAMP signaling and promote skeletal muscle anabolism and muscle gain by activating the Akt / mTOR and ERK1 / 2 signaling pathways. Simultaneously, UCN2 can induce motor inhibition and reduce food and water intake. These findings open new avenues for treating muscle mass loss.

[0004] Neurocrine Biosciences' NBI-69734, a UCN2 peptide, has been used clinically in the treatment of congestive heart failure (CHF) and in studies on its use in treating hypertension. However, NBI-69734 has a very short half-life (t1 / 2 = 15.36 min), requiring intravenous injection, which leads to poor patient compliance. Summary of the Invention

[0005] This disclosure provides a novel long-acting corticotropin-releasing factor 2 receptor (CRF2R) agonist and a pharmaceutical composition thereof; at the same time, this disclosure provides a method for preparing a medicament for treating CRF2R-related diseases using the above-mentioned agonist or composition, a method for treating diseases, or a use thereof, thereby greatly reducing the frequency of administration and improving patient compliance and experience.

[0006] In a first aspect, this disclosure provides an adrenocorticotropic hormone-releasing factor 2 receptor agonist comprising the amino acid sequence of Formula 1:

[0007] KIVLSLDVPIGLLQILLEQX 20 X 21 X 22 X 23 X 24 X 25 X 26 X 27 QAKTNAEILERV (Formula 1)

[0008] in,

[0009] X 20 Selected from A or E, X 21 Selected from R or K, X 22 Choose from A or Q, X 23 Selected from R or E, X 24 Selected from A or K, X 25 Selected from A or E, X 26 Selected from R or K, and / or X 27 Selected from E or Q;

[0010] Optionally, the carboxyl terminus of the amino acid sequence of Formula 1 is modified with NH2.

[0011] In some embodiments, in the above-mentioned corticotropin-releasing factor 2 receptor agonist, the amino acid sequence of Formula 1,

[0012] X 20 Let A and X be the two numbers. 21 Let R, X 22 Let A and X be the two numbers. 23 Let R, X 24 Let A and X be the two numbers. 25 Let A and X be the two numbers. 26 Let R and X be the numbers. 27 E;

[0013] The amino acid sequence of Formula 1 is shown in SEQ ID NO:2.

[0014] In some embodiments, in the above-mentioned corticotropin-releasing factor 2 receptor agonist, the amino acid sequence of Formula 1,

[0015] X 20 For E, X 21 Let K, X 22 For Q, X 23 For E, X 24 Let K, X 25 For E, X 26 Let K and X be the numbers. 27 For Q;

[0016] The amino acid sequence of Formula 1 is shown in SEQ ID NO:3.

[0017] In this invention, receptor agonists refer to any compound that can activate a receptor, and adrenocorticotropic hormone-releasing factor 2 receptor agonists include, but are not limited to, polypeptides, polypeptide derivatives (such as fatty acid-modified polypeptides), or pharmaceutically acceptable salts thereof.

[0018] In some embodiments, the aforementioned corticotropin-releasing factor 2 receptor agonist is a polypeptide derivative or a pharmaceutically acceptable salt thereof.

[0019] In some embodiments, in any of the above-described adrenocorticotropic hormone-releasing factor 2 receptor agonists, at least one amino acid residue in the amino acid sequence of Formula 1 is attached to a fatty acid side chain, preferably an amino acid K residue is attached to a fatty acid side chain, more preferably an amino acid K residue at position 1 and / or position 30 is attached to a fatty acid side chain.

[0020] In some embodiments, in any of the above-described adrenocorticotropic hormone-releasing factor 2 receptor agonists, the fatty acid side chain is linked by the ε-amino group of the amino acid K residue of the amino acid sequence of Formula 1, for example, by the ε-amino group of the amino acid K residue at position 1 and / or position 30 of the amino acid sequence of Formula 1.

[0021] In some embodiments, in any of the above-described corticotropin-releasing factor 2 receptor agonists, the fatty acid side chain is selected from...

[0022] One or more of the following, where x is any integer from 4 to 38;

[0023] Preferably, the fatty acid side chain is selected from:

[0024] HOOC(CH2) 14 CO-, HOOC(CH2) 15 CO-, HOOC(CH2) 16 CO-, HOOC(CH2) 17 CO-, HOOC(CH2) 18 CO-, HOOC(CH2) 19 CO-, HOOC(CH2) 20 CO-, HOOC(CH2) 21 CO- and HOOC(CH2) 22 One or more of CO-.

[0025] In some embodiments, in any of the above-described corticotropin-releasing factor 2 receptor agonists, the connection is a direct connection or an indirect connection (e.g., a connection via a connector).

[0026] In some embodiments, in any of the above-described corticotropin-releasing factor 2 receptor agonists, the fatty acid side chain is linked to an amino acid residue of the amino acid sequence of Formula 1 via a linker, for example, to an amino acid K residue, or more specifically, to an amino acid K residue at position 1 and / or position 30.

[0027] In some embodiments, in the above-mentioned corticotropin-releasing factor 2 receptor agonists, the linker is selected from... One or more of the following, where m is 0, 1, 2 or 3; n is 1 or 2; p is any integer from 1 to 5;

[0028] Preferably, the connector is:

[0029] In some embodiments, the corticotropin-releasing factor 2 receptor agonist is compound 1 of this disclosure, whose amino acid sequence is shown in SEQ ID NO:2, and the ε-amino group on the K residue of the first amino acid is bonded to (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-(γGlu)3-CO-(CH2) via an amide bond. 16 -CO2H (English name: 17-{[(22S,27S,32S)-22,27,32-tricarboxy-1,10,19,24,29-pentaoxo-9,18,23,28-tetraza-3,6,12,15-tetraoxadotriacont-32-yl]amino}-17-oxoheptadecane-1-carboxylic) acid, Chinese name: 17-{[(22S,27S,32S)-22,27,32-tricarboxylic acid-1,10,19,24,29-pentaoxylidene-9,18,23,28-tetraaza-3,6,12,15-tetraoxa-tetrazole-32-yl]amino}-17-oxylidene-heptadecane-1-carboxylic acid) linked by (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-(γGlu)3-CO-(CH2) 16 The structural formula of -CO2H is shown in formula (I):

[0030] In some embodiments, the corticotropin-releasing factor 2 receptor agonist is compound 2 of this disclosure, whose amino acid sequence is shown in SEQ ID NO:2, and the ε-amino groups on the K residues of amino acids at positions 1 and 30 are respectively bonded by amide bonds to (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-(γGlu)2-CO-(CH2). 14 -CO2H (English name: 15-{[(22S,27S)-22,27-dicarboxy-1,10,19,24-tetraoxo-3,6,12,15-tetraoxa-9,18,23-triazaheptacos-27-yl]amino}-15-oxopentadecane-1-carboxylic acid, Chinese name: 15-{[(22S,27S)-22,27-dicarboxy-1,10,19,24-tetraoxo-3,6,12,15-tetraoxa-9,18,23-triazaheptacos-27-yl]amino}-15-oxopentadecane-1-carboxylic acid) linked by (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-(γGlu)2-CO-(CH2) 14 The structural formula of -CO2H is shown in formula (II):

[0031] In some embodiments, the corticotropin-releasing factor 2 receptor agonist is compound 3 of this disclosure, whose amino acid sequence is shown in SEQ ID NO:2, and the ε-amino group on the K residue of the first amino acid is bonded to (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-(γGlu)2-CO-(CH2) via an amide bond. 14 -CO2H linkage, (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-(γGlu)2-CO-(CH2) 14 The structural formula of -CO2H is shown in formula (II); the ε-amino group on the K residue of the 30th amino acid is bonded to (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-(γGlu)-CO-(CH2) via an amide bond. 14-CO2H (English name: 16-{[(22S)-22-carboxy-1,10,19-trioxo-9,18-diaza-3,6,12,15-tetraoxadocos-22-yl]amino}-16-oxohexadecanoic acid, Chinese name: 16-{[(22S)-22-carboxyl-1,10,19-trioxo-9,18-diaza-3,6,12,15-tetraoxadocos-22-yl]amino}-16-oxo-ylidene hexadecanoic acid) linked by (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-(γGlu)-CO-(CH2) 14 The structural formula of -CO2H is shown in formula (III):

[0032] In some embodiments, the corticotropin-releasing factor 2 receptor agonist is compound 4 of this disclosure, whose amino acid sequence is shown in SEQ ID NO:3, and the ε-amino group on the K residue of the first amino acid is bonded to (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-(γGlu)3-CO-(CH2) via an amide bond. 16 -CO2H linkage, (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-(γGlu)3-CO-(CH2) 16 The structural formula of -CO2H is shown in formula (I).

[0033] In some embodiments, the corticotropin-releasing factor 2 receptor agonist is compound 5 of this disclosure, whose amino acid sequence is shown in SEQ ID NO:3, and the ε-amino groups on the K residues of amino acids at positions 1 and 30 are respectively bonded by amide bonds to (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-(γGlu)2-CO-(CH2). 14 -CO2H linkage, (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-(γGlu)2-CO-(CH2) 14 The structural formula of -CO2H is shown in formula (II).

[0034] In some embodiments, the corticotropin-releasing factor 2 receptor agonist is compound 6 of this disclosure, whose amino acid sequence is shown in SEQ ID NO:3, and the ε-amino group on the K residue of the first amino acid is bonded to (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-(γGlu)2-CO-(CH2) via an amide bond. 14 -CO2H linkage, (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-(γGlu)2-CO-(CH2) 14 The structural formula of -CO2H is shown in formula (II); the ε-amino group on the K residue of the 30th amino acid is bonded to (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-(γGlu)-CO-(CH2) via an amide bond. 14 -CO2H linkage, (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-(γGlu)-CO-(CH2) 14 The structural formula of -CO2H is shown in formula (III).

[0035] In some embodiments, the corticotropin-releasing factor 2 receptor agonist is compound 7 of this disclosure, whose amino acid sequence is shown in SEQ ID NO:2, and the ε-amino group on the K residue of the first amino acid is bonded to (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-(γGlu)2-CO-(CH2) via an amide bond. 16 -CO2H linkage, (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-(γGlu)2-CO-(CH2) 16 The structural formula of -CO2H is shown in formula (IV):

[0036] In some embodiments, the corticotropin-releasing factor 2 receptor agonist is compound 8 of this disclosure, whose amino acid sequence is shown in SEQ ID NO:2, and the ε-amino group on the K residue of the first amino acid is bonded to (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-γGlu-CO-(CH2) via an amide bond. 16 -CO2H linkage, (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-γGlu-CO-(CH2) 16 The structural formula of -CO2H is shown in formula (V):

[0037] In some embodiments, the corticotropin-releasing factor 2 receptor agonist is compound 9 of this disclosure, whose amino acid sequence is shown in SEQ ID NO:2, and the ε-amino group on the K residue of the first amino acid is bonded by an amide bond to (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-(γGlu)3-CO-(CH2). 18 -CO2H linkage, (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-(γGlu)3-CO-(CH2) 18 The structural formula of -CO2H is shown in formula (VI):

[0038] In some embodiments, the corticotropin-releasing factor 2 receptor agonist is compound 10 of this disclosure, having the amino acid sequence shown in SEQ ID NO:2, and wherein the ε-amino group at the K residue of the first amino acid is bonded by an amide bond to (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-βAsp-CO-(CH2). 16 -CO2H linkage, (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-βAsp-CO-(CH2) 16 The structural formula of -CO2H is shown in formula (VII):

[0039] In some embodiments, the corticotropin-releasing factor 2 receptor agonist is compound 11 of this disclosure, whose amino acid sequence is shown in SEQ ID NO:2, and the ε-amino group on the K residue of the first amino acid is bonded to (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-(γGlu)2-CO-(CH2) via an amide bond. 18 -CO2H linkage, (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-(γGlu)2-CO-(CH2) 18 The structural formula of -CO2H is shown in formula (VIII):

[0040] In some embodiments, the corticotropin-releasing factor 2 receptor agonist is compound 12 of this disclosure, having the amino acid sequence shown in SEQ ID NO:2, and wherein the ε-amino group at the K residue of the first amino acid is bonded to (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-βAsp-CO-(CH2) via an amide bond. 18-CO2H linkage, (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-βAsp-CO-(CH2) 18 The structural formula of -CO2H is shown in equation (IX):

[0041] In a second aspect, this disclosure provides a polypeptide having an amino acid sequence as shown in Formula 1 above. Preferably, the amino acid sequence of the polypeptide is shown in SEQ ID NO:2 or 3.

[0042] In a third aspect, this disclosure provides a nucleic acid molecule that encodes the aforementioned polypeptide.

[0043] In a fourth aspect, this disclosure provides a recombinant vector comprising the aforementioned nucleic acid molecules.

[0044] In a fifth aspect, this disclosure provides a recombinant cell comprising the aforementioned nucleic acid molecule or the aforementioned recombinant vector; preferably, the recombinant cell is capable of expressing and optionally secreting the aforementioned polypeptide.

[0045] In a sixth aspect, this disclosure provides a method for preparing any of the above-described corticotropin-releasing factor 2 receptor agonists or the above-described polypeptides;

[0046] The method includes the step of preparing the adrenocorticotropic hormone-releasing factor 2 receptor agonist or polypeptide using chemical and / or biological methods;

[0047] Preferably, the chemical method includes liquid-phase or solid-phase peptide synthesis; the biological method includes molecular biology methods and / or cell biology methods.

[0048] In a seventh aspect, this disclosure provides a pharmaceutical composition comprising any of the above-described corticotropin-releasing factor 2 receptor agonists or the above-described polypeptides, and pharmaceutically acceptable excipients.

[0049] The pharmaceutical compositions disclosed herein may be administered via any suitable route known in the art, including but not limited to: oral, nasal, intradermal, subcutaneous, intravenous, intramuscular, intrabronchial, intrapleural, intraperitoneal, intraarterial, lymphatic, and / or cerebrospinal fluid administration.

[0050] In some embodiments, the pharmaceutical compositions of this disclosure further comprise other active ingredients, including but not limited to glucagon-like peptide-1 receptor (GLP-1R) agonists, amylin receptor agonists, glucose-dependent insulinotropic peptide receptor (GIPR) agonists or antagonists, glucagon receptor (GCGR) agonists, or ingredients having agonistic or antagonistic activity against two or more of the above receptors simultaneously (e.g., GLP-1R / GIPR dual agonists, GLP-1R agonist / GIPR antagonist, GLP-1R / GCGR dual agonist, GLP-1R / GIPR / GCGR triple agonist, GLP-1R agonist / GIPR antagonist / GCGR agonist, GLP-1R / amylin receptor agonist, GLP-1R / GIPR / amylin receptor triple agonist, GLP-1R agonist / GIPR antagonist / amylin receptor agonist).

[0051] In some embodiments, the pharmaceutical compositions disclosed herein further comprise a glucagon-like peptide-1 receptor agonist, such as Ecnoglutide.

[0052] In some embodiments, the pharmaceutical compositions disclosed herein may be used in combination with one or more other pharmaceutical compositions. In some embodiments, the other one or more pharmaceutical compositions include, but are not limited to, glucagon-like peptide-1 receptor (GLP-1R) agonists, amylin receptor agonists, glucose-dependent insulinotropic peptide receptor (GIPR) agonists or antagonists, glucagon receptor (GCGR) agonists, or components having simultaneous agonistic or antagonistic activity against two or more of the above receptors (e.g., GLP-1R / GIPR dual agonists, GLP-1R agonist / GIPR antagonist, GLP-1R / GCGR dual agonists, GLP-1R / GIPR / GCGR triple agonists, GLP-1R agonist / GIPR antagonist / GCGR agonist, GLP-1R / amylin receptor agonist, GLP-1R / GIPR / amylin receptor triple agonist, GLP-1R agonist / GIPR antagonist / amylin receptor agonist). In some embodiments, the additional one or more pharmaceutical compositions comprise a glucagon-like peptide-1 receptor agonist, such as Ecnoglutide.

[0053] In some embodiments, different pharmaceutical compositions may be administered simultaneously, sequentially, or separately to patients in need. In some embodiments, different pharmaceutical compositions may be administered sequentially to patients in need, for example, on days 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 1 month, 2 months, 3 months, or longer, either simultaneously, sequentially, or separately, once, twice, three times, or more per day.

[0054] In an eighth aspect, this disclosure provides the use of any of the above-described corticotropin-releasing factor 2 receptor agonists, the above-described polypeptides, or any of the above-described pharmaceutical compositions in the preparation of medicaments for the prevention and / or treatment of corticotropin-releasing factor 2 receptor-mediated diseases.

[0055] In some implementations, the disease described above is one or more of heart failure, hypertension, obesity, sarcopenia, diabetes, and chronic kidney disease.

[0056] In some implementations, the disease described above is one or more of heart failure, hypertension, obesity, sarcopenia, diabetes, chronic kidney disease, and related diseases.

[0057] In a ninth aspect, this disclosure also provides a method for preventing and / or treating diseases mediated by corticotropin-releasing factor 2 receptor, comprising administering to a patient in need a therapeutically effective amount of any of the above-described corticotropin-releasing factor 2 receptor agonists, the above-described polypeptides, or any of the above-described pharmaceutical compositions.

[0058] In some implementations, the disease described above is one or more of the following: heart failure, hypertension, obesity, sarcopenia, diabetes, and chronic kidney disease.

[0059] In some implementations, the disease described above is one or more of heart failure, hypertension, obesity, sarcopenia, diabetes, chronic kidney disease, and related diseases.

[0060] The medicament disclosed herein may be administered to the patient (e.g., a mammal, such as a human) via any suitable route known in the art, including but not limited to: oral, nasal, intradermal, subcutaneous, intravenous, intramuscular, intrabronchial, intrapleural, intraperitoneal, intraarterial, lymphatic, and / or cerebrospinal fluid administration.

[0061] In some embodiments, in any of the methods described above, the administration cycle of the corticotropin-releasing factor 2 receptor agonist, the polypeptide, or any of the pharmaceutical compositions described above is once or more daily, weekly, bi-weekly, tri-weekly, monthly, 2-monthly, 3-monthly, 4-monthly, 5-monthly, 6-monthly, 7-monthly, 8-monthly, 9-monthly, 10-monthly, 11-monthly, or 12-monthly, for example, daily, weekly, bi-weekly, tri-weekly, monthly, 2-monthly, 3-monthly, 4-monthly, or monthly. Apply 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, or 50 times every 5 months, 6 months, 7 months, 8 months, 9 months, 10 months, 11 months, or 12 months.

[0062] This disclosure also provides a kit comprising any of the above-described corticotropin-releasing factor 2 receptor agonists, or the above-described polypeptides, or any of the above-described pharmaceutical compositions, and optionally instructions for use.

[0063] This disclosure provides any of the above-described corticotropin-releasing factor 2 receptor agonists, the above-described polypeptides, any of the above-described pharmaceutical compositions, or the above-described kits for the prevention and / or treatment of diseases mediated by corticotropin-releasing factor 2 receptors. Attached Figure Description

[0064] Figure 1 shows the activity of the compound on CRF1R cells.

[0065] Figure 2 shows the weight reduction effect of the compound.

[0066] Figure 3 shows the effect of the compound on inhibiting food intake.

[0067] Figure 4 shows the weight loss and food intake inhibition effects of a single dose of the compound in rats, where A represents the weight loss effect and B represents the food intake inhibition effect.

[0068] Figure 5 shows the weight loss effect of the compound combined with Ecnoglutide.

[0069] Figure 6 shows the fat loss and muscle gain effects of the compound combined with Ecnoglutide.

[0070] Figure 7 shows the weight loss effect of different doses of the compound combined with Ecnoglutide.

[0071] Figure 8 shows the fat reduction and muscle gain effects of different doses of the compound combined with Ecnoglutide.

[0072] Figure 9 shows the effects of multiple administrations of the compound on weight loss and food intake inhibition in DIO model mice (E1-E5 groups), where A represents weight loss and B represents food intake inhibition.

[0073] Figure 10 shows the fat reduction and muscle gain effects of the compound after multiple administrations to DIO model mice (E1-E5 groups).

[0074] Figure 11 shows the effects of multiple administrations of the compound on weight loss and food intake inhibition in DIO model mice (F1-F4 groups), where A represents weight loss and B represents food intake inhibition.

[0075] Figure 12 shows the fat reduction and muscle gain effects of the compound after multiple administrations to DIO model mice (F1-F4 groups).

[0076] Figure 13 shows the weight loss effect of the DIO model mice in Example 7.

[0077] Figure 14 shows the fat loss and muscle gain effects of the DIO model mice in Example 7, where A represents the lean body mass change rate and B represents the fat mass change rate.

[0078] Figure 15 shows the body fat ratio of the DIO model mice in Example 7. Detailed Implementation

[0079] Unless otherwise specified, the experimental methods used in the following examples are conventional methods.

[0080] Unless otherwise specified, all materials and reagents used in the following examples are commercially available.

[0081] The present disclosure will be further described below with reference to specific embodiments. It should be understood that the following embodiments are for illustrative purposes only and are not intended to limit the scope of the present disclosure.

[0082] definition

[0083] Unless otherwise stated, the terms used herein have the following definitions.

[0084] In this document, the standard single letters used for natural amino acids are employed. All amino acid residues in the peptides disclosed herein are L-configured unless otherwise specified.

[0085] The terms “polypeptide” or “protein” may be used interchangeably. A “polypeptide” or “protein” is any chain containing two or more amino acids, regardless of post-translational modifications (e.g., glycosylation or phosphorylation), including naturally occurring or non-naturally occurring (e.g., artificially synthesized) amino acids or amino acid analogs, wherein the amino acids in any chain are covalently linked by peptide bonds.

[0086] The terms "fatty acid modified peptide" or "peptide derivative" refer to peptides that contain fatty acid side chains (including fatty acid-like structural side chains) that are modified.

[0087] The terms “comprising,” “including,” or “containing” should be understood to include the specified components but do not exclude any other components.

[0088] The terms “patient,” “object,” and “individual” are used interchangeably and include both human and non-human animals, including mammals such as monkeys, rats, mice, cows, pigs, goats, sheep, dogs, and cats.

[0089] When "giving" and "treating" are used to refer to animals, humans, experimental subjects, cells, tissues, organs, or biological fluids, it means contacting an exogenous drug, therapeutic agent, diagnostic agent, or composition with the animal, human, subject, cell, tissue, organ, or biological fluid. "Giving" and "treating" can refer to, for example, methods of treatment, pharmacokinetic methods, diagnostic methods, research methods, and experimental methods. Treating cells includes contacting a reagent with cells and contacting a reagent with a fluid, wherein the fluid contacts the cells. "Giving" and "treating" also mean, for example, in vitro and ex vivo treatment of cells by means of a reagent, diagnostic agent, conjugated composition, or other cells.

[0090] As used herein, “prevention” or “treatment” includes delaying the development of disease-related symptoms and / or reducing the severity of such symptoms that the disease will or is expected to develop. The term also includes alleviating existing symptoms, preventing additional symptoms, and the underlying causes of alleviating or preventing these symptoms. Therefore, the term implies that a beneficial outcome has been attributed to a vertebrate subject suffering from a disease, such as a human.

[0091] As used herein, the term "therapeutic effective dose" or "effective dose" refers to the amount of a corticotropin-releasing factor 2 receptor agonist administered alone or in combination with another therapeutic agent to cells, tissues, or a subject, which effectively prevents or alleviates the disease or condition to be treated. Therapeutic effective dose further refers to the amount of the corticotropin-releasing factor 2 receptor agonist sufficient to cause symptom relief, such relief being, for example, the treatment, cure, prevention, or alleviation of an associated medical condition, or an increase in the rate of treatment, cure, prevention, or alleviation of the symptom. The effective dose for a specific subject can vary depending on a variety of factors, such as the disease to be treated, the patient's overall health condition, the route and dosage of administration, and the severity of side effects. The effective dose may be the maximum dose or administration regimen that avoids significant side effects or toxicity. When administered to an individual as a single active ingredient, the therapeutic effective dose refers to that single ingredient. When administered in combination, the therapeutic effective dose refers to the combined amount of active ingredients that produce the therapeutic effect, regardless of whether they are administered in combination, continuously, or simultaneously. The effective therapeutic dose will typically reduce symptoms by at least 10%; typically by at least 20%; preferably by at least about 30%; more preferably by at least 40% and most preferably by at least 50%.

[0092] Adrenocorticotropic hormone-releasing factor 2 receptor agonists

[0093] This disclosure discloses a corticotropin-releasing factor 2 receptor agonist that exhibits significantly enhanced CRF2R activation activity compared to wild-type human urocortin 2 (hUCN2) and control compounds, while showing no agonistic activity against CRF1R. It is a fatty acid-modified polypeptide or polypeptide derivative.

[0094] Experiments have confirmed that compounds 1, 2, 3, 4, 7, 8, 9, 10, 11, and 12 disclosed herein exhibit better activity than hUCN2 and the control compound in experiments on cells overexpressing human CRF2R receptor, but have no activity in experiments on cells overexpressing human CRF1R receptor.

[0095] It should be understood that the corticotropin-releasing factor 2 receptor agonists of this disclosure may also be provided in the form of salts. Pharmaceutically acceptable salts include salts in anionic and salts in cationic forms. Some examples of anionic salts include hydrochloride, citrate, chloride, and acetate. Preferably, the salt is an acetate. Some examples of cationic salts include salts in which the cation is selected from alkali metals (e.g., sodium and potassium), alkaline earth metals (e.g., calcium), etc.

[0096] In addition, the corticotropin-releasing factor 2 receptor agonist disclosed herein can also interact with metal ions (e.g., Mn). 2+ and Zn 2+The derivative can form a coordination complex, thus existing in the form of a complex. Since the corticotropin-releasing factor 2 receptor agonist of this disclosure contains a hydroxyl or carboxylic acid, the derivative can also react with a suitable carboxylic acid or alcohol to form an ester, thus existing in the form of an ester. The corticotropin-releasing factor 2 receptor agonist of this disclosure can also exist in the form of a prodrug, which can be converted in vivo or in vitro to one of the parent compounds. Generally, at least one biological activity of the corticotropin-releasing factor 2 receptor agonist is reduced in the prodrug form and can be activated by conversion of the prodrug to release the corticotropin-releasing factor 2 receptor agonist or its metabolites. Some examples of prodrugs include the use of a protecting group, which can be removed in situ to release the active compound or to inhibit the clearance of the drug in vivo.

[0097] Synthetic corticotropin-releasing factor 2 receptor agonists

[0098] The disclosed corticotropin-releasing factor 2 receptor agonists can be prepared using chemical and / or biological methods. Chemical methods are preferred, for example, using liquid-phase or solid-phase peptide synthesis techniques to synthesize the disclosed corticotropin-releasing factor 2 receptor agonists. Biological methods include molecular biology methods and cell biology methods.

[0099] The method for preparing the corticotropin-releasing factor 2 receptor agonist disclosed herein may include the following steps:

[0100] The disclosed corticotropin-releasing factor 2 receptor agonist was synthesized stepwise or by fragment assembly according to the polypeptide sequence using liquid-phase or solid-phase polypeptide synthesis methods; or

[0101] A nucleic acid construct encoding a polypeptide sequence of an adrenocorticotropic hormone-releasing factor 2 receptor agonist is introduced into a host cell, and then cultured under certain conditions for a period of time to obtain a polypeptide product of the adrenocorticotropic hormone-releasing factor 2 receptor agonist from the host cell culture. For example, the polypeptide can be obtained by expressing it in a prokaryotic host (e.g., *Escherichia coli*) or a eukaryotic host (e.g., yeast, higher plants, or animals) using recombinant technology, and then modifying the fatty acid side chain of the polypeptide to obtain the adrenocorticotropic hormone-releasing factor 2 receptor agonist of this disclosure; or

[0102] The polypeptide product of the adrenocorticotropic hormone-releasing factor 2 receptor agonist was expressed in a cell-free system using a nucleic acid construct encoding a polypeptide sequence of an adrenocorticotropic hormone-releasing factor 2 receptor agonist, and the polypeptide was modified with fatty acid side chains to obtain the adrenocorticotropic hormone-releasing factor 2 receptor agonist of this disclosure.

[0103] In some embodiments, the corticotropin-releasing factor 2 receptor agonist of this disclosure is prepared by solid-phase peptide synthesis on a suitable resin. Solid-phase peptide synthesis steps are well known in the art, for example, by attaching an N-terminal protected amino acid and its carboxyl terminus to an inert solid support carrying a cleavable linker. This solid support can be any polymer that allows initial amino acid coupling, such as Rink Amide MBHA resin. In some embodiments, starting with Rink Amide MBHA resin, amino acids with Fmoc-protecting groups are sequentially linked according to the solid-phase synthesis method to obtain a protected linear peptide resin, during which the Fmoc-protecting groups are sequentially removed, and a peptide linking reaction is performed using TBTU as a condensing agent to obtain a protected fatty acid-modified peptide resin. The peptide is then cleaved from the resin, followed by simultaneous removal of side-chain protecting groups, separation and purification by column chromatography, and freeze-drying to obtain a powdered purified peptide derivative.

[0104] In some embodiments, the polypeptide sequence of the corticotropin-releasing factor 2 receptor agonist of this disclosure is prepared using recombinant technology. In this case, this disclosure also provides a nucleic acid molecule encoding the polypeptide sequence of the corticotropin-releasing factor 2 receptor agonist of this disclosure, the nucleotide sequence of which may be a codon-optimized sequence according to the host to which it is to be transferred; the nucleic acid molecule may be a DNA fragment or an RNA fragment, which can typically be obtained by amplification using a PCR instrument or by artificial synthesis.

[0105] In some embodiments, the polypeptide sequence of the corticotropin-releasing factor 2 receptor agonist disclosed herein is prepared using recombinant technology. In this case, the present disclosure also provides a recombinant vector comprising the above-described nucleic acid molecule; the recombinant vector comprises a cloning vector and an expression vector, the cloning vector being used to replicate the relevant sequence, and the expression vector being used to express the relevant gene. The vector can be any vector commonly used in the art, such as plasmids, bacteriophages, granules, small chromosomes, or viruses. In addition to the nucleic acid encoding the polypeptide sequence of the corticotropin-releasing factor 2 receptor agonist, the expression vector may include not only a promoter for initiating transcription of the gene encoding the polypeptide sequence, but also a signal peptide sequence, a terminator for terminating transcription of the gene encoding the polypeptide sequence, and an enhancer sequence.

[0106] The method used to construct the recombinant expression vector can be any known method. The promoter described above, the nucleic acid of the gene encoding the polypeptide sequence, and other DNA segments (e.g., terminators, enhancers, if present) can be introduced in a predetermined order into a suitably selected vector used as a basis. For example, the recombinant vector can be constructed using restriction endonucleases and ligases.

[0107] In some embodiments, the polypeptide sequence of the corticotropin-releasing factor 2 receptor agonist disclosed herein is prepared using recombinant technology. In this case, the present disclosure also provides a recombinant cell comprising the above-described recombinant vector, wherein the recombinant cell expresses the polypeptide sequence of the corticotropin-releasing factor 2 receptor agonist disclosed herein, with or without induction. In some embodiments, the method for constructing the recombinant cell includes: transforming the recombinant expression vector into an expression host cell, culturing the cell, and adding an inducer to induce expression (if necessary) to obtain the polypeptide sequence of the corticotropin-releasing factor 2 receptor agonist. Further, the expression host cell is a prokaryotic or eukaryotic cell, such as Escherichia coli, yeast, plant cells, animal cells, etc.

[0108] More specifically, the above-mentioned method for constructing recombinant cells includes the following steps:

[0109] (1) Amplification of the gene encoding the polypeptide sequence of the adrenocorticotropic hormone-releasing factor 2 receptor agonist;

[0110] (2) Construction of recombinant expression vectors;

[0111] (3) The recombinant expression vector is transformed or transfected into the expression host cell;

[0112] (4) Positive clones are obtained by selective screening.

[0113] The disclosed polypeptides can be secreted extracellularly or expressed on the cell surface or inside the cell.

[0114] In some embodiments, preparing the polypeptide sequence of the disclosed corticotropin-releasing factor 2 receptor agonist using recombinant technology includes the following steps:

[0115] (1) The above recombinant cells are cultured and an inducing agent is added to induce (if necessary) the expression of the polypeptide sequence of the adrenocorticotropic hormone-releasing factor 2 receptor agonist disclosed herein, to obtain cell culture;

[0116] (2) Optionally, the polypeptide sequence of the adrenocorticotropic hormone-releasing factor 2 receptor agonist disclosed herein is isolated and purified from the cell culture (e.g., cells, cell culture supernatant).

[0117] Bioactivity

[0118] The disclosed corticotropin-releasing factor 2 receptor agonist has activating activity against receptor CRF2R and is a CRF2R agonist.

[0119] In this disclosure, EC is used 50 The value serves as a numerical measure of agonist potency for a given receptor (i.e., CRF2R). EC50 The value refers to the concentration at which the 50% maximal effect is achieved. In the same assay targeting a specific receptor, EC... 50 Compounds with lower values ​​can be considered to have higher potency to the receptor.

[0120] This disclosure investigated the CRF2R agonist activity of corticotropin-releasing factor 2 receptor agonists. In cellular activity assays for human CRF2R agonism, compounds 1, 2, 3, 4, 7, 8, 9, 10, 11, and 12 exhibited better activity than hUCN2 and the control compound in cells overexpressing the human CRF2R receptor. Meanwhile, in cellular activity assays for human CRF1R agonism, only hUCN1 showed good activity in cells overexpressing the human CRF1R receptor; the other molecules showed no activity.

[0121] Furthermore, pharmacodynamic experiments in SD rats revealed that, compared with the solvent group, different molecules exhibited varying degrees of inhibition on body weight and food intake; among them, compared with the control compounds, compounds 1, 2, 3, 4, 7, 8, 9, and 10 had comparable or enhanced effects on weight loss and inhibition of food intake.

[0122] Furthermore, efficacy studies in DIO mice revealed that compounds 1, 7, 9, 10, 11, and 12 exhibited significant weight-loss effects compared to the solvent group, increasing lean body mass and reducing fat content in obese mice. Compared to Ecnoglutide alone, the combination of compounds 1, 2, and 4 with Ecnoglutide showed stronger weight-loss effects; and compared to Ecnoglutide alone, the combination of compound 1 with Ecnoglutide demonstrated the strongest lean body mass and fat loss / muscle gain effects. Based on these findings, it was further discovered that the combination of compound 1 with Ecnoglutide showed a stronger, dose-dependent weight-loss effect compared to Ecnoglutide alone, with the combination group exhibiting significantly enhanced fat loss and muscle gain.

[0123] Pharmaceutical compositions of corticotropin-releasing factor 2 receptor agonists

[0124] The disclosed corticotropin-releasing factor 2 receptor agonist or polypeptide can be formulated into a pharmaceutical composition wherein the corticotropin-releasing factor 2 receptor agonist or polypeptide is present in a therapeutically effective amount.

[0125] In addition to the active ingredient, an adrenocorticotropic hormone-releasing factor 2 receptor agonist or peptide, the pharmaceutical compositions described herein also contain pharmaceutically acceptable excipients. Those skilled in the art are familiar with pharmaceutically acceptable excipients, such as non-toxic fillers, stabilizers, diluents, carriers, solvents, or other pharmaceutical excipients. Examples include diluents and excipients such as microcrystalline cellulose and mannitol; fillers such as starch and sucrose; binders such as starch, cellulose derivatives, alginate, gelatin, and / or polyvinylpyrrolidone; disintegrants such as calcium carbonate and / or sodium bicarbonate; absorption enhancers such as quaternary ammonium compounds; surfactants such as hexadecyl alcohol; carriers and solvents such as water, physiological saline, kaolin, and soap clay; and lubricants such as talc, calcium / magnesium stearate, and polyethylene glycol.

[0126] The disclosed corticotropin-releasing factor 2 receptor agonist or peptide can be used alone or in combination with any compound beneficial for the prevention and / or treatment of one or more of heart failure, hypertension, obesity, sarcopenia, diabetes, chronic kidney disease and related diseases in a pharmaceutical composition, which is expected to enhance the beneficial therapeutic effects of the disclosed corticotropin-releasing factor 2 receptor agonist. The aforementioned beneficial compounds include, but are not limited to, glucagon-like peptide-1 receptor (GLP-1R) agonists, amylin receptor agonists, glucose-dependent insulinotropic peptide receptor (GIPR) agonists or antagonists, glucagon receptor (GCGR) agonists, or components having simultaneous agonistic or antagonistic activity against two or more of the above receptors (e.g., GLP-1R / GIPR dual agonists, GLP-1R agonist / GIPR antagonist, GLP-1R / GCGR dual agonist, GLP-1R / GIPR / GCGR triple agonist, GLP-1R agonist / GIPR antagonist / GCGR agonist, GLP-1R / amylin receptor agonist, GLP-1R / GIPR / amylin receptor triple agonist, GLP-1R agonist / GIPR antagonist / amylin receptor agonist). A preferred beneficial compound is a glucagon-like peptide-1 receptor agonist, such as ecnoglutide.

[0127] The pharmaceutical compositions disclosed herein may also be used in combination with one or more other drugs, such combination is expected to have a synergistic effect in the prevention and / or treatment of one or more of heart failure, hypertension, obesity, sarcopenia, diabetes, chronic kidney disease and related diseases. The aforementioned one or more drugs include, but are not limited to, glucagon-like peptide-1 receptor (GLP-1R) agonists, amylin receptor agonists, glucose-dependent insulinotropic peptide receptor (GIPR) agonists or antagonists, glucagon receptor (GCGR) agonists, or components having simultaneous agonistic or antagonistic activity against two or more of the above receptors (e.g., GLP-1R / GIPR dual agonists, GLP-1R agonist / GIPR antagonist, GLP-1R / GCGR dual agonist, GLP-1R / GIPR / GCGR triple agonist, GLP-1R agonist / GIPR antagonist / GCGR agonist, GLP-1R / amylin receptor agonist, GLP-1R / GIPR / amylin receptor triple agonist, GLP-1R agonist / GIPR antagonist / amylin receptor agonist). A preferred beneficial compound is a glucagon-like peptide-1 receptor agonist, such as ecnoglutide.

[0128] Medical conditions

[0129] By administering an effective amount of an adrenocorticotropic hormone-releasing factor 2 receptor agonist or peptide or pharmaceutical composition as described herein, it can be used for the prevention and / or treatment of one or more of heart failure, hypertension, obesity, sarcopenia, diabetes, chronic kidney disease and related diseases.

[0130] Example 1: Preparation of polypeptide derivatives

[0131] A solid-phase organic synthesis method was employed, utilizing the Fmoc-protected amino acid strategy, to synthesize, cleave, and purify the peptide derivative, yielding the target product. Taking compound 1 in Table 2 as an example, the preparation process is as follows:

[0132] 1. Solid-phase synthesis:

[0133] Using Rink Amide MBHA resin (100-200 mesh, 1% DVB, CAS 431041-83-7) and the Fmoc / tBu process, amino acids were sequentially condensed and linked from the C-terminus to the N-terminus (from right to left) according to the amino acid sequence of the polypeptide derivative, as shown in Table 1, to finally form a linear polypeptide resin.

[0134] Table 1 List of Synthesis Procedures

[0135] The following amino acids are coupled sequentially:

[0136] A-01Fmoc-Val-OH,A-02Fmoc-Arg(Pbf)-OH,A-03Fmoc-Glu(OtBu)-OH,A-04Fmoc-Leu-OH,A-05Fmoc-Ile-OH,A-06Fmoc-G lu(OtBu)-OH,A-07Fmoc-Ala-OH,A-08Fmoc-Asn(Trt)-OH,A-09Fmoc-Thr(tBu)-OH,A-10Fmoc-Lys(Boc)-OH,A-11Fmoc-A la-OH,A-12Fmoc-Gln(Trt)-OH,A-13Fmoc-Glu(OtBu)-OH,A-14Fmoc-Arg(Pbf)-OH,A-15Fmoc-Ala-OH,A-16Fmoc-Ala-OH ,A-17Fmoc-Arg(Pbf)-OH,A-18Fmoc-Ala-OH,A-19Fmoc-Arg(Pbf)-OH,A-20Fmoc-Ala-OH,A-21Fmoc-Gln(Trt)-OH,A-22F moc-Glu(OtBu)-OH,A-23Fmoc-Leu-OH,A-24Fmoc-Leu-OH,A-25Fmoc-Ile-OH,A-26Fmoc-Gln(Trt)-OH,A-27Fmoc-Leu-OH ,A-28Fmoc-Leu-OH,A-29Fmoc-Gly-OH,A-30Fmoc-Ile-OH,A-31Fmoc-Pro-OH,A-32Fmoc-Val-OH,A-33Fmoc-Asp(OtBu)-O H, A-34Fmoc-Leu-OH, A-35Fmoc-Ser(tBu)-OH, A-36Fmoc-Leu-OH, A-37Fmoc-Val-OH, A-38Fmoc-Ile-OH, A-39Boc-Lys(Fmoc)-OH, A-40Fmoc-AEEA-OH, A-41Fmoc-AEEA-OH, A-42Fmoc-Glu-OtBu, A-43Fmoc-Glu-OtBu, A-44Fmoc-Glu-OtBu, A-45C18 diacid; finally, this peptide resin is formed. The peptide resin is washed, transferred, and dried to constant weight for lysis.

[0137] 2. Peptide resin cleavage:

[0138] Preparation of lysis reagent: Calculate the amount of lysis reagent (TFA:H2O:EDT:TIS = 95:1:2:2 (volume ratio)) based on a ratio of 1g peptide resin to 10ml ± 2ml of lysis reagent. Add the required amounts of lysis reagent (H2O, TFA, EDT, TIS) sequentially to the lysis reaction flask, maintaining the temperature of the lysis reagent at 0–10℃. Add the lysis reagent to the peptide resin with stirring. After the system temperature stabilizes, maintain the temperature at 25–30℃ and stir for 2.5 hours.

[0139] The lysis solution was filtered out and precipitated with 5 times the liquid volume of ice-cold diethyl ether. The precipitate was filtered out and washed 3 times with 3 times the liquid volume of ice-cold diethyl ether. The solution was then dried under reduced pressure at room temperature to obtain a solid crude product.

[0140] 3. Purification and lyophilization:

[0141] The crude product was ground into a fine powder. Purified water was prepared and slowly added to the powder while stirring, along with acetonitrile aqueous solution. After the crude product was completely added and dissolved, the mixture was filtered through a 0.45 μm microporous membrane. Crude product purification was performed using a C-18 column packed with mobile phases: A: 0.1% TFA / H₂O, B: 0.1% TFA / ACN. Separation and purification were carried out at room temperature using a suitable gradient. The target product was collected, analyzed, and classified. The impurity purity requirement was ≥90%. Unqualified target substances were collected and purified again using a suitable gradient. The qualified main peak was freeze-dried under reduced pressure to obtain a powdered purified polypeptide derivative.

[0142] The remaining compounds in Table 2 were synthesized in a similar manner, with the control compound being the UCN2 derivative in WO 2023 / 285347 A1; and Ecnoglutide being the GLP-1 (glucagon-like peptide-1) receptor agonist in WO 2019 / 201328 A1.

[0143] Table 2 UCN2 polypeptide derivatives

[0144] In Table 2, the ε-amino group of lysine is bonded to (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-(γGlu)3-CO-(CH2) via an amide bond. 16-CO2H (English name: 17-{[(22S,27S,32S)-22,27,32-tricarboxy-1,10,19,24,29-pentaoxo-9,18,23,28-tetraza-3,6,12,15-tetraoxadotriacont-32-yl]amino}-17-oxoheptadecane-1-carboxylicacid) The Chinese names are 17-{[(22S,27S,32S)-22,27,32-tricarboxylic acid-1,10,19,24,29-pentaoxylidene-9,18,23,28-tetraaza-3,6,12,15-tetraoxa-tetrazo-dodecane-32-yl]amino}-17-oxylidene-heptadecane-1-carboxylic acid and (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-(γGlu)2-CO-(CH2). 14 -CO2H (English name: 15-{[(22S,27S)-22,27-dicarboxy-1,10,19,24-tetraoxo-3,6,12,15-tetraoxa-9,18,23-triazaheptacos-27-yl]amino}-15-oxopentadecane-1-carboxylic acid; Chinese name: 15-{[(22S,27S)-22,27-dicarboxy-1,10,19,24-tetraoxo-3,6,12,15-tetraoxa-9,18,23-triazaheptacos-27-yl]amino}-15-oxopentadecane-1-carboxylic acid), (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-(γGlu)-CO-(CH2) 14 -CO2H (English name: 16-{[(22S)-22-carboxy-1,10,19-trioxo-9,18-diaza-3,6,12,15-tetraoxadocos-22-yl]amino}-16-oxohexadecanoic acid; Chinese name: 16-{[(22S)-22-carboxyl-1,10,19-trioxo-9,18-diaza-3,6,12,15-tetraoxadocos-22-yl]amino}-16-oxo-ylidene hexadecanoic acid), (2-(2-(2-Aminoethoxy)ethoxy)acetyl)2-(γGlu)3-CO-(CH2) 18-CO2H (English name: 19-{[(22S,27S,32S)-22,27,32-tricarboxy-10,19,24,29-tetraoxo-9,18,23,28-tetraza-3,6,12,15-tetraoxadotriacont-32-yl]amino}-19-oxononadecane-1-carboxylic acid, Chinese name: 19-{[(22S,27S,32S)-22,27,32-tricarboxy-10,19,24,29-tetraoxo-9,18,23,28-tetraaza-3,6,12,15-tetraoxadotriacont-32-yl]amino}-19-oxononadecane-1-carboxylic acid) is linked.

[0145] When using, the polypeptide derivative can be dissolved in the solvent according to the required concentration for the experiment. Unless otherwise specified, PBS solution will be used as the solvent.

[0146] The abbreviations used in the above preparation process have the following meanings: AA: Amino Acid, amino acid; AEEA: 2-(2-(2-aminoethoxy)ethoxy)acetic acid; Boc: t-Butyloxycarbonyl, tert-Butyloxycarbonyl; ACN: Acetonitrile; DMF: N,N-Dimethylformamide, N,N-Dimethylformamide; DIEA: N,N-Diisopropylethylamine, N,N-Diisopropylethylamine; EDT: 1,2-Ethanedithiol, 1,2-Ethylenedithiol; Fmoc: 9-fluorenylmethyloxycarbonyl, 9-fluorenylmethoxycarbonyl; OtBu: tert-Butyl ester; Pbf: 2,2,4,6,7-Pentamethyldihydrobenzofuran-5-sulfonyl chloride, 2,2,4,6,7-pentamethyldihydrobenzofuran-5-sulfonyl Pip: Piperidine, piperidine TBTU: O-(Benzotriazol-l-yl)-N,N,N',N',-tetramethyluronium Tetrafluoroborate, O-benzotriazol-N,N,N',N'-tetramethylurea tetrafluoroborate tBu: tertiary butyl, tert-butyl TFA: Trifluoroacetic acid, trifluoroacetic acid TIS: Triisopropylsilane, triisopropylsilane Trt: Triphenylmethyl, triphenylmethyl γGlu: γ-glutamic acid

[0147] Example 2: Detection of cellular activity of peptide derivatives against human CRF2R agonism

[0148] 1. Detection of cellular activity of peptide derivatives against human CRF2R agonism

[0149] 1.1 The activity of UCN2 derivatives in activating CRF2R was investigated by a human CRF2R-CRE-Luciferase-HEK293 (human CRF2R: NCBI Reference Sequence: NM_001202475.1) reporter gene assay.

[0150] Construction of CRF2R-CRE-Luciferase-HEK293: HEK293 cells (Cell Resource Center, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences, product catalog number 1101HUM-PUMC000010) were transfected with plasmid pGL4.29[luc2P / CRE / Hygro]Vector (Promega, catalog number E8471) containing a multi-copy cAMP response element (CRE) driven luciferase expression cassette and CRF2R_pcDNA3.1(+) (vector source: Invitrogen, V79020) to obtain a stable CRF2R-CRE-Luciferase-HEK293 cell line containing the luciferase expression plasmid.

[0151] In a 96-well plate, add 100 μL of CRF2R-CRE-Luciferase-HEK293 cells (3*10) cultured in DMEM + 10% FBS medium to each well. 5 Cells / mL) were incubated overnight at 37°C in a 5% CO2 incubator, then the old medium was discarded. 50 μL of fresh DMEM + 10% FBS medium was added to each well, followed by 50 μL of different concentrations of the analyte, resulting in 10 concentrations (50 nM, 16.7 nM, 5.56 nM, 1.85 nM, and 0.62 nM) through a 3-fold serial dilution. Incubation continued for 24 hours. 50 μL of Bright-Glo reagent (Promega) was added, and the plate was gently tapped to mix the solution. Chemiluminescence values ​​were read using a microplate reader after 3 minutes. % Activity = Sample Reading / Max Control Reading (mean) * 100% (Max control reading is the average reading of the highest hUCN2 concentration points, totaling 8 points). Graphs were plotted using GraphPad Prism 10 with four parameters, and EC was calculated. 50 Values. The results are shown in Table 3.

[0152] Table 3

[0153] Experimental results showed that compounds 1-6 in this disclosure all exhibited human CRF2R receptor agonist activity; surprisingly, compounds 1, 2, 3, and 4 showed better activity than hUCN2 and the control compound in cell experiments overexpressing human CRF2R receptor.

[0154] 1.2. Activity was detected using the same method as in 1.1. The final results were expressed as hUCN2 EC. 50 To use as a reference and normalize to 1, calculate the relative EC of each molecule. 50 (i.e., the test compound EC) 50 / hUCN2 EC 50 As shown in Table 4.

[0155] Table 4: Relative to EC 50

[0156] As shown in Table 4, similar to compounds 1, 2, 3, and 4, compounds 7-12 also exhibited better activity than hUCN2 and the control compounds in experiments on cells overexpressing the human CRF2R receptor.

[0157] 2. Detection of cellular activity of peptide derivatives against human CRF1R agonism

[0158] The activity of UCN2 derivatives or hUCN1 (DNPSLSIDLTFHLLRTLLE LARTQSQRERAEQNRIIFDSV-NH2, SEQ ID NO:6) in activating CRF1R was investigated using a human CRF1R-CRE-Luciferase-HEK293 (human CRF1R: NCBI Reference Sequence: NM_001145146.2) reporter gene assay.

[0159] Construction of CRF1R-CRE-Luciferase-HEK293: HEK293 cells (Cell Resource Center, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences, product catalog number 1101HUM-PUMC000010) were transfected with plasmid pGL4.29[luc2P / CRE / Hygro]Vector (Promega, catalog number E8471) containing a multi-copy cAMP response element (CRE) driven luciferase expression cassette and CRF1R_pcDNA3.1(+) (vector source: Invitrogen, V79020) to obtain a stable CRF1R-CRE-Luciferase-HEK293 cell line containing the luciferase expression plasmid.

[0160] In a 96-well plate, add 100 μL of CRF1R-CRE-Luciferase-HEK293 (3 x 10⁻⁶ cells / well) cultured in DMEM + 10% FBS medium to each well. 5 Cells / mL) were collected and incubated overnight at 37°C in a 5% CO2 incubator, then the old culture medium was discarded. 50 μL of fresh DMEM + 10% FBS medium was added to each well, followed by 50 μL of different concentrations of the analyte, resulting in 10 concentrations (1000 nM, 333.3 nM, 111.1 nM, 37.04 nM, and 12.35 nM) through a 3-fold serial dilution. Incubation continued for 24 hours. 50 μL of Bright-Glo reagent (Promega) was added, and the plate was gently tapped to mix the solution. The chemiluminescence value was read using a microplate reader after 3 minutes. Activity calculation formula: % activity = (sample read / mean Max control read) * 100% (The mean Max control read is the mean of the highest concentration hUCN1 readings, for a total of 8 points).

[0161] The activity of the polypeptide derivative on CRF1R cells is shown in Figure 1.

[0162] Figure 1 shows that only hUCN1 exhibited good activity in the experiment of overexpressing human CRF1R receptor cells, while other molecules were inactive.

[0163] Example 3: Pharmacodynamic Experiment in SD Rats

[0164] 1.1. SPF-grade male SD rats (8-10 weeks old, approximately 250g) were used in this study after quarantine. The animal housing conditions were: room temperature 20℃~23℃, relative humidity 40%~50%. During the quarantine period and the experiment, the animals were fed Co60 rat breeding feed, and purified water was provided in bottles for free access. The animals were divided into 8 groups (n=5) based on average body weight, named A1-A8. The compounds and dosages administered to each group are shown in Table 5. The solvent was PBS, pH 7.4. Subcutaneous injection was used, with a single dose of 5mL / kg. The experiment lasted 4 days, with each day of administration recorded as Day 1 / Time 0 (T0). Day 0 recorded the initial body weight and the addition of the initial feed. From Day 1 to Day 3, the animal's body weight and remaining food were recorded daily, and the relative body weight change rate and food intake were calculated. Any abnormalities were recorded and reported. The rats remained in good condition throughout the experiment, and no abnormalities were observed. Weight change rate = (BW) Tn -BW T0 ) / BW T0 *100%, BW represents body weight, T0 and Tn represent administration and n hours after administration, respectively.

[0165] The weight-loss and food intake inhibition effects of different molecules are shown in Figures 2 and 3, respectively. The points in the figures are represented using the mean ± standard error (SEM) method. Food intake is the average amount of food consumed per animal in each group before a certain time point after drug administration.

[0166] Table 5. Active ingredients and dosages of test samples in single-dose screening experiments on SD rats.

[0167] As shown in Figures 2 and 3, compared with the A1 solvent group, different molecules exhibited varying degrees of inhibition on body weight and food intake. Among them, compounds 1, 2, 3, and 4 showed comparable or enhanced effects on weight loss and inhibition of food intake compared to the control compounds.

[0168] 1.2 The efficacy of the compound after a single dose in SD rats was tested in a manner similar to that in 1.1, as follows:

[0169] SPF-grade male SD rats (6-8 weeks old, 220-240g) were used in this study after quarantine. The animals were housed at room temperature of 20℃-26℃ and relative humidity of 40%-70%. During quarantine and the experiment, they were fed Co60 rat breeding feed and provided purified water via bottles, allowing free access to water. The animals were divided into 6 groups (n=5) based on average body weight, named A1'-A6'. The compounds and dosages for each treatment group are shown in Table 6. Subcutaneous injection was used, with a single dose administered over 4 days. The day of administration was recorded as Day1 / Time0 (T0). Day0 recorded the initial body weight and the addition of the initial feed. From Day1 to Day3, the animal's body weight and remaining food were recorded daily, and the relative body weight change rate and food intake were calculated. Any abnormalities were recorded and reported. The rats remained in good condition throughout the experiment, and no abnormalities were observed. Body weight change rate = (BW...) / (T0 / T0 ... Tn -BW T0 ) / BW T0 *100%, BW represents body weight, T0 and Tn represent drug administration and n hours after administration, respectively. Points in the figure are represented using the mean ± standard error (SEM) method. Food intake is the average amount of food consumed per animal in each group before a certain time point after drug administration. The efficacy comparison of different drug molecules is shown in Figure 4.

[0170] Table 6. Active ingredients and dosages of test samples in SD rat experiments.

[0171] As can be seen from the results in Figure 4, in the single-dose rat experiment, compared with the solvent group, all compounds of the present invention had significant effects on weight reduction and food intake inhibition. Among them, compound 1 had better effects on weight reduction and food intake inhibition than other molecules.

[0172] Example 4: Pharmacodynamic experiments of compounds 1, 2, and 4 in combination with Ecnoglutide injection in DIO mice.

[0173] SPF-grade male DIO mice (C57BL / 6J mice, 17-18 weeks old, 40-60g, fed a high-fat diet for 12 weeks starting at 5 weeks of age) were used for this study. The animal housing conditions were: room temperature 20℃~23℃, relative humidity 40%~50%. During the DIO model establishment and the experiment, the animals were fed a high-fat diet (D12492), and purified water was provided via water bottles for free access. The animals were randomly divided into 5 groups (n=5) based on body weight, named B1-B5. The compounds and dosages administered to each group are shown in Table 7. Solvent 1 (8mM phosphate buffer solution (PB) + 15mg / mL propylene glycol + 5.5mg / mL phenol, pH 7.4) and solvent 2 (phosphate buffer saline (PBS), pH 7.4) were the solvents for Ecnoglutide and the compound of this application, respectively. The drugs were administered subcutaneously at a volume of 5 mL / kg, once daily for 21 consecutive days. The experiment lasted 23 days, with the day of administration designated as Day 1 (D1). Day 0 recorded the initial body weight of the animals. From Day 1 to Day 22, the body weight of the experimental animals was recorded daily. On Day 22, at the experimental endpoint, NMR was used to detect body fat and lean body mass in the mice, and the rate of change in body weight, lean body mass (lean body weight / body weight), and fat content (fat / body weight) were calculated. Any abnormalities were reported. The DIO mice remained in good condition throughout the experiment, and no abnormalities were observed. Rate of change in body weight = (BW...) / (BW...) Dn -BW D1 ) / BW D1 *100%, BW represents body weight, D1 and Dn represent the time of administration and day N after administration, respectively. Statistical analysis was performed using a one-way ANOVA method, where ns represents no statistical difference, * represents p<0.05, ** represents p<0.01, *** represents p<0.001, and **** represents p<0.0001.

[0174] Figures 5 and 6 show the weight loss and body composition results of DIO mice, respectively. All points in the figures are represented using the mean ± standard error (SEM) method.

[0175] Table 7. Active ingredients and dosages of test samples in single-dose screening experiments on DIO mice.

[0176] As shown in Figure 5, the combination of compounds 1, 2, and 4 with Ecnoglutide had a stronger weight loss effect compared with Ecnoglutide alone. As shown in Figure 6, the combination of compound 1 with Ecnoglutide had the highest lean body mass and the lowest fat mass compared with Ecnoglutide alone.

[0177] Example 5: Pharmacodynamic study of compound 1 in DIO mice - dose-dependent effect

[0178] SPF-grade male DIO mice (C57BL / 6J mice, 25-26 weeks old, 40-60g, fed a high-fat diet for 21 weeks starting at 5 weeks of age) were used for this study. The animal housing conditions were: room temperature 20℃~23℃, relative humidity 40%~50%. During the DIO model establishment and the experiment, the animals were fed a high-fat diet (D12492), and purified water was provided via water bottles for free access. The animals were randomly divided into 5 groups (n=5) based on body weight, named C1-C5. The compounds and dosages administered to each group are shown in Table 8. Solvent 1 (8mM phosphate buffer solution (PB) + 15mg / mL propylene glycol + 5.5mg / mL phenol, pH 7.4) and solvent 2 (phosphate buffer saline (PBS), pH 7.4) were the solvents for Ecnoglutide and the compound of this application, respectively. The drugs were administered subcutaneously at a volume of 5 mL / kg, once daily for 21 consecutive days. The experiment lasted 23 days, with the day of administration designated as Day 1 (D1). Day 0 recorded the initial body weight of the animals, and Days 1-22 recorded the body weight daily. On Day 22, at the experimental endpoint, NMR was used to detect body fat and lean body mass in the mice. The rate of change in body weight, lean body mass (lean body weight / body weight), and fat content (fat / body weight) were calculated, and any abnormalities were reported. The DIO mice remained in good condition throughout the experiment, and no abnormalities were observed. The rate of change in body weight = (BW...) / (BW...) Dn -BW D1 ) / BW D1*100%, BW represents body weight, D1 and Dn represent the time of administration and day N after administration, respectively. Statistical analysis was performed using a one-way ANOVA method, where ns represents no statistical difference, * represents p<0.05, ** represents p<0.01, *** represents p<0.001, and **** represents p<0.0001.

[0179] Figures 7 and 8 show the weight loss and body composition results of DIO mice, respectively. All points in the figures are represented using the mean ± standard error (SEM) method.

[0180] Table 8. Dose-dependent efficacy study in DIO mice: active ingredient and dosage of the test product.

[0181] As shown in Figure 7, the combination of compound 1 and Ecnoglutide had a stronger weight loss effect than Ecnoglutide alone, and this effect was dose-dependent. As shown in Figure 8, the combination therapy group had a significantly enhanced fat loss and muscle gain effect compared to Ecnoglutide alone.

[0182] Example 6: Pharmacodynamic experiment of the compound after multiple administrations in DIO mice

[0183] Experimental methods:

[0184] SPF-grade male DIO mice (17-18 weeks old, 40-60g, C57BL / 6J mice fed a high-fat diet from 5 weeks of age) were used in this study. The experimental animals were housed at room temperature of 20℃–26℃ and relative humidity of 30%–70%. During the establishment of the DIO model and throughout the experiment, the animals were fed a high-fat diet (D12492) and provided purified water via water bottles, allowing free access to water. The animals were randomly divided into 9 groups (n=5) based on body weight, named E1-E5 and F1-F4. The active ingredients and dosages of the administered compounds for each group are shown in Tables 9 and 10. The drugs were administered subcutaneously once daily for 14 consecutive days. The experiment lasted 15 days, with the day of administration designated as Day 1 (D1). Day 0 recorded the initial body weight of the animals, and body weight was recorded daily from Day 1 to Day 15. On Day 15, at the end of the experiment, NMR was used to detect the body fat and lean body mass of the mice. The rate of change in body weight, body fat percentage, and any abnormalities were calculated and reported. The DIO mice remained in good condition throughout the experiment, and no abnormalities were observed. Rate of change in body weight = (BW) Dn -BW D1 ) / BW D1*100%, BW represents body weight, D1 and Dn represent the time of administration and day N after administration, respectively. Lean body mass = (Lean mass / BW)100%, fat mass = (Fat mass / BW). Points in the figure are expressed using the mean ± standard error (SEM) method. Cumulative food intake is the average total food intake per animal in each group before a certain time point after administration. Statistical analysis was performed using the One-Way ANOVA method, where ns represents no statistical difference, * represents p<0.05, ** represents p<0.01, *** represents p<0.001, and **** represents p<0.0001. The changes in body weight reduction, food intake, and body fat content in DIO mice in groups E1-E5 are shown in Figures 9 and 10, respectively. The changes in body weight reduction and food suppression in groups F1-F4 are shown in Figure 11, and the effect on body fat content in DIO model mice is shown in Figure 12.

[0185] Table 9. Active ingredients and dosages of test samples in DIO mice.

[0186] Table 10 Active ingredients and dosages of DIO mouse test samples

[0187] As can be seen from the results in Figures 9 and 11, in the multiple-dose administration experiment in obese mice, the invented compounds showed a significant weight loss effect compared to the solvent group. More importantly, the invented compounds significantly increased the lean body mass of obese mice while reducing the fat content (Figures 10 and 12).

[0188] Example 7: Pharmacodynamic experiment of compound 1 in maintaining weight loss after discontinuation of Ecnoglutide

[0189] Experimental methods:

[0190] SPF-grade male DIO mice (18-19 weeks old, 40-60g; C57BL / 6J mice were fed a high-fat diet from 5 weeks of age) were used in this study. The experimental animals were housed at room temperature of 20℃–26℃ and relative humidity of 30%–70%. During the experiment, they were fed a high-fat diet (D12492) and provided purified water via water bottles for free access. After quarantine, the animals were divided into two groups, named G1 (n=5) and G2 (n=15). The active ingredients and dosages of the administered compounds in the two groups are shown in Table 11. The drugs were administered subcutaneously once daily for 14 consecutive days. The day of administration was designated Day 1 (D1). On Day 14, the G2 group was further subdivided according to body weight and body fat percentage, named G3-G5 (n=5). The active ingredients and dosages of the administered compounds are shown in Table 12. The mice were administered subcutaneously once daily for 14 days (Day 15-Day 28). From Day 1 to Day 28, the mice's weight was recorded daily, and the rate of weight change was calculated. On Day 14 and Day 29, MRI scans were performed to measure lean body mass and fat content, and any abnormalities were recorded and reported. The mice remained healthy throughout the experiment, and no abnormalities were observed. Rate of weight change = (BW) Dn -BW D1 ) / BW D1 *100%, Lean body mass change rate = (Lean D29 -Lean D14 ) / LeanD 14 *100%, Fat Change Rate = (Fat D29 -Fat D14 ) / Fat D14 *100%, Lean body mass percentage = (Lean fat percentage) D15 / Fat D15 (BW represents body weight, D1 and Dn represent the time of drug administration and day N after drug administration, respectively. Points in the figure are expressed as mean ± standard error (SEM). Statistical analysis was performed using one-way ANOVA, where ns represents no statistical difference, * represents p < 0.05, ** represents p < 0.01, *** represents p < 0.001, and **** represents p < 0.0001. The effect of weight reduction in the DIO model mice is shown in Figure 13, and the effect of changes in body fat content in the DIO model mice is shown in Figures 14 and 15.)

[0191] Table 11 Active ingredients and dosages of DIO mouse test samples

[0192] Table 12 Active ingredients and dosages of DIO mouse test samples

[0193] Conclusion: As shown in Figure 13, after discontinuing Ecnoglutide, compared with the solvent group, the low- and high-dose compounds 1 controlled the weight rebound and maintained the weight at a lower level better. More importantly, as shown in Figures 14 and 15, after discontinuing Ecnoglutide, both the low- and high-dose compounds 1 increased lean body mass and continuously reduced fat mass, and the lean body fat ratio increased dose-dependently.

Claims

1. A corticotropin-releasing factor 2 receptor agonist comprising the amino acid sequence of Formula 1: KIVLSLDVPIGLLQILLEQX 20 X 21 X 22 X 23 X 24 X 25 X 26 X 27 QAKTNAEILERV (Formula 1) in, X 20 selected from A or E, X 21 selected from R or K, X 22 selected from A or Q, X 23 selected from R or E, X 24 selected from A or K, X 25 selected from A or E, X 26 selected from R or K, and / or X 27 selected from E or Q; Optionally, the carboxyl terminus of the amino acid sequence of Formula 1 is modified with NH2.

2. The corticotropin-releasing factor 2 receptor agonist of claim 1, wherein: In the amino acid sequence of Formula 1, X 20 is A, X 21 is R, X 22 is A, X 23 is R, X 24 is A, X 25 is A, X 26 is R, and X 27 is E.

3. The corticotropin-releasing factor 2 receptor agonist of claim 1, wherein: In the amino acid sequence of Formula 1, X 20 is E, X 21 is K, X 22 is Q, X 23 is E, X 24 is K, X 25 is E, X 26 is K, and X 27 is Q.

4. A corticotropin-releasing factor 2 receptor agonist according to any one of claims 1 to 3, characterized in that: In the amino acid sequence of Formula 1, at least one amino acid residue is attached to a fatty acid side chain, preferably an amino acid K residue is attached to a fatty acid side chain, and more preferably an amino acid K residue at position 1 and / or position 30 is attached to a fatty acid side chain.

5. A corticotropin-releasing factor 2 receptor agonist according to any one of claims 1 to 4, characterized in that: The ε-amino group of the amino acid K residue in the amino acid sequence of Formula 1 is linked to the side chain of the fatty acid.

6. A corticotropin-releasing factor 2 receptor agonist according to any one of claims 1 to 5, characterized in that: The fatty acid side chain is selected from... One or more of the following, where x is any integer from 4 to 38; Preferably, the fatty acid side chain is selected from: HOOC(CH2) 14 CO-, HOOC(CH2) 15 CO-, HOOC(CH2) 16 CO-, HOOC(CH2) 17 CO-, HOOC(CH2) 18 CO-, HOOC(CH2) 19 CO-, HOOC(CH2) 20 CO-, HOOC(CH2) 21 CO-, HOOC(CH2) 22 one or more of CO-, HOOC(CH2) 7. A corticotropin-releasing factor 2 receptor agonist according to any one of claims 1 to 6, characterized in that: The fatty acid side chain is linked to amino acid residues of the amino acid sequence of Formula 1 via a linker; Preferably, the linker is selected from One or more of the following, where m is 0, 1, 2 or 3; n is 1 or 2; p is any integer from 1 to 5; Preferably, the linker is:

8. A corticotropin-releasing factor 2 receptor agonist according to any one of claims 1 to 7, characterized in that: The agonist is selected from compounds 1 to 12.

9. A polypeptide having an amino acid sequence as shown in Formula 1 of any one of claims 1-3; preferably, the amino acid sequence of the polypeptide is shown in SEQ ID NO:2 or 3.

10. A nucleic acid molecule encoding the polypeptide of claim 9.

11. A recombinant vector comprising the nucleic acid molecule of claim 10.

12. A recombinant cell comprising the nucleic acid molecule of claim 10 or the recombinant vector of claim 11; preferably, the recombinant cell is capable of expressing and optionally secreting the polypeptide of claim 9.

13. A method for preparing the corticotropin-releasing factor 2 receptor agonist according to any one of claims 1-8 or the polypeptide according to claim 9; The method includes the step of preparing the adrenocorticotropic hormone-releasing factor 2 receptor agonist or polypeptide using chemical and / or biological methods; Preferably, the chemical method includes liquid-phase or solid-phase peptide synthesis; the biological method includes molecular biology methods and / or cell biology methods.

14. A pharmaceutical composition comprising the adrenocorticotropic hormone-releasing factor 2 receptor agonist of any one of claims 1-8 or the polypeptide of claim 9, and pharmaceutically acceptable excipients.

15. The pharmaceutical composition of claim 14, wherein: The pharmaceutical composition further comprises a glucagon-like peptide-1 receptor (GLP-1R) agonist, an amylin receptor agonist, a glucose-dependent insulinotropic peptide receptor (GIPR) agonist or antagonist, a glucagon receptor (GCGR) agonist, or a component having agonist or antagonist activity against two or more of the above receptors simultaneously (e.g., GLP-1R / GIPR dual agonist, GLP-1R agonist / GIPR antagonist, GLP-1R / GCGR dual agonist, GLP-1R / GIPR / GCGR triple agonist, GLP-1R agonist / GIPR antagonist / GCGR agonist, GLP-1R / amylin receptor agonist, GLP-1R / GIPR / amylin receptor triple agonist, GLP-1R agonist / GIPR antagonist / amylin receptor agonist); preferably, the pharmaceutical composition further comprises a glucagon-like peptide-1 receptor agonist, such as Ecnoglutide.

16. The use of the corticotropin-releasing factor 2 receptor agonist of any one of claims 1-8, the polypeptide of claim 9, or the pharmaceutical composition of claim 14 or 15 in the preparation of a medicament for the prevention and / or treatment of corticotropin-releasing factor 2 receptor-mediated diseases.

17. Use according to claim 16, characterized in that: The disease mentioned is one or more of the following: heart failure, hypertension, obesity, sarcopenia, diabetes, chronic kidney disease, and related diseases.

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