A disulfide cyclic peptide compound, and a preparation method and application thereof

Abstract

The application belongs to the technical field of medicine, and particularly relates to a disulfide ring peptide compound of a melanocortin receptor agonist based on a polypeptide small molecule, the disulfide ring peptide compound has the following structure: R2-R1-c(Cys-D-Ala-His-D-Phe-Arg-Trp-Cys)-R3-R4; wherein R1 is selected from Arg; R2 is selected from an acetyl group or a palmitoyl group; R3 is selected from Ile, Pro or is absent; and R4 is selected from an amino group; when R3 is absent, R2 is a palmitoyl group. The disulfide ring peptide compound provided in the application has good selectivity for melanocortin receptors (MCR1, MCR3, MCR4, MCR5).

Description

[0001] This invention is a divisional application of Chinese Patent Application No. 2024118964772, filed on December 20, 2024, entitled "A disulfide cyclic peptide compound and its preparation method and application," the entire contents of which are incorporated herein by reference. Technical Field

[0002] This invention belongs to the field of pharmaceutical technology, and in particular relates to a disulfide cyclic peptide compound, its preparation method, and its application. Background Technology

[0003] The melanocortin system includes melanocortin and its receptor (MCR), neuropeptide Y (NPY), and endogenous melanocortin antagonists agouti and agouti-related protein (AgRP). Melanocortin hormones are a large class of polypeptides regulating different functions, including α-, β-, and γ-melanocyte stimulating hormones (MSH) and adrenocorticotropin (ACTH). They all originate from tissue-specific post-translational processing of pro-opiomelanocortin (POMC). The melanocortin receptor system includes five MCRs (MC1R~MC5R) and two receptor helper proteins (MRAPs). Except for MC2R, which specifically binds to ACTH, MC1R, MC3R, MC4R, and MC5R can bind to all melanocyte stimulating hormones, but their affinity varies among different receptors.

[0004] The function of MCR receptors is related to their specific expression in tissues. MC2R, primarily located in the adrenal cortex, is a key component of the hypothalamic-pituitary-adrenal axis, stimulating the biosynthesis of glucocorticoids. Melanocortin receptors, MC3R and MC4R, are involved in energy metabolism regulation. MC4R, due to its loss of function, manifests as an obesity-like phenotype in both mice and humans, and is considered a key factor in regulating individual energy metabolism. In addition to MC4R, MC3R is widely distributed in the brain, primarily expressed in the hypothalamus, mainly the arcuate nucleus and ventromedial hypothalamus. These structures are involved in regulating energy homeostasis, metabolism, and appetite. Many studies have shown that MC3R and MC4R can function independently, playing complementary rather than redundant roles in controlling energy balance. MC5R is widely expressed in peripheral organs and tissues. MC5R appears to play a crucial role in immune and inflammatory responses and is essential for temperature control and exocrine function.

[0005] Several melanocortin peptide analogs have been developed as potential drugs for treating skin diseases, obesity, anorexia, and type 2 diabetes. Setmelanotide is a cyclic high-affinity peptide with a G protein signaling spectrum biased towards Gq / 11 (phospholipase C activation) compared to natural α-MSH, exhibiting 20-fold selectivity for the MC4R receptor subtype. As a targeted therapy, setmelanotide restores the function of the damaged MC4R pathway, re-establishing energy expenditure and appetite control in patients with rare genetic obesity disorders, reducing hunger and weight loss. However, setmelanotide also has certain side effects, including injection site reactions, skin pigmentation (darker patches than surrounding skin), headaches, and gastrointestinal side effects (such as nausea, diarrhea, and abdominal pain). Reproductive system adverse reactions have also been reported during treatment. Depression and suicidal ideation have also been reported during setmelanotide use.

[0006] Therefore, it is still necessary to develop a ligand for the melanocortin receptor (MC1R-MC5R) that has better selectivity, stronger stability, and fewer side effects. Summary of the Invention

[0007] The purpose of this invention is to provide a disulfide cyclic peptide compound, its preparation method, and its application.

[0008] To achieve the above-mentioned objectives, the technical solution of this invention is as follows:

[0009] In a first aspect of the present invention, a disulfide cyclic peptide compound is provided having the following structure: R2-R1-c(Cys-D-Ala-His-D-Phe-Arg-Trp-Cys)-R3-R4; wherein R2 and R4 are each independently selected from amino, acetyl, or palmitoyl groups; R1 and R3 are each independently selected from Ile, Arg, D-Phe, Pro, or omitted, R1 and R3 are not omitted simultaneously, and when R1 or R3 is omitted, at least one of R2 and R4 is a palmitoyl group.

[0010] In one aspect of the invention, the disulfide cyclic peptide compound has the following structural formula:

[0011]

[0012] The definitions of R1, R2, R3, and R4 are as described above.

[0013] In one aspect of the invention, R3 is selected from Ile, D-Phe, Pro or missing, and R1 is Arg.

[0014] In one aspect of the invention, R2 is selected from acetyl or palmitoyl groups, and R4 is an amino group.

[0015] In one aspect of the invention, when R3 is missing, R4 is a palmitoyl group.

[0016] In one aspect of the invention, the disulfide cyclic peptide compound is selected from at least one of the following compounds:

[0017]

[0018]

[0019] In one aspect of the present invention, a disulfide cyclic peptide compound is provided, wherein the disulfide cyclic peptide compound is compound WP302-1.

[0020] Specifically, compound WP302-1 has the following structure:

[0021]

[0022] Specifically, compound WP302-1 is Ac-Arg-c(Cys-D-Ala-His-D-Phe-Arg-Trp-Cys)-D-Phe-NH2.

[0023] Specifically, compound WP302-1 has the amino acid sequence shown in SEQ ID NO.1.

[0024] Specifically, the molecular formula of compound WP302-1 is C58 H 77 O 10 N 19 S2.

[0025] In one aspect of the present invention, a disulfide cyclic peptide compound is provided, wherein the disulfide cyclic peptide compound is compound WP302-2.

[0026] Specifically, compound WP302-2 has the following structure:

[0027]

[0028] Specifically, compound WP302-2 is Ac-Arg-c(Cys-D-Ala-His-D-Phe-Arg-Trp-Cys)-Pro-NH2.

[0029] Specifically, compound WP302-2 has the amino acid sequence shown in SEQ ID NO.2.

[0030] Specifically, the molecular formula of compound WP302-2 is C 54 H 75 O 10 N 19 S2.

[0031] In one aspect of the present invention, a disulfide cyclic peptide compound is provided, said disulfide cyclic peptide compound being compound WP302-3.

[0032] Specifically, compound WP302-3 has the following structure:

[0033]

[0034] Specifically, compound WP302-3 is Ac-Arg-c(Cys-D-Ala-His-D-Phe-Arg-Trp-Cys)-Ile-NH2.

[0035] Specifically, compound WP302-3 has the amino acid sequence shown in SEQ ID NO.3.

[0036] Specifically, the molecular formula of compound WP302-3 is C 55 H 79 O 10 N 19 S2.

[0037] In one aspect of the present invention, a disulfide cyclic peptide compound is provided, wherein the disulfide cyclic peptide compound is compound WP302-4.

[0038]

[0039] Specifically, compound WP302-4 is Palm-Arg-c(Cys-D-Ala-His-D-Phe-Arg-Trp-Cys)-NH2.

[0040] Specifically, compound WP302-4 has the amino acid sequence shown in SEQ ID NO.4.

[0041] Specifically, the molecular formula of compound WP302-4 is C 63 H 96 N 18 O9S2.

[0042] In one aspect of the present invention, a disulfide cyclic peptide compound is provided, wherein the disulfide cyclic peptide compound is compound WP302-5.

[0043] Specifically, compound WP302-5 has the following structure:

[0044]

[0045] Specifically, compound WP302-5 is Palm-Arg-c(Cys-D-Ala-His-D-Phe-Arg-Trp-Cys)-D-Phe-NH2.

[0046] Specifically, compound WP302-5 has the amino acid sequence shown in SEQ ID NO.5.

[0047] Specifically, the molecular formula of compound WP302-5 is C 72 H 105 N 19 O 10 S2.

[0048] In one aspect of the present invention, a disulfide cyclic peptide compound is provided, wherein the disulfide cyclic peptide compound is compound WP302-6.

[0049] Specifically, compound WP302-6 has the following structure:

[0050]

[0051] Specifically, compound WP302-6 is Palm-Arg-c(Cys-D-Ala-His-D-Phe-Arg-Trp-Cys)-Pro-NH2.

[0052] Specifically, compound WP302-6 has the amino acid sequence shown in SEQ ID NO.6.

[0053] Specifically, the molecular formula of compound WP302-6 is C 68 H 103 N 19 O 10 S2.

[0054] In one aspect of the present invention, a disulfide cyclic peptide compound is provided, wherein the disulfide cyclic peptide compound is compound WP302-7.

[0055] Specifically, compound WP302-7 has the following structure:

[0056]

[0057] Specifically, compound WP302-7 is Palm-Arg-c(Cys-D-Ala-His-D-Phe-Arg-Trp-Cys)-Ile-NH2.

[0058] Specifically, compound WP302-7 has the amino acid sequence shown in SEQ ID NO.7.

[0059] Specifically, the molecular formula of compound WP302-7 is C 69 H 107 N 19 O 10 S2.

[0060] In a second aspect of the present invention, a method for preparing the aforementioned disulfide cyclic peptide compound is provided, the method comprising the following steps:

[0061] Step 1: Select a resin and remove the Fmoc protecting group to obtain a resin with the Fmoc protecting group removed;

[0062] Step 2: Weigh Fmoc-R3-OH or Fmoc-Cys(Trt)-OH and PyBop, and perform a coupling reaction on the resin with the Fmoc protecting group removed to obtain the coupling resin.

[0063] Step 3: According to the polypeptide sequence, for the coupling resin, couple sequentially from the C-terminus to the N-terminus to obtain a linear peptide resin;

[0064] Step 4: The protecting group is removed by reacting the linear peptide resin with the lysis buffer to obtain the polypeptide;

[0065] Step 5: Dissolve the polypeptide in water, adjust the pH and add H2O2 to react and obtain the disulfide cyclic peptide compound.

[0066] In a third aspect of the present invention, the use of the aforementioned disulfide cyclic peptide compound or the disulfide cyclic peptide compound prepared by the aforementioned method in the preparation of a pharmaceutical is provided.

[0067] In one aspect of the invention, the medicament includes a medicament for treating or preventing acute or chronic inflammatory diseases, autoimmune diseases, transplant rejection, metabolic diseases with weight gain, metabolic diseases with weight loss, diabetes, diabetic complications, cancer, cancerous hyperplasia, reproductive system diseases, peripheral or central nervous system diseases, cardiovascular diseases, or respiratory system diseases.

[0068] Specifically, the acute or chronic inflammatory diseases include, but are not limited to, systemic inflammation, inflammatory bowel disease, encephalitis, sepsis, and septic shock.

[0069] Specifically, the autoimmune diseases include, but are not limited to, systemic lupus erythematosus, rheumatoid arthritis, gouty arthritis, psoriatic arthritis, axial spondyloarthritis, myasthenia gravis, multiple sclerosis, psoriasis, pemphigus, vitiligo, somnambulism, neuromyelitis optica, hyperthyroidism, hypothyroidism, autoimmune gastritis, autoimmune hepatitis, primary cholangitis, Crohn's disease, ulcerative colitis, celiac disease, lupus nephritis, pulmonary hemorrhage-nephritis syndrome, autoimmune oophoritis, autoimmune orchitis, polymyositis, vasculitis, or diffuse connective tissue diseases such as Sjögren's syndrome.

[0070] Specifically, the organs involved in transplant rejection include, but are not limited to, the kidney, heart, liver, pancreas and islets of Langerhans, parathyroid glands, heart and lungs, bone marrow, and cornea.

[0071] Specifically, the metabolic diseases associated with weight gain include, but are not limited to, obesity, eating disorders, and Pu-Wei syndrome.

[0072] Specifically, the metabolic diseases associated with weight loss include, but are not limited to, anorexia or bulimia.

[0073] Specifically, the diabetic complications include, but are not limited to, diabetic nephropathy, diabetic retinopathy, diabetic cataract, diabetic foot, diabetic cardiovascular complications, diabetic cerebrovascular disease, and diabetic neuropathy.

[0074] Specifically, the cancers mentioned include, but are not limited to, dermatomas, neuroblastomas, rectal cancer, colon cancer, familiar adenomatous polyposis and hereditary nonpolyposis colorectal cancer, esophageal cancer, lip cancer, laryngeal cancer, hypopharyngeal cancer, tongue cancer, salivary gland cancer, gastric cancer, adenocarcinoma, medullary thyroid carcinoma, papillary thyroid carcinoma, kidney cancer, renal parenchymal carcinoma, ovarian cancer, cervical cancer, uterine corpus cancer, endometrial cancer, choriocarcinoma, pancreatic cancer, prostate cancer, bladder cancer, testicular cancer, breast cancer, and urinary tract cancer. Melanoma, brain tumor, lymphoma, head and neck cancer, acute lymphoblastic leukemia, chronic lymphoblastic leukemia, acute myeloid leukemia, chronic myeloid leukemia, hepatocellular carcinoma, gallbladder cancer, bronchial carcinoma, small cell lung cancer, non-small cell lung cancer, multiple myeloma, basal sarcoma, teratoma, retinoblastoma, choroidal melanoma, seminoma, rhabdomyosarcoma, osteosarcoma, chondrosarcoma, myoma, liposarcoma, fibrosarcoma, Ewing sarcoma, and plasmacytoma.

[0075] Specifically, the cancerous hyperplasia includes, but is not limited to, squamous epithelial dysplasia and glandular epithelial dysplasia.

[0076] Specifically, the reproductive system diseases include, but are not limited to, abnormal urination, pyuria, abnormal urethral discharge, pain, lumps, sexual dysfunction, male infertility, female infertility, polycystic ovary syndrome, menstrual disorders, dysmenorrhea, abnormal pregnancy, and uterine lesions.

[0077] Specifically, the peripheral or central nervous system diseases include, but are not limited to, depression, bipolar disorder or manic depression, acute and chronic anxiety states, schizophrenia, Alzheimer's disease, vascular dementia, Parkinson's disease, acute and chronic multiple sclerosis or acute and chronic pain, and brain injury caused by stroke, hypoxia, or craniocerebral trauma.

[0078] Specifically, the cardiovascular diseases include, but are not limited to, coronary heart disease, stroke, ischemic stroke, cerebral hemorrhage, subarachnoid hemorrhage, heart failure, hypertensive heart disease, rheumatic heart disease, cardiomyopathy, abnormal heart rhythm, congenital heart disease, valvular heart disease, carditis, aortic aneurysm, peripheral artery disease, thromboembolic disease, and venous thrombosis.

[0079] Specifically, the respiratory diseases include, but are not limited to, upper respiratory tract infections, acute bronchitis, acute pharyngitis, pneumonia, chronic bronchitis, chronic obstructive pulmonary disease, pulmonary tuberculosis, lung tumors, bronchiectasis, lung abscess, pulmonary interstitial fibrosis, pulmonary embolism, acute respiratory distress syndrome, and cor pulmonale.

[0080] In one aspect of the invention, the application is achieved by the binding of the disulfide cyclic peptide compound to the melanocortin receptor.

[0081] In one aspect of the invention, the melanocortin receptor includes at least one of MC1R, MC3R, MC4R, and MC5R.

[0082] In a fourth aspect of the present invention, a pharmaceutical composition is provided comprising the aforementioned disulfide cyclic peptide compound or an analogue thereof, the analogue comprising a derivative of the disulfide cyclic peptide compound, a pharmaceutically acceptable salt thereof, a tautomer thereof, or a stereoisomer thereof.

[0083] In one aspect of the invention, the pharmaceutical composition further includes a pharmaceutically acceptable carrier or excipient.

[0084] In one aspect of the invention, a pharmaceutically acceptable carrier or excipient refers to any formulation or carrier medium representative of the present invention capable of delivering an effective amount of the active substance without interfering with the bioactivity of the active substance and without toxic side effects on the host or subject, including water, oil, vegetables and minerals, ointment bases, lotion bases, ointment bases, etc. These bases include suspending agents, thickeners, transdermal penetration enhancers, etc.

[0085] The beneficial effects of this invention are as follows:

[0086] This invention provides the first preparation of a disulfide cyclic peptide compound with the following structure: R2-R1-c(Cys-D-Ala-His-D-Phe-Arg-Trp-Cys)-R3-R4. The disulfide cyclic peptide compound provided by this invention, as a ligand for one or more melanocortin receptors, can bind well to the melanocortin receptor and produce a good effect. Attached Figure Description

[0087] Figure 1 The MS spectrum of compound WP302-1 from Example 1 is shown below.

[0088] Figure 2 The HPLC chromatogram of compound WP302-1 from Example 1 is shown below.

[0089] Figure 3 The MS spectrum of compound WP302-2 in Example 2;

[0090] Figure 4 The HPLC chromatogram of compound WP302-2 in Example 2 is shown below.

[0091] Figure 5 The MS spectrum of compound WP302-3 in Example 3 is shown below.

[0092] Figure 6 The HPLC chromatogram of compound WP302-3 in Example 3 is shown below.

[0093] Figure 7 The MS spectrum of compound WP302-4 in Example 4;

[0094] Figure 8 The HPLC chromatogram of compound WP302-4 in Example 4 is shown below.

[0095] Figure 9 The MS spectrum of compound WP302-5 in Example 5 is shown below.

[0096] Figure 10 The HPLC chromatogram of compound WP302-5 in Example 5 is shown below.

[0097] Figure 11 The MS spectrum of compound WP302-6 in Example 6;

[0098] Figure 12 The HPLC chromatogram of compound WP302-6 in Example 6 is shown below.

[0099] Figure 13 The MS spectrum of compound WP302-7 from Example 7 is shown below.

[0100] Figure 14 The HPLC chromatogram of compound WP302-7 from Example 7 is shown below.

[0101] Figure 15 The HPLC chromatogram of compound WP300 in Comparative Example 1 is shown.

[0102] Figure 16 The MS spectrum of compound WP300 in Comparative Example 1 is shown. Detailed Implementation

[0103] The following non-limiting embodiments are intended to enable those skilled in the art to gain a more comprehensive understanding of the present invention, but do not limit the invention in any way. The following content is merely an exemplary description of the scope of protection claimed by the present invention, and those skilled in the art can make various changes and modifications to the invention based on the disclosed content, which should also fall within the scope of protection claimed in this application.

[0104] Terms and abbreviations:

[0105]

[0106] The present invention will be further described below by way of specific embodiments. Unless otherwise specified, all chemical reagents used in the embodiments of the present invention were obtained through conventional commercial means. Unless otherwise specified, all contents mentioned below are mass contents. Unless otherwise specified, it is understood that the process was carried out at room temperature.

[0107] Example 1: Ac-Arg-c(Cys-D-Ala-His-D-Phe-Arg-Trp-Cys)-D-Phe-NH2 (SEQ ID NO. 1)

[0108] The title peptide was synthesized using a fluorenylmethoxycarbonyl (Fmoc) chemical method in a 100 mL reactor.

[0109] 1 g of Ramage Amide AM resin with a loading of 0.30 mmol / g was selected, swollen, and then the Fmoc protecting group was removed. 0.35 g of Fmoc-D-Phe-OH, 468 mg of PyBop (benzotriazol-1-yl-oxytripyrrolidinephosphide hexafluorophosphate), and 0.34 mL of DIEA were weighed and dissolved in 25 mL of DMF at 0 °C. After complete dissolution, the solution was added to the reaction column and reacted at room temperature for 1 hour. The reaction progress was determined by a negative result using the ninhydrin detection method. After the coupling reaction was completed, the reaction solution was dried under vacuum, washed with 30 mL of DMF, reacted with 40 mL of 20% hexahydropyridine DMF solution for 5 min, washed once with DMF, and after washing, another 30 mL of 20% hexahydropyridine DMF solution was added, reacted for 10 min, dried under vacuum, washed three times with DMF, twice with DCM, and once with DMF.

[0110] According to the polypeptide sequence, coupling was performed sequentially from the C-terminus to the N-terminus until Fmoc-Arg(Pbf)-OH was de-Fmoc and the amino group was blocked by acetic anhydride and pyridine to form acetylation. The side chain protecting groups of Arg, Cys, His, and Trp are Pbf, Trt, Trt, and Boc, respectively. All amino acids were protected with Fmoc at the α-amino group.

[0111] The peptide was reacted with a linear peptide resin using a lysis buffer (TFA: triisopropylsilane: water = 95:2.5:2.5) to obtain approximately 386.2 mg of peptide with all side-chain protecting groups removed.

[0112] The peptide was dissolved in 400 mL of pure water, the pH was adjusted to 8.0, 1 mL of H2O2 was added, and the reaction was carried out at room temperature. After 30 min of reaction, the peptide was completely converted into a cyclic peptide by analytical HPLC.

[0113] The crude linear peptide aqueous solution was filtered through a 0.45 μm filter membrane. The purified crude peptide was then purified using high-performance liquid chromatography (HPLC): a DAC-HB50 dynamic axial compression column was used with gradient elution using mobile phases A (0.05% trifluoroacetic acid aqueous solution) and B (0.05% trifluoroacetic acid-acetonitrile solution). The sample was detected using a UV detector, and the peptide solution of the target peak was collected in fractions. After HPLC purification, 80 mL of a finished peptide liquid with a purity greater than 95% was obtained. The liquid was equilibrated with 1 / 1000 acetic acid and acetonitrile, concentrated by rotary evaporation to obtain 30 mL of liquid. This liquid was then pre-lyophilized and lyophilized to finally obtain 45.7 mg of refined peptide, namely compound WP302-1.

[0114] The structure of compound WP302-1 is shown below:

[0115]

[0116] The MS and HPLC mass spectra of compound WP302-1 are as follows: Figure 1 and Figure 2 As shown.

[0117] Example 2: Ac-Arg-c(Cys-D-Ala-His-D-Phe-Arg-Trp-Cys)-Pro-NH2 (SEQ ID NO. 2)

[0118] The title peptide was synthesized using a fluorenylmethoxycarbonyl (Fmoc) chemical method in a 100 mL reactor.

[0119] 1 g of Ramage Amide AM resin with a loading of 0.30 mmol / g was selected, swollen, and then the Fmoc protecting group was removed. 0.53 g of Fmoc-Cys(Trt)-OH, 468 mg of PyBop (benzotriazol-1-yl-oxytripyrrolidinephosphide hexafluorophosphate), and 0.34 mL of DIEA were weighed and dissolved in 25 mL of DMF at 0 °C. After complete dissolution, the solution was added to the reaction column and reacted at room temperature for 1 hour. The reaction progress was determined by a negative result using the ninhydrin detection method. After the coupling reaction was completed, the reaction solution was dried under vacuum, washed with 30 mL of DMF, reacted with 40 mL of 20% hexahydropyridine DMF solution for 5 min, washed once with DMF, and after washing, 30 mL of 20% hexahydropyridine DMF solution was added again, reacted for 10 min, dried under vacuum, washed 3 times with DMF, 2 times with DCM, and 1 time with DMF.

[0120] According to the polypeptide sequence, coupling was performed sequentially from the C-terminus to the N-terminus until Fmoc-Arg(Pbf)-OH was de-Fmoc and the amino group was blocked by acetic anhydride and pyridine to form acetylation. The side chain protecting groups of Arg, Cys, His, and Trp are Pbf, Trt, Trt, and Boc, respectively. All amino acids were protected with Fmoc at the α-amino group.

[0121] The peptide was reacted with a linear peptide resin using a lysis buffer (TFA: triisopropylsilane: water = 95:2.5:2.5) to obtain approximately 368.9 mg of peptide with all side-chain protecting groups removed.

[0122] The peptide was dissolved in 400 mL of pure water, the pH was adjusted to 8.0, 1 mL of H2O2 was added, and the reaction was carried out at room temperature. After 30 min of reaction, the peptide was completely converted into a cyclic peptide by analytical HPLC.

[0123] The crude linear peptide aqueous solution was filtered through a 0.45 μm filter membrane. The purified crude peptide was then purified using high-performance liquid chromatography (HPLC): a DAC-HB50 dynamic axial compression column was used with gradient elution using mobile phases A (0.05% trifluoroacetic acid aqueous solution) and B (0.05% trifluoroacetic acid-acetonitrile solution). The sample was detected using a UV detector, and the peptide solution of the target peak was collected in fractions. After HPLC purification, 80 mL of a finished peptide liquid with a purity greater than 95% was obtained. This liquid was equilibrated with 1 / 1000 acetic acid and acetonitrile, concentrated by rotary evaporation to obtain 30 mL of liquid. The liquid was then pre-lyophilized and lyophilized to finally obtain 42.5 mg of refined peptide, namely compound WP302-2.

[0124] The structure of compound WP302-2 is shown below:

[0125]

[0126] The MS and HPLC mass spectra of compound WP302-2 are as follows: Figure 3 and Figure 4 As shown.

[0127] Example 3: Ac-Arg-c(Cys-D-Ala-His-D-Phe-Arg-Trp-Cys)-Ile-NH2 (SEQ ID NO. 3)

[0128] The title peptide was synthesized using a fluorenylmethoxycarbonyl (Fmoc) chemical method in a 100 mL reactor.

[0129] 1 g of Ramage Amide AM resin with a loading of 0.30 mmol / g was selected, swollen, and then the Fmoc protecting group was removed. 0.32 g of Fmoc-Ile-OH, 468 mg of PyBop (benzotriazol-1-yl-oxytripyrrolidinephosphide hexafluorophosphate), and 0.34 mL of DIEA were dissolved in 250 mL of DMF at 0 °C. After complete dissolution, the solution was added to the reaction column and reacted at room temperature for 1 hour. The reaction progress was determined by a negative result using the ninhydrin detection method. After the coupling reaction was completed, the reaction solution was dried under vacuum, washed with 30 mL of DMF, and then reacted with 40 mL of 20% hexahydropyridine DMF solution for 5 min. The solution was washed once with DMF, and after washing, another 30 mL of 20% hexahydropyridine DMF solution was added, reacted for 10 min, and then dried under vacuum. The solution was washed three times with DMF, twice with DCM, and once with DMF.

[0130] According to the polypeptide sequence, coupling was performed sequentially from the C-terminus to the N-terminus until Fmoc-Arg(Pbf)-OH was de-Fmoc and the amino group was blocked by acetic anhydride and pyridine to form acetylation. The side chain protecting groups of Arg, Cys, His, and Trp are Pbf, Trt, Trt, and Boc, respectively. All amino acids were protected with Fmoc at the α-amino group.

[0131] The peptide was reacted with a linear peptide resin using a lysis buffer (TFA: triisopropylsilane: water = 95:2.5:2.5) to obtain approximately 396.3 mg of peptide with all side-chain protecting groups removed.

[0132] The peptide was dissolved in 400 mL of pure water, the pH was adjusted to 8.0, 1 mL of H2O2 was added, and the reaction was carried out at room temperature. After 30 min of reaction, the peptide was completely converted into a cyclic peptide by analytical HPLC.

[0133] The crude linear peptide aqueous solution was filtered through a 0.45 μm filter membrane. The purified crude peptide was then purified using high-performance liquid chromatography (HPLC): a DAC-HB50 dynamic axial compression column was used with gradient elution of mobile phases A (0.05% trifluoroacetic acid aqueous solution) and B (0.05% trifluoroacetic acid-acetonitrile solution). The sample was detected using a UV detector, and the peptide solution of the target peak was collected in fractions. After HPLC purification, 75 mL of a finished peptide liquid with a purity greater than 95% was obtained. This liquid was equilibrated with 1 / 1000 acetic acid and acetonitrile, concentrated by rotary evaporation to obtain 30 mL of liquid. The liquid was then pre-lyophilized and lyophilized to finally obtain 35.6 mg of refined peptide, namely compound WP302-3.

[0134] The structure of compound WP302-3 is shown below:

[0135]

[0136] The MS and HPLC mass spectra of compound WP302-3 are as follows: Figure 5 and Figure 6 As shown.

[0137] Example 4: Palm-Arg-c(Cys-D-Ala-His-D-Phe-Arg-Trp-Cys)-NH2 (SEQ ID NO.4)

[0138] The title peptide was synthesized using a fluorenylmethoxycarbonyl (Fmoc) chemical method in a 100 mL reactor.

[0139] 1 g of Ramage Amide AM resin with a loading of 0.30 mmol / g was selected, swollen, and then the Fmoc protecting group was removed. 0.53 g of Fmoc-Cys(Trt)-OH, 468 mg of PyBop (benzotriazol-1-yl-oxytripyrrolidinephosphide hexafluorophosphate), and 0.34 mL of DIEA were dissolved in 25 mL of DMF at 0 °C. After complete dissolution, the solution was added to the reaction column and reacted at room temperature for 1 hour. The reaction progress was determined by a negative result using the ninhydrin detection method. After the coupling reaction was completed, the reaction solution was dried under vacuum, washed with 30 mL of DMF, reacted with 40 mL of 20% hexahydropyridine DMF solution for 5 min, washed once with DMF, and after washing, 30 mL of 20% hexahydropyridine DMF solution was added again, reacted for 10 min, dried under vacuum, washed 3 times with DMF, 2 times with DCM, and 1 time with DMF.

[0140] According to the polypeptide sequence, coupling was performed sequentially from the C-terminus to the N-terminus until Fmoc-Arg(Pbf)-OH was obtained and Fmoc was removed. Then, palmitic acid was used to react with the α-amino group of Arg to form an amide bond. The side chain protecting groups of Arg, Cys, His, and Trp are Pbf, Trt, Trt, and Boc, respectively. All amino acids were protected with Fmoc at the α-amino position.

[0141] The peptide was reacted with a linear peptide resin using a lysis buffer (TFA: triisopropylsilane: water = 95:2.5:2.5) to obtain approximately 4.5.2 mg of peptide with all side-chain protecting groups removed.

[0142] The peptide was dissolved in 450 mL of pure water, the pH was adjusted to 8.0, 1 mL of H2O2 was added, and the reaction was carried out at room temperature. After 30 min of reaction, the peptide was completely converted into a cyclic peptide by analytical HPLC.

[0143] The crude linear peptide aqueous solution was filtered through a 0.45 μm filter membrane. The purified crude peptide was then purified using high-performance liquid chromatography (HPLC): a DAC-HB50 dynamic axial compression column was used with gradient elution of mobile phases A (0.05% trifluoroacetic acid aqueous solution) and B (0.05% trifluoroacetic acid-acetonitrile solution). The sample was detected using a UV detector, and the peptide solution of the target peak was collected in fractions. After HPLC purification, 120 mL of a finished peptide liquid with a purity greater than 95% was obtained. The liquid was equilibrated with 1 / 1000 acetic acid and acetonitrile, concentrated by rotary evaporation to obtain 50 mL of liquid. This liquid was then pre-lyophilized and lyophilized to finally obtain 32.7 mg of refined peptide, namely compound WP302-4.

[0144] The structure of compound WP302-4 is shown below:

[0145]

[0146] The MS and HPLC mass spectra of compound WP302-4 are as follows: Figure 7 and Figure 8 As shown.

[0147] Example 5: Palm-Arg-c(Cys-D-Ala-His-D-Phe-Arg-Trp-Cys)-D-Phe-NH2 (SEQ ID NO. 5)

[0148] The title peptide was synthesized using a fluorenylmethoxycarbonyl (Fmoc) chemical method in a 100 mL reactor.

[0149] 1 g of Ramage Amide AM resin with a loading of 0.30 mmol / g was selected, swollen, and then the Fmoc protecting group was removed.

[0150] Weigh 0.34 g of Fmoc-D-Phe-OH, 468 mg of PyBop (benzotriazol-1-yl-oxytripyrrolidinephosphide hexafluorophosphate), and 0.34 mL of DIEA. Dissolve them in 25 mL of DMF at 0 °C. After complete dissolution, add the solution to the reaction column and react at room temperature for 1 hour. The reaction progress is judged by a negative result using the ninhydrin detection method. After the coupling reaction is complete, dry the reaction solution and wash with 30 mL of DMF. Add 40 mL of 20% hexahydropyridine DMF solution and react for 5 min. Wash once with DMF. After washing, add another 30 mL of 20% hexahydropyridine DMF solution and react for 10 min. Dry the solution and wash three times with DMF, twice with DCM, and once with DMF.

[0151] According to the polypeptide sequence, coupling was performed sequentially from the C-terminus to the N-terminus until Fmoc-Arg(Pbf)-OH was obtained and Fmoc was removed. Then, palmitic acid was used to react with the α-amino group of Arg to form an amide bond. The side chain protecting groups of Arg, Cys, His, and Trp are Pbf, Trt, Trt, and Boc, respectively. All amino acids were protected with Fmoc at the α-amino position.

[0152] The peptide was reacted with a linear peptide resin using a lysis buffer (TFA: triisopropylsilane: water = 95:2.5:2.5) to obtain approximately 488.89 mg of peptide with all side-chain protecting groups removed.

[0153] The peptide was dissolved in 500 mL of pure water, the pH was adjusted to 8.0, 1 mL of H2O2 was added, and the reaction was carried out at room temperature. After 30 min of reaction, the peptide was completely converted into a cyclic peptide by analytical HPLC.

[0154] The crude linear peptide aqueous solution was filtered through a 0.45 μm filter membrane. The purified crude peptide was then purified using high-performance liquid chromatography (HPLC): a DAC-HB50 dynamic axial compression column was used with gradient elution using mobile phases A (0.05% trifluoroacetic acid aqueous solution) and B (0.05% trifluoroacetic acid-acetonitrile solution). The sample was detected using a UV detector, and the peptide solution of the target peak was collected in fractions. After HPLC purification, 150 mL of a finished peptide liquid with a purity greater than 95% was obtained. This liquid was equilibrated with 1 / 1000 acetic acid and acetonitrile, concentrated by rotary evaporation to obtain 65 mL of liquid. The liquid was then pre-lyophilized and lyophilized to finally obtain 35.1 mg of refined peptide, namely compound WP302-5.

[0155] The structure of compound WP302-5 is shown below:

[0156]

[0157] The MS and HPLC mass spectra of compound WP302-5 are as follows: Figure 9 and Figure 10 As shown.

[0158] Example 6: Palm-Arg-c(Cys-D-Ala-His-D-Phe-Arg-Trp-Cys)-Pro-NH2 (SEQ ID NO. 6)

[0159] The title peptide was synthesized using a fluorenylmethoxycarbonyl (Fmoc) chemical method in a 100 mL reactor.

[0160] 1 g of Ramage Amide AM resin with a loading of 0.30 mmol / g was selected, swollen, and then the Fmoc protecting group was removed. 0.30 g of Fmoc-Pro-OH, 468 mg of PyBop (benzotriazol-1-yl-oxytripyrrolidine phosphorus hexafluorophosphate), and 0.34 mL of DIEA were dissolved in 25 mL of DMF at 0 °C. After complete dissolution, the solution was added to the reaction column and reacted at room temperature for 1 hour. The reaction progress was determined by a negative result using the ninhydrin detection method. After the coupling reaction was completed, the reaction solution was dried under vacuum, washed with 30 mL of DMF, reacted with 40 mL of 20% hexahydropyridine DMF solution for 5 min, washed once with DMF, and after washing, 30 mL of 20% hexahydropyridine DMF solution was added again, reacted for 10 min, dried under vacuum, washed 3 times with DMF, 2 times with DCM, and 1 time with DMF.

[0161] According to the polypeptide sequence, coupling was performed sequentially from the C-terminus to the N-terminus until Fmoc-Arg(Pbf)-OH was obtained and Fmoc was removed. Then, palmitic acid was used to react with the α-amino group of Arg to form an amide bond. The side chain protecting groups of Arg, Cys, His, and Trp are Pbf, Trt, Trt, and Boc, respectively. All amino acids were protected with Fmoc at the α-amino position.

[0162] The peptide was reacted with a linear peptide resin using a lysis buffer (TFA: triisopropylsilane: water = 95:2.5:2.5) to obtain approximately 436.12 mg of peptide with all side-chain protecting groups removed.

[0163] The peptide was dissolved in 450 mL of pure water, the pH was adjusted to 8.0, 1 mL of H2O2 was added, and the reaction was carried out at room temperature. After 30 min of reaction, the peptide was completely converted into a cyclic peptide by analytical HPLC.

[0164] The crude linear peptide aqueous solution was filtered through a 0.45 μm filter membrane. The purified crude peptide was then purified using high-performance liquid chromatography (HPLC): a DAC-HB50 dynamic axial compression column was used with gradient elution using mobile phases A (0.05% trifluoroacetic acid aqueous solution) and B (0.05% trifluoroacetic acid-acetonitrile solution). The sample was detected using a UV detector, and the peptide solution of the target peak was collected in fractions. After HPLC purification, 120 mL of a finished peptide liquid with a purity greater than 95% was obtained. The liquid was equilibrated with 1 / 1000 acetic acid and acetonitrile, concentrated by rotary evaporation to obtain 50 mL of liquid. This liquid was then pre-lyophilized and lyophilized to finally obtain 29.6 mg of refined peptide, namely compound WP302-6.

[0165] The structure of compound WP302-6 is shown below:

[0166]

[0167] The MS and HPLC mass spectra of compound WP302-6 are shown below. Figure 11 and Figure 12 As shown.

[0168] Example 7: Palm-Arg-c(Cys-D-Ala-His-D-Phe-Arg-Trp-Cys)-Ile-NH2 (SEQ ID NO. 7)

[0169] The title peptide was synthesized using a fluorenylmethoxycarbonyl (Fmoc) chemical method in a 100 mL reactor.

[0170] 1 g of Ramage Amide AM resin with a loading of 0.30 mmol / g was selected, swollen, and then the Fmoc protecting group was removed. 0.31 g of Fmoc-Ile-OH, 468 mg of PyBop (benzotriazol-1-yl-oxytripyrrolidinephosphide hexafluorophosphate), and 0.34 mL of DIEA were dissolved in 25 mL of DMF at 0 °C. After complete dissolution, the solution was added to the reaction column and reacted at room temperature for 1 hour. The reaction progress was determined by a negative result using the ninhydrin detection method. After the coupling reaction was completed, the reaction solution was dried under vacuum, washed with 30 mL of DMF, reacted with 40 mL of 20% hexahydropyridine DMF solution for 5 min, washed once with DMF, and after washing, 30 mL of 20% hexahydropyridine DMF solution was added again, reacted for 10 min, dried under vacuum, washed three times with DMF, twice with DCM, and once with DMF.

[0171] According to the polypeptide sequence, coupling was performed sequentially from the C-terminus to the N-terminus until Fmoc-Arg(Pbf)-OH was obtained and Fmoc was removed. Then, palmitic acid was used to react with the α-amino group of Arg to form an amide bond. The side chain protecting groups of Arg, Cys, D-His, and Trp are Pbf, Trt, Trt, and Boc, respectively. All amino acids were protected with Fmoc at the α-amino position.

[0172] The peptide was reacted with a linear peptide resin using a lysis buffer (TFA: triisopropylsilane: water = 95:2.5:2.5) to obtain approximately 469.52 mg of peptide with all side-chain protecting groups removed.

[0173] The peptide was dissolved in 480 mL of pure water, the pH was adjusted to 8.0, 1 mL of H2O2 was added, and the reaction was carried out at room temperature. After 30 min of reaction, the peptide was completely converted into a cyclic peptide by analytical HPLC.

[0174] The crude linear peptide aqueous solution was filtered through a 0.45 μm filter membrane. The purified crude peptide was then purified using high-performance liquid chromatography (HPLC): a DAC-HB50 dynamic axial compression column was used with gradient elution using mobile phases A (0.05% trifluoroacetic acid aqueous solution) and B (0.05% trifluoroacetic acid-acetonitrile solution). The sample was detected using a UV detector, and the peptide solution containing the target peak was collected in fractions. After HPLC purification, 135 mL of a finished peptide liquid with a purity greater than 95% was obtained. This liquid was equilibrated with 1 / 1000 acetic acid and acetonitrile, concentrated by rotary evaporation to obtain 55 mL of liquid. The liquid was then pre-lyophilized and lyophilized to finally obtain 36.8 mg of refined peptide, namely compound WP302-7.

[0175] The structure of compound WP302-7 is shown below:

[0176]

[0177] The MS and HPLC mass spectra of compound WP302-7 are shown below. Figure 13 and Figure 14 As shown.

[0178] Comparative Example 1: Ac-Arg-c(Cys-DAla-His-DPhe-Arg-Trp-Cys)-NH2(SEQ ID NO.8)

[0179] The title peptide was synthesized using a fluorenylmethoxycarbonyl (Fmoc) chemistry in a 5 L reactor. Rink Amide 4-methyldiphenylmethylamine (MBHA) resin (Novabiochem®, San Diego, CA) with 0.451 mmol / g substitution was used. The Fmoc amino acids used in the synthesis were Fmoc-Arg(pbf)-OH, Fmoc-Cys(Trt)-OH, Fmoc-D-Ala-OH, Fmoc-His(Trt)-OH, Fmoc-D-Phe-OH, and Fmoc-Trp(Boc)-OH. The synthesis was carried out on a 50 mmol scale. The Fmoc groups were removed by treatment with 20% piperidine in N,N-dimethylformamide (DMF) for 30 min. In each coupling step, Fmoc amino acids (3 eq, 150 mmol), N,N-diisopropylcarbodiimide (DIC) (3 eq, 150 mmol), and 1-hydroxybenzotriazole (HOBT) (3 eq, 150 mmol) were coupled in DMF. The resin was subjected to the following cycle in the reactor: (1) washing with DMF, (2) removing the Fmoc protecting group by treatment with 20% piperidine in DMF for 30 min, (3) washing with DMF, and (4) coupling with Fmoc amino acids in the presence of DIC and HOBT for 1 h. The resin was successfully coupled according to the sequence of the title peptide. After the peptide chain was assembled and the final Fmoc protecting group was removed, it was acetylated with a solution of acetic anhydride, N-methylmorpholine (NMM), and DMF (v / v / v: 6 / 10 / 84). The resin was thoroughly washed with dichloromethane (DCM) and methanol and then dried to obtain a dry resin.

[0180] To cleave the title peptide, the resin was treated with a solution of TFA, phenol, EDT, H2O, and anisole (v / v / v / v / v: 87.5 / 2.5 / 2.5 / 2.5 / 5) at room temperature for 3 hours. The resin was filtered, and the filtrate was poured into diethyl ether. The precipitate was collected by centrifugation. The precipitate was washed three times with diethyl ether and centrifuged to obtain the final precipitate, which was then vacuum dried to obtain the crude peptide.

[0181] The crude peptide was dissolved in a 0.1% TFA aqueous solution, and then a 10 g / L iodine methanol solution was added dropwise to the solution for oxidative bridging. The addition of iodine was stopped when the solution changed from colorless to pale yellow. After standing for 3 minutes, vitamin C was added dropwise. The addition was stopped when the solution changed from pale yellow to colorless, indicating that the oxidation was complete. The oxidation was performed using a Luna C18 100A (650) reversed-phase preparative HPLC system. 650 The crude product after oxidation was purified using a 2350 mm (Varian) column. The column was eluted for approximately 1 hour using a linear gradient of 80% A:20% B to 50% A:50% B, where A was a 0.1% TFA aqueous solution and B was a mixture of 0.1% TFA in 80% acetonitrile and 20% water. The resulting TFA salt product (over 90% purity) was then processed on a reversed-phase preparative HPLC system using a Luna C18 100A (650 mm) column. 650 A 2350 mm (Varian) column was used for Ac salt replacement, eluting the column with a linear gradient from 100% A:0% B to 40% A:60% B for approximately 1 hour. A was a 0.5% aqueous HAc solution, and B was a mixture of 0.5% HAc, 80% acetonitrile, and 20% water. Analysis was performed using HPLC. Figure 5 After freeze-drying the qualified product, 21635 mg of a white solid with 95% purity was obtained, with a yield of 34.7%, which is compound WP300.

[0182] The structure of compound WP300 is shown below:

[0183]

[0184] The MS and HPLC mass spectra of compound WP300 are as follows: Figure 16 and Figure 15 As shown.

[0185] Experiment 1: Serum stability assay

[0186] 1. Add 594 µL of serum to a 96-well plate and pre-incubate at 37°C for 5 minutes.

[0187] 2. Add 6 µL of test sample / positive control to a 96-well plate and mix thoroughly using a pipette. Continue the reaction at 37°C with constant shaking for 48 hours.

[0188] 3. For the test sample, at 0, 0.5, 1.5, 3, 6, 24, and 48 hours, take 50 µL and add it to a solution containing 150 µL of 0.1% FA (formic acid) in methanol as an internal standard. For the positive control, at 0, 15, 30, 60, and 120 minutes, take 50 µL and add it to a solution containing 150 µL of 0.1% FA in methanol as an internal standard.

[0189] 4. Shake and mix for 10 minutes, then centrifuge at 6000 x g for 10 minutes. Inject the supernatant into an LC-MS / MS system for analysis.

[0190] The measurement results are shown in Table 1:

[0191] Table 1

[0192]

[0193] Experiment 2: EC50 Measurement

[0194] Intracellular cyclic AMP (cAMP) levels were measured using an electrochemiluminescence (ECL) assay (Meso Scale Discovery®, Gaithersburg, MD; hereinafter referred to as MSD).

[0195] 1. Remove stable cell lines MC1, MC3, MC4, and MC5 from the liquid nitrogen storage system. Thaw rapidly in a 37°C thermostatic water bath, then transfer the cell suspension to 15mL centrifuge tubes using a pipette, and add 10mL of complete culture medium. (MC1 cells: F12K + 10% FBS + 400 μg / mL G418; MC3 cells: F12K + 10% FBS + 100 μg / mL Hygromycin B + 200 μg / mL Zeocin; MC4 cells: F12K + 10% FBS + 100 μg / mL Hygromycin B + 8 μg / mL L Prouromycin; MC5 cells: F12K + 10% FBS + 100 μg / mL Hygromycin B + 400 μg / mL G418).

[0196] 2. After centrifugation at 1000 rpm for 4 minutes, discard the supernatant, resuspend the cell pellet in 5 mL of complete culture medium, transfer to a T75 culture flask, add 15 mL of culture medium, and incubate at 37°C in a 5% CO2 incubator. The cells are passaged once and then used for this cell experiment.

[0197] 3. When the cell density reaches 80%-90%, discard the culture medium and wash the cells with 5 mL of phosphate buffer. 4. Remove the phosphate buffer, add 3 mL of trypsin, and incubate at 37°C (carbon dioxide) for 2-5 minutes.

[0198] 5. Add 10 mL of complete culture medium to collect cells, centrifuge at 1000 rpm for 4 min and discard the supernatant.

[0199] 6. Adjust the cell suspension to the appropriate density using Stimulation Buffer. MC1, MC3, and MC5 cells: 1000 cells / well. MC4 cells: 2000 cells / well.

[0200] Transfer 10 μL of cell solution to an assay plate. Centrifuge at 600 rpm for 3 minutes and incubate at room temperature for 60 minutes. After incubation, add 5 μL of Eu-cAMP tracer and 5 μL of L Light™ anti-cAMP solution from the cAMP assay kit to the assay plate. Centrifuge again at 600 rpm for 3 minutes and incubate at room temperature for 60 minutes.

[0201] cAMP signals were read using a multi-functional microplate reader, and the data were processed, analyzed, and reported as EC50 values ​​using a GraphPad Prism 6. The experimental results are shown in Tables 2 to 5.

[0202] Table 2: MC1 activation experiment

[0203]

[0204] Table 3: MC3 activation experiment

[0205]

[0206] Table 4: MC4 activation experiment

[0207]

[0208] Table 5: MC5 activation experiment

[0209]

[0210] The cellular-level effect of the compound WP302 provided by this invention upon binding to the melanocortin receptor is measured using intracellular cyclic adenosine monophosphate (cAMP) levels. EC50 represents the concentration of the agonist compound required to achieve 50% of the maximum response. The results show that the compound provided by this invention produces a favorable effect on the melanocortin receptor.

[0211] This invention provides a series of compounds with the general formula R2-R1-c(Cys-D-Ala-His-D-Phe-Arg-Trp-Cys)-R3-R4; and from the above experimental data, it can be seen that as the molecular weight of the N-terminal substituent increases, the activity of the MC5 target decreases and the selectivity increases, that is, the target is related to the N-terminal substituent.

[0212] The above detailed description is a specific illustration of one feasible embodiment of the present invention, and this embodiment is not intended to limit the patent scope of the present invention. It should be noted that all equivalent implementations or modifications made without departing from the present invention should be included within the scope of the technical solution of the present invention. Therefore, the protection scope of the present invention should be determined by the appended claims.

Claims

1. A disulfide cyclic peptide compound, characterized in that, The following compounds: 。 2. A method for preparing the disulfide cyclic peptide compound of claim 1, characterized in that, The method includes the following steps: Step 1: Select a resin and remove the Fmoc protecting group to obtain a resin with the Fmoc protecting group removed; Step 2: Weigh Fmoc-R3-OH or Fmoc-Cys(Trt)-OH and PyBop, and perform a coupling reaction on the resin with the Fmoc protecting group removed to obtain the coupling resin. Step 3: According to the polypeptide sequence, for the coupling resin, couple sequentially from the C-terminus to the N-terminus to obtain a linear peptide resin; Step 4: The protecting group is removed by reacting the linear peptide resin with the lysis buffer to obtain the polypeptide; Step 5: Dissolve the polypeptide in water, adjust the pH and add H2O2 to react and obtain the disulfide cyclic peptide compound.

3. The use of the disulfide cyclic peptide compound of claim 1 or the disulfide cyclic peptide compound prepared by the method of claim 2 in the preparation of a drug for treating obesity.

4. The application according to claim 3, characterized in that, The application is achieved by the binding of the disulfide cyclic peptide compound to the melanocortin receptor.

5. The application according to claim 4, characterized in that, The melanocortin receptor is MC4R.

6. A pharmaceutical composition, characterized in that, The pharmaceutical composition comprises the disulfide cyclic peptide compound of claim 1 or a pharmaceutically acceptable salt thereof.

7. The pharmaceutical composition according to claim 6, characterized in that, The pharmaceutical composition also includes a pharmaceutically acceptable carrier or excipient.

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