Triple-target peptide and derivative thereof

Abstract

The present invention relates to the field of peptide drugs, and particularly to a triple-target peptide and a derivative thereof. The triple-target peptide provided by the present invention exerts effects of lowering blood glucose, reducing blood lipids, and weight loss. Based on the above studies, the triple-target peptide provided by the present invention is further long-acting modified, and while retaining excellent effects for the specific indications described above, also has long-acting advantages, which can reduce dosing frequency, improve medication convenience, and enhance patient compliance. Experimental results show that the peptide and the long-acting derivative thereof provided by the present invention can significantly lower blood glucose, reduce blood lipids and / or induce weight loss, and can effectively prevent, treat, or ameliorate diabetes, obesity, obesity-related diseases, or metabolic disorders.

Description

Tri-target peptides and their derivatives

[0001] This application claims priority to Chinese Patent Application No. 202411862982.5, filed on December 16, 2024, entitled “Three-target polypeptides and their derivatives”, the entire contents of which are incorporated herein by reference. Technical Field

[0002] This invention relates to the field of polypeptide drugs, and particularly to three-target polypeptides and their derivatives. Background Technology

[0003] 1. Obesity, lipid-lowering

[0004] (1) Disease overview:

[0005] Globally, over one billion people, or one in eight, suffer from obesity. It is estimated that by 2035, the number of overweight and obese individuals (BMI ≥ 25 kg / m²) worldwide may exceed 4 billion. In 2020, this number already exceeded 2.6 billion. The proportion of the overweight and obese population increased from 14% in 1990 to 24% in 2020. Data from the past 35 years shows a significant upward trend in global obesity rates. Based on the provided data, the following key points regarding the epidemiology of obesity and overweight in China and the United States can be summarized:

[0006] China's overweight and obesity problem is becoming increasingly serious. The latest data shows that more than half of Chinese adults (50.7%) are overweight or obese; the overweight rate for children under 6 years old is 6.8%, and the obesity rate is 3.6%; for adolescents aged 6-17, the overweight rate is 11.1%, and the obesity rate is 7.9%. There are significant regional differences in the overweight and obese population in China, with a generally higher rate in the north than in the south. The prevalence is higher among men and the elderly. In recent years, the rate of overweight and obesity has been rising faster in rural areas than in urban areas, and the urban-rural gap is gradually narrowing.

[0007] Overweight and obesity are equally serious problems in the United States. Data from 2015-2016 shows that 39.8% of adults were obese, and 31.8% of teenagers were overweight or obese. It is projected that by 2030, nearly half of American adults will be obese. Obesity rates in the United States vary across races, regions, genders, and ages. Obesity rates are higher among African Americans and Hispanics than among whites and Asians, and are generally higher in Southern states.

[0008] Overweight and obesity lead to a variety of health problems and are risk factors for many chronic diseases such as diabetes and heart disease. In both China and the United States, overweight and obesity have become leading causes of death and disability. The pathogenesis of obesity is complex and related to multiple factors, including genetics, lifestyle, and socioeconomic status. Increased consumption of fast food and decreased physical activity are significant contributing factors. Both China and the United States urgently need to implement effective public health policies, strengthen public education, promote healthy lifestyles, curb the obesity epidemic, and reduce the disease burden it causes.

[0009] (2) Progress of mainstream treatment options and pipeline projects:

[0010] The "Expert Consensus on the Prevention and Treatment of Obesity in Chinese Residents" points out that when lifestyle interventions are ineffective, patients with a BMI ≥ 30 kg / m² or a BMI ≥ 27 kg / m² accompanied by obesity-related diseases (such as hypertension and type 2 diabetes) may consider drug treatment. The "Guidelines for Medical Nutrition Therapy for Overweight / Obesity in China (2021)" recommends that adult patients with a BMI ≥ 27 kg / m² and obesity-related complications use medication for weight loss in addition to lifestyle interventions. Some medications for treating diabetes, such as GLP-1 receptor agonists, are also recommended for obese or overweight diabetic patients, but the available options are very limited.

[0011] The 2022 American Gastroenterological Association (AGA) guidelines recommend that for obese or overweight adults with weight-related complications, if lifestyle interventions are ineffective, medication should be added to lifestyle interventions, rather than using continuous lifestyle interventions alone. FDA-approved weight-loss drugs include orlistat, lorcaserin, phentermine / topiramate extended-release, liraglutide, semaglutide, and naltrexone / bupropion extended-release. Based on weight-loss efficacy, the 2022 AGA guidelines recommend semaglutide, phentermine / topiramate extended-release, liraglutide, and naltrexone / bupropion extended-release for obese or overweight adult patients with complications. The 2015 American Endocrine Society guidelines state that patients with a BMI ≥ 30 kg / m² or a BMI ≥ 27 kg / m² with complications such as hypertension, hyperlipidemia, or type 2 diabetes may consider drug therapy. The use of weight-loss drugs requires evaluation of efficacy and safety; generally, it is recommended that at least 5% weight loss be achieved before continuing medication.

[0012] In summary, both Chinese and American guidelines recommend the use of medication in addition to lifestyle interventions for obese patients who do not respond well to them. Currently, only orlistat is approved for weight loss in China, while the United States has a variety of weight loss drugs available and recommends drugs with novel mechanisms such as GLP-1 receptor agonists. Clinical use of weight loss drugs requires a careful balance of benefits and risks, and regular assessments of efficacy and safety are necessary.

[0013] GLP-1 receptor agonists are currently a hot target in weight loss drug development. GLP-1 drugs approved for marketing in China include Novo Nordisk's liraglutide (indications include overweight / obesity) and semaglutide (approved only for type 2 diabetes). Several domestic pharmaceutical companies are developing GLP-1 drugs for weight loss indications, such as Huadong Medicine, Livzon Pharmaceutical Group, Hanyu Pharmaceutical, and Tonghua Dongbao. Oral and long-acting formulations, as well as drugs with new mechanisms such as GLP-1 / GIP dual agonists, are expected to further enrich the selection of weight loss drugs. Examples include Novo Nordisk's oral semaglutide and Eli Lilly's Tirzepatide. With the rising incidence of obesity and the approval of new drugs, the Chinese weight loss drug market is expected to grow rapidly, potentially reaching 38.3 billion yuan by 2030.

[0014] Both Chinese and American guidelines recommend drug intervention as an important means of obesity treatment. GLP-1 receptor agonists are currently the most popular target for weight loss drug development, and oral and long-acting formulations are expected to improve patient compliance. Domestic pharmaceutical companies are actively investing in this area, and the market prospects are broad. However, attention should be paid to the selection of drug indications and their rational use to avoid blind or inappropriate use.

[0015] (3) Unmet clinical needs:

[0016] The following clinical needs remain unmet in the treatment of obesity in the United States regarding drug efficacy:

[0017] In the United States, the number of approved weight-loss drugs is limited. Currently, the FDA has approved fewer than 10 drugs for the long-term treatment of obesity, including orlistat and semaglutide. Compared to the vast obese population, the existing drug options are limited and cannot meet diverse clinical needs. Some weight-loss drugs have poor efficacy. Early weight-loss drugs, such as amphetamines, while effective, were highly addictive and had significant side effects, and most have been banned. The weight-loss efficacy of existing approved drugs also needs further improvement to better control weight. Taking semaglutide as an example, according to the provided information, although semaglutide, as a new type of weight-loss drug, has significant efficacy, it also has some major drawbacks:

[0018] ① Increased risk of gastrointestinal adverse reactions

[0019] A recent study in JAMA shows that GLP-1 agonists, including semaglutide, may increase the risk of gastrointestinal diseases such as pancreatitis, gastroparesis, and intestinal obstruction by 4 to 9 times.

[0020] Common adverse reactions to oral smegglutide tablets include nausea, vomiting, diarrhea, constipation, and stomach pain. The higher the dose, the more pronounced the side effects.

[0021] ②Injection administration is inconvenient to carry

[0022] Although it has the advantage of one injection per week, injection is still less convenient than oral administration.

[0023] ③ Potential risk of abuse

[0024] There is a risk that it may be misused by non-obese individuals for non-medical weight loss purposes.

[0025] ④ Long-term safety needs to be observed.

[0026] As a novel drug, the safety of semaglutide for long-term use still needs further evaluation.

[0027] Therefore, although smegglutinin has shown excellent efficacy in weight loss, issues such as the risk of gastrointestinal adverse reactions still need to be addressed to better meet clinical needs. At the same time, the potential risk of its misuse should also be noted.

[0028] Furthermore, the huge market demand for weight-loss drugs (estimated at $200 billion by 2035) and the resulting drug shortages affect patient access. For example, the high demand for new weight-loss drugs like semaglutide leads to supply shortages, impacting patients' ability to use the medication normally. Increasing production capacity will take time. The drugs are also expensive, placing a heavy burden on patients. New weight-loss drugs like semaglutide can cost up to $10,000 per year, with limited health insurance coverage, resulting in a significant financial burden on patients and affecting drug accessibility and adherence. Finally, the development of new weight-loss drugs remains crucial. New weight-loss drugs with better efficacy, higher safety, and easier use are needed to meet the diverse needs of patients, specifically targeting different types of obesity and patient characteristics.

[0029] Compared to the United States, China has a much larger unmet clinical need for weight loss indications. The number of obese people in China increased from 191 million in 2017 to 230 million in 2021, and is projected to reach 329 million by 2030. Currently, China has more obese people than the United States, ranking first in the world.

[0030] Furthermore, there are differences between China and the US in their approaches to lifestyle interventions for obesity. The US has established a health insurance system covering the entire lifespan and specific population groups, while China lacks a widely accepted lifestyle intervention program for obesity management. The US started clinical trials for new obesity drugs earlier, at more advanced stages, and has approved more drugs for market. While China has conducted more clinical trials in recent years, approval for market access still takes time. In summary, significant differences exist between China and the US in obesity incidence, accessibility to treatment, and medical insurance support, leading to different clinical needs for obesity treatment. China needs to accelerate the research and approval of new drugs and expand patient accessibility to meet the treatment needs of its large obese population.

[0031] (4) Obesity-related diseases

[0032] Obesity and obstructive sleep apnea (OSA)

[0033] Obesity is the most important risk factor for OSA. There is a bidirectional causal relationship between obesity and OSA. While obesity leads to OSA, OSA can also lead to further weight gain through various mechanisms such as appetite hormone imbalance, fatigue, and low mood. Obesity and OSA often coexist and share similar complications such as diabetes, hypertension, and cardiovascular disease. Weight loss can improve OSA, and OSA treatment can also help control weight and metabolic disorders.

[0034] In China, being overweight and obese can increase the risk of OSA by 1.5 to 2.4 times. In the United States, approximately 70% of adults with OSA are obese. The higher the BMI, the higher the prevalence of OSA. Weight gain exacerbates OSA. Studies show that for every 10% increase in weight, the risk of OSA increases by 30%, and the acute shoulder index (AHI) increases by 32%. Conversely, a 10% weight loss can decrease the AHI by 26%. In children, obesity is also a major risk factor for OSA. Obese children have a 46% higher risk of OSA than children of normal weight. The impact of obesity on OSA is even more significant during adolescence.

[0035] In China, the overall prevalence of obstructive sleep apnea (OSA) is approximately 4%, with a higher prevalence among obese individuals. A survey in Shanghai showed an OSA prevalence of 3.62% in people over 30 years of age. In the United States, OSA affects approximately 25% of adults, with this figure reaching as high as 45% in obese individuals. With the increasing prevalence of obesity, the OSA prevalence is expected to rise further. Furthermore, abdominal obesity is more closely associated with OSA. Fat deposits around the abdominal cavity and pharynx narrow the upper airway, making it more prone to collapse and thus leading to OSA.

[0036] In summary, obesity is the most significant risk factor for OSA, and the two factors are mutually reinforcing, creating a vicious cycle that places a heavy burden on individual and public health. Screening for and treating OSA while simultaneously preventing and controlling obesity is crucial for controlling the prevalence of both diseases.

[0037] Obesity and Cardiovascular Disease (CVD)

[0038] Obesity is one of the major risk factors for CVD. In China, the risk of CVD is significantly higher in overweight and obese individuals than in those of normal weight. Obesity can increase the risk of CVD by 1.5-2.4 times. With the continued rise in obesity rates in both China and the United States, the burden of CVD will also increase. In 2019, 549,500 CVD deaths in China were attributed to high BMI. Obesity leads to CVD through a variety of direct and indirect pathophysiological mechanisms. Abdominal obesity is more closely related to CVD. Weight loss can improve metabolic indicators and reduce the risk of CVD.

[0039] 2. Diabetes, blood sugar lowering

[0040] (1) Disease overview:

[0041] The prevalence of diabetes in China has been steadily rising. It was 9.7% in 2007, 10.4% in 2013, and reached 11.2% in 2017. It is projected that by 2045, the number of people with diabetes in China will reach 174.4 million. A national epidemiological survey conducted from 2015 to 2017 showed that the prevalence of diabetes among people aged 18 and above in China was 12.8%, and the prevalence of prediabetes was 35.2%. The estimated total number of people with diabetes in China is 129.8 million. The prevalence of diabetes is higher among men, the elderly, and urban residents in China. It is higher in southern regions than in northern regions. Obesity is a major risk factor. In the United States, the overall prevalence of diabetes in 2019 was 11.3%, or 37.3 million people. Of these, 8.7% were diagnosed, and 23% were undiagnosed.

[0042] In the United States, 29.2% of people aged 65 and older have diabetes. In 2014, the global prevalence of diabetes among adults aged 18 and older was 8.5%. Between 2000 and 2019, the standardized mortality rate from diabetes worldwide increased by 3%.

[0043] In both China and the United States, the control of risk factors among diabetic patients is not ideal, and diabetes and its complications place a huge burden on the healthcare system, necessitating strengthened prevention and control measures.

[0044] (2) Progress of mainstream treatment regimens and pipelines under development

[0045] Currently, diabetes treatment drugs are divided into two main categories: oral medications and injectable formulations.

[0046] Oral hypoglycemic agents mainly include insulin secretagogues, non-insulin secretagogues, dipeptidyl peptidase-4 inhibitors (DPP-4 inhibitors), and sodium-glucose cotransporter 2 inhibitors (SGLT-2 inhibitors).

[0047] Injectable formulations include insulin and insulin analogs, and glucagon-like peptide-1 receptor agonists (GLP-1 receptor agonists).

[0048] GLP-1 receptor agonists were initially used to treat type 2 diabetes, but their indications have since expanded to include obesity, cardiovascular disease, non-alcoholic steatohepatitis, Alzheimer's disease, and chronic kidney disease. The penetration rate of GLP-1 drugs among type 2 diabetes patients in China is low, indicating significant potential for future growth. Besides single-target GLP-1 drugs, multi-target combinations of GLP-1 with GIP, GCG, and other receptors are also a hot research area. The GLP-1 / GIP dual-target drug telpotetide has been approved for marketing, demonstrating superior efficacy in lowering blood sugar and weight loss. Triple-target GLP-1 / GCG and GLP-1 / GIP / GCG drugs are also expected to further improve efficacy. Eli Lilly's GLP-1 / GIP / GCG triple-target drug Retatrutide has shown outstanding clinical efficacy, resulting in an average weight loss of 24.2% after 48 weeks of treatment. Several domestic companies have also developed multi-target GLP-1 drugs, such as Federal Pharmaceuticals' UBT251 and Huadong Medicine's DR10624. China's insulin market is growing rapidly, and diabetes medications have become the second most used drug globally. The risk of developing diabetes in obese individuals in China is five times higher than in non-obese individuals. GLP-1 inhibitors are expected to maintain strong growth in the diabetes and obesity fields. Summary of the Invention

[0049] In view of this, the present invention provides a three-target peptide and its derivatives. The peptides and their long-acting derivatives provided by the present invention can significantly lower blood sugar, lower lipids and / or reduce weight, and can effectively prevent, treat or improve diabetes, obesity, obesity-related diseases or metabolic abnormalities.

[0050] To achieve the above-mentioned objectives, the present invention provides the following technical solution:

[0051] In a first aspect, the present invention provides a three-target polypeptide or a pharmaceutically acceptable salt, solvate, hydrate, complex, chelate, derivative, variant, or analog thereof, wherein the polypeptide structure is shown in Formula I:

[0052] H-X2-QGTFTSDYSKYLDX 16 -X 17 -AAX 20 -EFX 23 -X 24 -WLX 27 -X 28 -X 29 -GPSSGAPPPSDRVYIHP

[0053] Formula I

[0054] in:

[0055] X2 is selected from G, Aib, or dS;

[0056] X 16 Selected from E or Aib;

[0057] X 17 Selected from Q, K or

[0058] X 20 Selected from K or

[0059] X 23 Selected from I or V;

[0060] X 24 Choose from A or E;

[0061] X 27 Selected from M or L;

[0062] X 28 Selected from N, E, D, or S;

[0063] X 29 Selected from T or G;

[0064] dS stands for D-SER;

[0065] This represents the Lys side chain -NH2 linked to the fatty acid chain;

[0066] Preferably, the Lys side chain -NH2 is connected to the fatty acid chain via a connector;

[0067] Preferably, the acyl group on the connector forms an amide bond with the Lys side chain -NH2, and the fatty acid chain HOOC-(CH2) n -CO- forms an amide bond with the amino group on the connector, and the structure of the connector is shown in Formula II:

[0068] Wherein, n is 12-20, preferably 14-18, and more preferably 16;

[0069] Where m is 0-4, preferably 1-3, and more preferably 2;

[0070] Preferably, the fatty acid chain comprises C18 diacid-Gamma-GLU-2OEG-.

[0071] In some specific embodiments of the present invention, the polypeptide has:

[0072] (I) An amino acid sequence as shown in any one of SEQ ID No. 1 to 31;

[0073] (II) An amino acid sequence that is functionally identical to the amino acid sequence described in (I) obtained by substituting, deleting, or adding one, two, or three amino acids to the amino acid sequence described in (I); or

[0074] (III) An amino acid sequence having 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% or more homology with the amino acid sequence described in (I) or (II).

[0075] In some specific embodiments of the present invention, the peptide chain ends of the polypeptide are free or chemically modified;

[0076] Preferably, the chemical modification includes chemical modification at the amino terminus or chemical modification at the carboxyl terminus;

[0077] Preferably, the chemical modification of the amino terminus includes acylation, sulfonation, alkylation, and PEG modification; the chemical modification of the carboxyl terminus includes amidation, sulfonamideation, and PEG modification.

[0078] Preferably, the chemical modification of the amino terminus is acetylation, benzoylation, or sulfonation of the amino group; the alkylation of the amino terminus is C1-6 alkylation or aralkylation; and the chemical modification of the carboxyl terminus is that the OH group in the carboxyl group is replaced by NH2, or the carboxyl group is replaced by sulfonamide, or the OH group in the carboxyl group is connected to a functionalized PEG molecule.

[0079] In a second aspect, the present invention also provides a medicament comprising the polypeptide or a pharmaceutically acceptable salt, solvate, hydrate, complex, chelate, derivative, variant or analog thereof, and at least one of a pharmaceutically acceptable excipient, excipient, carrier and solvent.

[0080] Preferably, it may also contain other active ingredients.

[0081] Thirdly, the present invention also provides the use of the said polypeptide or its pharmaceutically acceptable salts, solvates, hydrates, complexes, chelates, derivatives, variants or analogs, or polypeptides targeting GLP-1R, GCGR and MASR, in the preparation of drugs for lowering blood sugar, lowering blood lipids and / or losing weight, or drugs for preventing, treating and / or improving diabetes, obesity, obesity-related diseases or metabolic disorders.

[0082] Fourthly, the present invention also provides a method for lowering blood sugar, lowering blood lipids, and / or losing weight, or a method for preventing, treating, and / or improving diabetes, obesity, obesity-related diseases, or metabolic abnormalities, by administration, ingestion, or use of any of the following:

[0083] (I) The polypeptide or a pharmaceutically acceptable salt, solvate, hydrate, complex, chelate, derivative, variant, or analog thereof; or

[0084] (II) The drug.

[0085] The three-target peptides provided by this invention exert hypoglycemic, lipid-lowering, and weight-loss effects. Based on the aforementioned research, by extending the duration of action of the three-target peptides provided by this invention, the excellent effects for specific indications are retained while also possessing the advantage of long-acting action. This reduces the frequency of dosing, improves the convenience of medication, and enhances patient compliance. Experimental results show that the peptides provided by this invention and their extended-acting derivatives can significantly lower hypoglycemia, lower lipids, and / or cause weight loss, and can effectively prevent, treat, or improve diabetes, obesity, obesity-related diseases, or metabolic abnormalities. Attached Figure Description

[0086] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the accompanying drawings used in the description of the embodiments or the prior art will be briefly introduced below.

[0087] Figure 1 shows the changes in fasting blood glucose during drug administration; *** P<0.001 vs. Control; # P<0.05, ## P<0.01, ### P<0.001, vs. Vehicle;

[0088] Figure 2 shows random blood glucose changes during drug administration; *** P<0.001 vs. Control; ## P<0.01, ### P<0.001, vs. Vehicle;

[0089] Figure 3 shows the changes in glycated hemoglobin 6 weeks after drug administration; *** P<0.001 vs. Control; ## P<0.01, ### P<0.001, vs. Vehicle;

[0090] Figure 4 shows the effect on blood lipids;

[0091] Figure 5 shows the effect on body weight;

[0092] Figure 6 shows the effect on the rate of change in body weight (vs. Vehicle);

[0093] Figure 7 shows the effect on the rate of change in body weight (vs. Day 0);

[0094] Figure 8 shows the effects on fat weight, body fat index, and Lee's index;*** P<0.001 vs. Control; # P<0.05, ## P<0.01, ### P<0.001, vs.Vehicle; LY: LY3437943; Tirze: Tirzepatide;

[0095] Figure 9 shows the effect on the fat percentage in mice; * P<0.05, ** P<0.01 vs. Control; # P<0.05vs.Vehicle; LY: LY3437943; Tirze: Tirzepatide;

[0096] Figure 10 shows the weekly changes in food intake during the drug administration period;

[0097] Figure 11 shows the effect on fasting blood glucose; *** P<0.001 vs. Control; # P<0.05, ## P<0.01, ### P<0.001, vs. Vehicle;

[0098] Figure 12 shows the effect on random blood glucose; *** P<0.001 vs. Control; # P<0.05, ## P<0.01, ### P<0.001, vs. Vehicle;

[0099] Figure 13 shows the effect on glycated hemoglobin; *** P<0.001 vs. Control; # P<0.05, ## P<0.01, ### P<0.001, vs.Vehicle; LY: LY3437943; Tirze: Tirzepatide;

[0100] Figure 14 shows the effect on blood lipids; *** P<0.001 vs. Control; # P<0.05, ## P<0.01, ### P<0.001, vs.Vehicle; LY: LY3437943; Tirze: Tirzepatide;

[0101] Figure 15 shows the pharmacokinetic study of peptide 31. Detailed Implementation

[0102] This invention discloses three-target peptides and their derivatives. Those skilled in the art can refer to the content of this document and appropriately modify the process parameters to achieve the desired results. It should be particularly noted that all similar substitutions and modifications are obvious to those skilled in the art and are considered to be included in this invention. The methods and applications of this invention have been described through preferred embodiments. Those skilled in the art can clearly modify or appropriately change and combine the methods and applications described herein without departing from the content, spirit, and scope of this invention to realize and apply the technology of this invention.

[0103] This invention provides a three-target polypeptide or a pharmaceutically acceptable salt, solvate, hydrate, complex, chelate, derivative, variant, or analog thereof, wherein the polypeptide structure is shown in Formula I:

[0104] H-X2-QGTFTSDYSKYLDX 16 -X 17 -AAX 20 -EFX 23 -X 24 -WLX 27 -X 28 -X 29 -GPSSGAPPPSDRVYIHP

[0105] Formula I

[0106] in:

[0107] X2 is selected from G, Aib, or dS;

[0108] X 16 Selected from E or Aib;

[0109] X 17 Selected from Q, K or

[0110] X 20 Selected from K or

[0111] X 23 Selected from I or V;

[0112] X 24 Choose from A or E;

[0113] X 27 Selected from M or L;

[0114] X 28 Selected from N, E, D, or S;

[0115] X 29 Selected from T or G;

[0116] dS stands for D-SER;

[0117] This represents the Lys side chain -NH2 linked to the fatty acid chain;

[0118] Preferably, the Lys side chain -NH2 is connected to the fatty acid chain via a connector;

[0119] Preferably, the acyl group on the connector forms an amide bond with the Lys side chain -NH2, and the fatty acid chain HOOC-(CH2) n -CO- forms an amide bond with the amino group on the connector, and the structure of the connector is shown in Formula II:

[0120] Wherein, n is 12-20, preferably 14-18, and more preferably 16;

[0121] Where m is 0-4, preferably 1-3, and more preferably 2;

[0122] Preferably, the fatty acid chain comprises C18 diacid-Gamma-GLU-2OEG-.

[0123] The polypeptide has the following characteristics:

[0124] (I) An amino acid sequence as shown in any one of SEQ ID No. 1 to 31;

[0125] (II) An amino acid sequence that is functionally identical to the amino acid sequence described in (I) obtained by substituting, deleting, or adding one, two, or three amino acids to the amino acid sequence described in (I); or

[0126] (III) An amino acid sequence having 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% or more homology with the amino acid sequence described in (I) or (II).

[0127] The peptide chain ends of the polypeptide are either free or chemically modified;

[0128] Preferably, the chemical modification includes chemical modification at the amino terminus or chemical modification at the carboxyl terminus;

[0129] Preferably, the chemical modification of the amino terminus includes acylation, sulfonation, alkylation, and PEG modification; the chemical modification of the carboxyl terminus includes amidation, sulfonamideation, and PEG modification.

[0130] Preferably, the chemical modification of the amino terminus is acetylation, benzoylation, or sulfonation of the amino group; the alkylation of the amino terminus is C1-6 alkylation or aralkylation; and the chemical modification of the carboxyl terminus is that the OH group in the carboxyl group is replaced by NH2, or the carboxyl group is replaced by sulfonamide, or the OH group in the carboxyl group is connected to a functionalized PEG molecule.

[0131] Preferably, the three targets include GLP-1R, GCGR, and MASR.

[0132] Beneficial effects include, but are not limited to:

[0133] (1) This molecule achieves synergistic effects on three targets: GLP-1R, GCGR, and MASR, working together to lower blood sugar, lower lipids, and reduce weight. The mechanism of action is as follows:

[0134] Once activated, GLP-1R enhances insulin secretion in a glucose concentration-dependent manner, inhibits glucagon secretion, delays gastric emptying, increases satiety, reduces food intake through central appetite suppression, and increases energy expenditure through central and peripheral mechanisms, thereby achieving effects such as lowering blood sugar and weight loss. When GCGR is activated, its glycemic effect can be countered by the insulin-stimulating and glucagon-inhibiting effects of GLP-1R. The blood sugar-raising effect of GCGR activation can also mitigate the hypoglycemic side effects caused by excessive GLP-1R effects. Furthermore, the increased fatty acid oxidation and thermogenesis, reduced fat deposition, and increased energy expenditure of GCGR, after activation, can synergistically reduce weight with GLP-1R. The synergistic effect of GLP-1R and GCGR can increase insulin secretion, enhance insulin sensitivity, improve insulin resistance and β-cell dysfunction, suppress appetite, and reduce weight, while significantly reducing the occurrence of hypoglycemic symptoms. Simultaneously, the activation of MasR can enhance insulin-induced skeletal muscle glucose uptake and improve insulin sensitivity, synergistically enhancing the hypoglycemic effect with the insulin-stimulating effect of GLP-1R.

[0135] (2) Advantages of long-term effectiveness:

[0136] The above three target molecules have been extended in duration, retaining the excellent effects for specific indications while also having the advantage of long-term effect, which can reduce the frequency of administration, improve the convenience of medication, and enhance patient compliance.

[0137] ① Improve patient compliance: Traditional peptide drugs require frequent administration, which is inconvenient for patients. Long-acting drugs can extend the dosing interval and improve patient compliance.

[0138] ② Maintaining effective blood drug concentration: Long-acting drugs can maintain a relatively stable effective blood drug concentration in the body, avoiding large fluctuations in blood drug concentration.

[0139] ③ Reduce adverse reactions: Long-acting formulations can avoid adverse reactions caused by excessively high peak blood drug concentrations.

[0140] ④ Improve efficacy: Compared with intermittent dosing, continuous exposure to the effective concentration range may improve efficacy.

[0141] ⑤ Reduce systemic exposure: Long-lasting formulation can reduce systemic exposure and lower the risk of systemic toxicity.

[0142] Long-lasting technological advantages

[0143] Commonly used methods for extending the duration of action include sustained-release formulations, chemical modification, and fusion proteins. Different extension technologies have different advantages and disadvantages, and the appropriate technical route must be selected based on factors such as the specific drug's physicochemical properties and route of administration. The molecule in this application employs chemical modification, using fatty acid chains to modify the peptide, which can enhance the compound's stability and prolong its half-life.

[0144] The raw materials and reagents used in the three-target peptides and their derivatives provided by this invention are all commercially available.

[0145] The present invention will be further illustrated below with reference to the embodiments:

[0146] Example 1 Molecular Design

[0147] Table 1 shows the active peptides and their long-acting derivatives designed targeting the three GLP1R / GCGR / MasR sites.

[0148] Table 1. Active peptides targeting GLP1R / GCGR / MasR and their long-acting derivatives

[0149] Note 1: The series of polypeptides and polypeptide derivatives involved in this invention are named in Table 1 using the naming method of "polypeptide 1, polypeptide 2...polypeptide 31".

[0150] Note 2: (dS) represents D-SER; The Lys side chain -NH2 is linked to the fatty acid chain C18diacid-Gamma-GLU-2OEG-. Taking polypeptide 29 as an example, its structure is shown in Formula I:

[0151] Example 2: Preparation method of polypeptide

[0152] Taking polypeptide 29 as an example, the preparation method is as follows:

[0153] 1. Synthesis

[0154] Washing: DMF×1, MTBE×1, DMF×2, 2 minutes each time.

[0155] Coupling: ① Resin:AA:Oxyma:DIC = 1:3:3.3:3.75 (molar ratio)

[0156] ②Resin:AA:HATU:DIEA=1:3:2.85:6 (molar ratio)

[0157] Deprotection solution: 20% Pip / 0.1M HoBt / DMF 5 min × 1, 20 min × 1

[0158] Kaisertest solution: The Kaisertest solution is made by dissolving 5g of ninhydrin in 100ml of LEtOH.

[0159] Indene detection method: Take a small test tube and add a small amount of EtOH. Take a few resin grains that have reacted into the test tube, add 3-5 drops of indene detection solution, heat at 110℃ for 3-5 minutes, and observe the color of the resin to determine positive / negative.

[0160] (1) Preparation of Fmoc-Pro-CTC Resin

[0161] Weigh 11.0 g (Sub = 0.65 mmol / g) of CTC Resin into a reaction column. Weigh 6.1 g of Fmoc-Pro-OH and add DCM to a beaker. Weigh 4.7 g of DIEA, dissolve it in a beaker, and add it to the reaction column. Add DCM and bubble evenly. React at RT for 2.5-3 h. Add MeOH / DIEA = 9 / 160 mL to seal the reaction and react for 0.5 h. Add DCM × 1 and DMF × 2. Fmoc-Pro-CTC Resin is obtained, with an estimated 6 mmol.

[0162] (2) Deprotection

[0163] Half of the resin, approximately 3 mmol of Fmoc-Pro-CTC Resin, was swollen for 0.5 h. 60 mL of deprotection solution was added to a 2 L synthesis column, and the column was bubbled with nitrogen and stirred for 5 min to allow the deprotection reaction to proceed. The solvent was then drained. Another 60 mL of deprotection solution was added to the 2 L synthesis column, and the column was bubbled with nitrogen and stirred for 20 min to allow the deprotection reaction to proceed. The solvent was then drained. The resin was washed under nitrogen bubbling and stirring conditions, and the solvent was drained. A Kaisertest test was performed on the resin, and it showed a positive result.

[0164] (3) Peptide chain elongation

[0165] Weigh 5.6 g of Fmoc-His(Trt)-OH and 1.5 g of Oxyma into a 100 mL beaker, add 50 mL of DMF, stir to dissolve, add 1.75 g of DIC, stir until dissolved, and pour the solution into the reaction column. Turn on nitrogen bubbling and stirring, control the reaction temperature at 25℃, and react for 1 h ± 0.5 h. Take a trace amount of resin for Kaisertest testing. If the resin shows a negative result, stop the reaction and drain the solvent. Wash the resin under nitrogen bubbling and stirring conditions, and drain the solvent.

[0166] Using the above-mentioned similar test method, sequentially couple Fmoc-Ile-OH, Fmoc-Tyr(tBu)-OH, Fmoc-Val-OH, Fmoc-Arg(Pbf)-OH, Fmoc-Asp (OtBu)-OH, Fmoc-Ser(tBu)-OH, Fmoc-Pro-OH, Fmoc-Pro-OH, Fmoc-Pro-OH, Fmoc-Ala-OH, Fmoc-Gly -OH, Fmoc-Ser(tBu)-OH, Fmoc-Ser(tBu)-OH, Fmoc-Pro-OH, Fmoc-Gly-OH, Fmoc-Gly-OH, Fmoc-Glu (OtBu)-OH, Fmoc-Leu-OH, Fmoc-Leu-OH, Fmoc-Trp-OH, Fmoc-Ala-OH, Fmoc-Val-OH, Fmoc-Phe-OH, F moc-Glu(OtBu)-OH, Fmoc-Lys(Boc)-OH, Fmoc-Ala-OH, Fmoc-Ala-OH, Fmoc-Lys(Alloc)-OH, Fmoc- Glu(OtBu)-OH, Fmoc-Asp(OtBu)-OH, Fmoc-Leu-OH, Fmoc-Tyr(tBu)-OH, Fmoc-Lys(Boc)-OH, Fmoc-S er(tBu)-OH, Fmoc-Tyr(tBu)-OH, Fmoc-Asp(OtBu)-OH, Fmoc-Ser(tBu)-OH, Fmoc-Thr(tBu)-OH, Fmo c-Phe-OH, Fmoc-Thr(tBu)-OH, Fmoc-Gly-OH, Fmoc-Gln(Trt)-OH, Fmoc-Aib-OH, Boc-His(Trt)-OH.

[0167] (4) Lys-coupled fatty acid chains

[0168] After coupling the main chain, the protecting group Alloc was removed using Pd(PPh3)4 / NMM / DCM. Following the same coupling ratio, Fmoc-AEEA-OH, Fmoc-AEEA-OH, Fmoc-Glu-OtBu, and tert-butyl octadecanoate were sequentially added to obtain the peptide resin. The resin was washed with DCM and anhydrous methanol under nitrogen bubbling and stirring conditions, and the solvent was drained. After vacuuming the reaction column for 1 hour, the peptide resin was dried in a vacuum drying oven (vacuum degree ≤ -0.08 MPa, oven temperature set to 15~30℃) until constant weight (constant weight standard: weight loss rate within 5 minutes measured by a rapid drying loss meter: ≤5%), yielding 14.0 g of peptide resin.

[0169] 2. Pyrolysis

[0170] (1) Preparation of lysis buffer

[0171] Prepare 200 mL of lysis buffer (TFA:TIS:MPS:H2O:DODT = 93%:2%:1%:2%:2% (v:v)), cool it to <0℃ in an ice bath, add it to a 500 mL beaker, and start stirring.

[0172] (2) Pyrolysis reaction

[0173] Under stirring conditions, 14.0 g of linear peptide resin was added to the above lysis solution, and the temperature of the lysis solution was controlled to be <10℃ during the resin addition process. Then the beaker was transferred to a water bath for heating and stirring. When the temperature of the reaction solution rose to 25℃, the reaction was started and the reaction was carried out for 3.0 h.

[0174] (3) Post-reaction treatment

[0175] After the pyrolysis reaction, the solution was filtered into a vacuum filtration flask, and the resin was washed twice with TFA, 50 mL each time. The filtrates were then combined.

[0176] Add the mixture to 2000 mL of pre-cooled MTBE at 2-8°C while stirring, and then let it stand for 30 minutes.

[0177] Centrifuge the sediment and collect the solid. Wash the solid three times with MTBE, each time using 1200L. Collect the solid and heat it in a vacuum drying oven (vacuum degree ≤ -0.08MPa, temperature 25℃) until constant weight to obtain 4.0g of crude peptide 29.

[0178] 3. Purification and freeze-drying

[0179] Crude peptide 29 was obtained by the above process. Sample preparation: Crude peptide 29 was dissolved in 0.1% TFA aqueous solution and acetonitrile at a ratio of 1:1, and then diluted with 0.1% TFA water at a ratio of 1:1 for purification.

[0180] Step 1 purification: Using crude peptide 29 solution as sample, a C18 reverse chromatographic column as stationary phase, 0.1% TFA water as aqueous phase, and acetonitrile as organic phase, gradient elution was performed, and the target peak was collected to obtain the first-step purified fraction of peptide 29.

[0181] The second purification step: Using the purified fraction from the first step as the sample, gradient elution was performed using a C18 reverse chromatographic column as the stationary phase, ammonium acetate aqueous solution as the aqueous phase, and acetonitrile as the organic phase. The target peak was collected, the collected fraction was concentrated, and the concentrate was lyophilized to obtain 406 mg of pure product.

[0182] Other polypeptides and their extended derivatives disclosed in this invention can be prepared by referring to the above method, and will not be elaborated upon here. The polypeptides and their derivatives of this invention were detected using conventional mass spectrometry methods, and the results are shown in Table 2.

[0183] Table 2 Mass Spectrometry Results of Peptides and Their Derivatives

[0184] Example 3 In vitro activity detection

[0185] 1. Assay of the activity of the test compound against the GLP-1R / GCGR receptor

[0186] (1) Cell Information

[0187] (2) Main reagents and consumables

[0188] (3) Culture medium preparation

[0189] ①Resuscitation medium: 10% FBS + 90% DMEM.

[0190] ②GCGR cell passage medium: 10% FBS + 90% DMEM + 200 μg / mL Hygromycin B + 2 μg / mL L Prouromycin.

[0191] ③GLP-1R cell passage medium: 10% FBS + 90% DMEM + 400 μg / mL Geneticin.

[0192] ④ Analytical medium: 0.5% FBS + 99.5% DMEM.

[0193] (4) Assay of the activity of the test compound against the GLP-1R / GCGR receptor

[0194] ① Digest GCGR / CRE-Luc / HEK293 and GLP-1R / CRE-Luc / HEK293 cells, resuspend them in analytical medium, and adjust the cell density to 0.3 × 10⁻⁶ cells / year.6 Cells / mL were then seeded into 96-well cell culture plates with an inoculation volume of 75 μL per well, and cultured overnight at 37°C with 5% CO2.

[0195] ② Dilute the compounds (the active polypeptides and their derivatives obtained in Example 2, endogenous control GLP-1(7-37), GCG) with analytical medium.

[0196] ③ Add 75 μL of the compound to each well and continue culturing at 37°C in 5% CO2 for 5 h.

[0197] ④ Remove the 96-well cell plate, equilibrate to room temperature for 20 minutes, and add 50 μL / well Bright-Glo LuciferaseAssay System.

[0198] ⑤ Place the 96-well plate into the microplate reader, shake at 510 rpm for 2 minutes, then read the plate. Select the Luminescence mode and detect the fluorescence intensity of each well.

[0199] ⑥ By plotting a four-parameter curve with concentration on the x-axis and fluorescence intensity on the y-axis, the EC50 value can be obtained.

[0200] 2. Determination of the activity of the test compound against the MasR receptor

[0201] (1) Preparation of KH solution and high potassium solution

[0202] Krebs-Henseleit (KH) solution (mM): NaCl 120, KCl 4.5, CaCl2 1.5, KH2PO4 1.2, MgSO4 1.2, NaHCO3 15, glucose 10, adjust pH to 7.4.

[0203] K+-Krebs-Henseleit (K+-KH) solution (mM): NaCl 60, KCl 63.5, CaCl2 1.5, KH2PO4 1.2, MgSO4 1.2, NaHCO3 15, glucose 10, adjust pH to 7.4.

[0204] (2) Preparation of vascular ring samples: After anesthetizing rats, the mesenteric artery was quickly removed and immersed in pre-cooled KH solution (filled with a mixture of 95% air and 5% CO2, pH=7.4). Under a stereomicroscope, the perivascular adhesions were carefully dissected and cut into 2mm segments for later use. The arterial vascular ring was hung on two L-shaped metal needles, one connected to a tension transducer and the other to a vascular fine-tuning device (to adjust the load tension). The vascular tension was recorded by the Taimeng BL-420N biosignal measurement system. The arterial vascular ring was immersed in a 1mL constant-temperature tissue bath containing KH solution, maintained at 37℃, and continuously purged with a mixture of 95% air and 5% CO2.

[0205] (3) Vascular function recording: Before the experiment, the vascular ring was pre-tensioned with 0.65g and equilibrated for about 90 minutes, during which fresh KH solution was replaced every 15 minutes. After equilibration, the arterial ring contractility was tested by constricting the blood vessels with high K+-KH solution (containing 60mM K+). Two contraction amplitudes with a difference of <10% were selected for the experiment.

[0206] (4) Preparation of vasoconcentration gradient vasodilation curve: After the blood vessels are in equilibrium, add 80 μL of vasoconstrictor phenylephrine (PE, purchased from Supelco, batch number: P2110532) to an 8 mL constant temperature tissue bath. After the blood vessels vasoconstrict and enter the plateau phase, add Ach or the drug to be tested sequentially from low to high concentration to plot the concentration gradient vasodilation curve.

[0207] Table 3 Target Activity Detection Results

[0208] The above experimental results show that peptides 25, 29, and 31 exhibit superior activity at the three targets of GCGR, GLP-1R, and MasR. Specifically, peptides 25 and 29 showed comparable activity to the positive control Glucagon at the GCG target, while peptide 31 showed slightly better activity than Glucagon. At the GLP-1R target, peptide 31 showed approximately 3.48 times greater activity than the positive control GLP-1(7-37), peptide 29 showed comparable activity to GLP-1(7-37), and peptide 25 showed slightly weaker activity at the GLP-1R target than GLP-1(7-37). For the MasR target, peptides 29 and 31 showed slightly better vasodilation rates (%) than the positive control Ang1-7, while peptide 25 showed comparable vasodilation rates at MasR to Ang1-7.

[0209] Example 4: Therapeutic effect of subcutaneous injection of long-acting polypeptide derivatives on dbdb diabetic mice

[0210] Methods: Eighty-four male BKS-DB mice were acclimatized and their blood glucose levels were measured randomly. The successfully modeling mice were randomly divided into 14 groups based on blood glucose levels: a model control group (Vehicle), a positive control group (Semaglutide), groups receiving peptide doses of 257, 14, 28, and 56 nmol / kg, groups receiving peptide doses of 2917.5, 35, 70, and 140 nmol / kg, and groups receiving peptide doses of 3117.5, 35, 70, and 140 nmol / kg, with six mice in each group. A blank control group (Control) of 10 mice was also included. Subcutaneous administration was performed twice weekly (Tuesday and Friday) for six consecutive weeks, with random and fasting blood glucose levels measured weekly. At the experimental endpoint, blood samples were collected for glycated hemoglobin and lipid levels, etc., using a Mindray BS-240VET biochemical analyzer. Glycated hemoglobin was measured using EDTA-anticoagulated plasma, and lipid levels were measured using serum.

[0211] Conclusion: Subcutaneous injection of the series of polypeptide derivatives of this invention has a significant hypoglycemic effect on dbdb diabetic mice. While reducing blood glucose, it can also reduce blood lipids.

[0212] The results are as follows:

[0213] 1. Effects on fasting blood glucose

[0214] After 6 weeks of administration, compared with the Control group, the Vehicle group showed a significant increase in fasting blood glucose. Compared with the Vehicle group, peptide 2556 nmol / kg significantly reduced fasting blood glucose (P<0.05), without dose dependence. Peptides 2970 and 140 nmol / kg significantly reduced fasting blood glucose in a dose-dependent manner (P<0.05), and peptide 31 significantly reduced fasting blood glucose in a dose-dependent manner (P<0.05, P<0.01). Peptides 29 and 31 at similar doses had a similar or better fasting blood glucose-lowering effect than semaglutide. The results are shown in Figure 1.

[0215] 2. Effects on random blood glucose

[0216] After 6 weeks of drug administration, compared with the Control group, the Vehicle group showed a significant increase in random blood glucose. Compared with the Vehicle group, peptide 2556 nmol / kg significantly reduced random blood glucose (P<0.01, P<0.001); peptide 2970 nmol / kg significantly reduced random blood glucose (P<0.05, P<0.01, P<0.001); and peptide 31 showed a dose-dependent significant reduction in random blood glucose (P<0.05, P<0.01, P<0.001). The results are shown in Figure 2.

[0217] 3. Effects on glycated hemoglobin

[0218] After 6 weeks of administration, HbA1c was significantly increased in the Vehicle group compared to the Control group. Compared to the Vehicle group, peptide 25 significantly reduced HbA1c in a dose-dependent manner (P<0.01), and peptides 29, 17.5, 70, and 140 nmol / kg significantly reduced HbA1c (P<0.05, P<0.01). Peptide 31 had comparable effects to the semaglutide group. The results are shown in Table 4 (mean ± standard deviation) and Figure 3 (mean ± standard error; the reason for less than 6 n in the figure is mortality during the 6-week administration period).

[0219] Table 4 Effects on Glycated Hemoglobin Note: — indicates that all animals died at the end of the experiment, and no test data was available.

[0220] 4. Effects on blood lipids

[0221] After 6 weeks of drug administration, compared with the control group, TG and LDL were significantly increased in the vehicle group; compared with the vehicle group, TG was significantly decreased in all drug administration groups, and LDL-C was significantly decreased in the 2556 nmol / kg peptide group, but the difference was not statistically significant. TC and HDL-C showed no significant changes in this model. The results are shown in Figure 4.

[0222] Example 5: Therapeutic effect of subcutaneous injection of peptide 31 on diet-induced obese mice

[0223] Methods: Sixty male DIO mice were fed a high-fat diet and acclimatized for 7 days. Mice were then randomly divided into six groups according to body weight: a model control group (Vehicle), a positive control group (Tirzepatide 30 nmol / kg) (Hangzhou Gutuo Biotechnology Co., Ltd., batch number: P220627-L007), a positive control group (LY3437943 10 nmol / kg) (Hangzhou Gutuo Biotechnology Co., Ltd., batch number: GTB10976-1219), and three dose groups of polypeptide 31 (3, 10, and 30 nmol / kg, corresponding to 0.015 mg / kg, 0.05 mg / kg, and 0.15 mg / kg, respectively), with 10 mice in each group. A blank control group of 10 mice was also included. The test drug and the positive control group were administered the drug subcutaneously, while the model control group and the blank control group were administered physiological saline subcutaneously. The drugs were administered twice a week, on Tuesdays and Fridays, for four consecutive weeks. Samples were collected after each week of administration. Body weight and food intake were recorded daily, and fasting and random blood glucose levels were measured weekly. After the last administration, blood samples were collected from 9 animals in each test group for pharmacokinetic analysis. Blood was collected from the jugular vein of 3 animals in each group at each time point, with EDTA-Na2 added as anticoagulant, centrifuged at 3000 rpm for 5 min, and plasma was collected. Blood collection time points were 0.5, 2, 4, 8, 12, 24, 48, 72, 96, 120, 144, and 168 hours after administration. The content of polypeptide 31 in mouse pharmacokinetic plasma samples was determined using LC-MS. At the endpoint, tissue samples were dissected, blood and serum were collected, and adipose tissue was harvested for assay.

[0224] Conclusion: Under the experimental conditions, subcutaneous injection of peptide 31 (3, 10, and 30 nmol / kg) significantly reduced the weight of DIO mice. At 30 nmol / kg, its weight reduction was comparable to or slightly stronger than that of LY3437943, and superior to Tirzepatide. Under the same weight-loss conditions, peptide 31 reduced fat mass, adipose tissue index, Lee's index, and fat loss effects comparable to LY3437943 and Tirzepatide, all of which restored the fat to normal levels. Its effect on appetite was comparable. Simultaneously, it significantly reduced glycated hemoglobin, with effects superior to or comparable to LY3437943 and Tirzepatide. Pharmacokinetic results showed that the half-life (T1 / 2) of peptide 31 ranged from 12.2 to 33.4 hours, reaching maximum plasma concentration 4 hours after injection. The minimum effective dose for peptide 31's weight-loss activity was 10 nmol / kg (0.05 mg / kg), which reduced body weight by 22.12% compared to the model group.

[0225] result:

[0226] 1. Effects on body weight:

[0227] After 4 weeks of administration, the vehicle group showed a significant increase in body weight compared to the control group (P<0.001); compared to the vehicle group, peptide 31 significantly reduced body weight in a dose-dependent manner, with animals receiving 30 nmol / kg of peptide 31 returning to normal body weight. Results are shown in Table 5 (mean ± standard deviation) and Figure 5.

[0228] Compared with the Vehicle group, by week 4 of administration, the weight loss rates of the peptide 3110 and 30 nmol / kg groups reached 26.45% and 47.56%, respectively; the weight loss rate of LY3437943 was 42.33%; and the weight loss rate of Tirzepatide was 38.63%. At 30 nmol / kg, the weight loss was comparable to or slightly stronger than that of LY3437943, and superior to that of Tirzepatide. The results are shown in Table 5 (mean ± standard deviation) and Figure 6.

[0229] Compared to before autologous administration (Day 0), by week 4 of administration, the weight loss rates in the peptide 3110 and 30 nmol / kg groups reached 22.19% and 44.38%, respectively. The weight loss rate in the LY3437943 group was 38.82%, and the weight loss rate in the Tirzepatide group was 34.96%. The weight loss effect of the peptide at 3130 nmol / kg was comparable to or slightly stronger than that of the LY3437943 group, and superior to that of the Tirzepatide group. The results are shown in Table 5 (mean ± standard deviation) and Figure 7.

[0230] Table 5. Effects on body weight (g) ▲ Two died; ▲▲ : One died (Note:) *** P < 0.001 vs. Control; # P < 0.05 ## P < 0.01, ### P < 0.001 vs. Vehicle

[0231] 2. Effects on body fat (abdominal fat) in DIO mice

[0232] Compared with the Control group, the Vehicle group showed significantly increased fat mass (P<0.001), adipose tissue index (P<0.001), and Lee's index (P<0.001). Compared with the Vehicle group, peptide 31 significantly reduced fat mass, adipose tissue index, and Lee's index in a dose-dependent manner. To achieve the same weight loss effect, peptide 31's effect in reducing fat mass, adipose tissue index, and Lee's index was comparable to LY3437943 and superior to Tirzepatide, restoring all to normal levels. Results are shown in Table 6 (mean ± standard deviation) and Figure 8 (mean ± standard error).

[0233] Table 6 Effects on fat changes ▲ Two died; ▲▲ 1 dead *** P<0.001 vs. Control; # P<0.05, ## P<0.01, ### P<0.001, vs. Vehicle

[0234] 3. Effects on fat content in DIO mice:

[0235] At the 4-week endpoint of drug administration, compared with the Control group, the Vehicle group showed a significant increase in fat (P<0.01). Compared with the Vehicle group, peptide 31 significantly reduced fat percentage in a dose-dependent manner, which was statistically significant at 30 nmol / kg (P<0.05). While achieving the same weight loss effect, its fat-reducing effect was comparable to that of LY3437943 and Tirzepatide, both of which restored the fat percentage of model mice to normal levels. The results are shown in Figure 9 (mean ± standard error).

[0236] 4. Effects on food intake

[0237] Under the condition of achieving the same weight loss effect, the effect of peptide 31 in inhibiting food intake was comparable to that of Tirzepatide and LY3437943. Compared with the Vehicle group, the food intake variation range of peptide 31 was -98.66% to 113.54%, that of LY3437943 was -98.66% to 22.73%, and that of Tirzepatide was -98.21% to 56.25%. The results are shown in Table 7 (mean ± standard deviation) and Figure 10.

[0238] Table 7. Effects on feed intake (g / d / animal) ▲ Two died.▲▲ 1 dead

[0239] 5. Effects on blood sugar

[0240] Compared with Vehicle, peptide 31 significantly reduced fasting blood glucose in a dose-dependent manner. The effect of peptide 31 at 30 nmol / kg was comparable to that of Tirzepatide. Compared with LY3437943, the glucose-lowering effect was more gradual, suggesting a lower risk of hypoglycemia. The results are shown in Table 8 (mean ± standard deviation) and Figure 11.

[0241] Compared with Vehicle, peptide 31 significantly reduced random blood glucose, and peptide 31 at 30 nmol / kg was more effective than Tirzepatide and LY3437943. The results are shown in Table 9 (mean ± standard deviation) and Figure 12.

[0242] Table 8. Effects on fasting blood glucose (mmol / L) * Two died; # 1 dead *** P<0.001 vs. Control; # P<0.05, ## P<0.01, ### P<0.001, vs. Vehicle

[0243] Table 9. Effects on random blood glucose (mmol / L) ▲ Two died; ▲▲ 1 dead *** P<0.001 vs. Control; # P<0.05, ## P<0.01, ### P<0.001, vs. Vehicle

[0244] 6. Effects on glycated hemoglobin

[0245] After 4 weeks of administration, all doses of peptide 31 reduced glycated hemoglobin, with statistically significant reductions at 10 nmol / kg and 30 nmol / kg (P<0.05, P<0.05). The reduction rates at 10 and 30 nmol / kg reached 9.46% and 8.88%, respectively, which was superior to Tirzepatide (reduction of 4.14%) and comparable to LY3437943 (reduction of 9.03%). The results are shown in Table 10 (mean ± standard deviation) and Figure 13 (mean ± standard error).

[0246] Table 10 Effects on Glycated Hemoglobin ▲ Two died; ▲▲ 1 dead *** P<0.001 vs. Control; # P<0.05, ## P<0.01, ### P<0.001, vs. Vehicle

[0247] 7. Effects on blood lipids

[0248] At the 4-week endpoint of drug administration, compared with the control group, serum TC and LDL levels were significantly increased in the Vehicle group (P<0.001); compared with the Vehicle group, peptide 31 significantly reduced TC, TG, and LDL levels in a dose-dependent manner (P<0.05, P<0.01, P<0.001), and its effect in reducing TC, TG, and LDL was comparable to or better than that of LY3437943 and Tirzepatide. The results are shown in Table 11 (mean ± standard deviation) and Figure 14 (mean ± standard error).

[0249] Table 11 Effects on blood lipid changes (mmol / L) ▲ Two died; ▲▲ 1 dead *** P<0.001 vs. Control; # P<0.05, ## P<0.01, ### P<0.001, vs. Vehicle

[0250] 8. Pharmacokinetics

[0251] The half-life (T1 / 2) of peptide 31 ranges from 12.2 to 33.4 hours. Maximum plasma concentration is reached 4 hours after subcutaneous injection. Exposure data from Cmax and AUClast indicate that the exposure level is lower than the dose-proportional level, approximately 0.6 times the dose-proportional level. (See Table 12 and Figure 15).

[0252] Table 12 Effects on pharmacokinetics

[0253] The foregoing has provided a detailed description of the three-target peptides and their derivatives provided by this invention. Specific examples have been used to illustrate the principles and implementation methods of this invention. The descriptions of the above embodiments are merely for the purpose of helping to understand the method and core ideas of this invention. It should be noted that those skilled in the art can make various improvements and modifications to this invention without departing from its principles, and these improvements and modifications also fall within the protection scope of the claims of this invention.

Claims

1. A three-target polypeptide or a pharmaceutically acceptable salt, solvate, hydrate, complex, chelate, derivative, variant, or analog thereof, characterized in that, The structure of the polypeptide is shown in Formula I: H-X2-Q-G-T-F-T-S-D-Y-S-K-Y-L-D-X 16 -X 17 -A-A-X 20 -E-F-X 23 -X 24 -W-L-X 27 -X 28 -X 29 -G-P-S-S-G-A-P-P-P-S-D-R-V-Y-I-H-P Formula I in: X2 is selected from G, Aib, or dS; X 16 Selected from E or Aib; X 17 Selected from Q, K or X 20 Selected from K or X 23 Selected from I or V; X 24 Choose from A or E; X 27 Selected from M or L; X 28 Selected from N, E, D, or S; X 29 Selected from T or G; dS stands for D-SER; This represents the Lys side chain -NH2 linked to the fatty acid chain; Preferably, the Lys side chain -NH2 is connected to the fatty acid chain via a connector; Preferably, the acyl group on the connector forms an amide bond with the Lys side chain -NH2, and the fatty acid chain HOOC-(CH2) n -CO- forms an amide bond with the amino group on the connector, and the structure of the connector is shown in Formula II: Wherein, n is 12-20, preferably 14-18, and more preferably 16; Where m is 0-4, preferably 1-3, and more preferably 2; Preferably, the fatty acid chain comprises C18 diacid-Gamma-GLU-2OEG-.

2. The polypeptide of claim 1 or a pharmaceutically acceptable salt, solvate, hydrate, complex, chelate, derivative, variant, or analogue thereof, characterized in that, The polypeptide has the following characteristics: (I) An amino acid sequence as shown in any one of SEQ ID No. 1 to 31; (II) An amino acid sequence that is functionally identical to the amino acid sequence described in (I) obtained by substituting, deleting, or adding one, two, or three amino acids to the amino acid sequence described in (I); or (III) An amino acid sequence having 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% or more homology with the amino acid sequence described in (I) or (II).

3. The polypeptide or its pharmaceutically acceptable salt, solvate, hydrate, complex, chelate, derivative, variant, or analogue as described in claim 1 or 2, characterized in that, The peptide chain ends of the polypeptide are either free or chemically modified; Preferably, the chemical modification includes chemical modification at the amino terminus or chemical modification at the carboxyl terminus; Preferably, the chemical modification of the amino terminus includes acylation, sulfonation, alkylation, and PEG modification; the chemical modification of the carboxyl terminus includes amidation, sulfonamideation, and PEG modification. Preferably, the chemical modification of the amino terminus is acetylation, benzoylation, or sulfonation of the amino group; the alkylation of the amino terminus is C1-6 alkylation or aralkylation; and the chemical modification of the carboxyl terminus is that the OH group in the carboxyl group is replaced by NH2, or the carboxyl group is replaced by sulfonamide, or the OH group in the carboxyl group is connected to a functionalized PEG molecule.

4. A drug, characterized in that, The drug comprises at least one of the following: a polypeptide as described in any one of claims 1-3, a pharmaceutically acceptable salt, solvate, hydrate, complex, chelate, derivative, variant, or analogue thereof, and a pharmaceutically acceptable excipient, excipient, carrier, and solvent; Preferably, it may also contain other active ingredients.

5. The use of the polypeptide or a pharmaceutically acceptable salt, solvate, hydrate, complex, chelate, derivative, variant or analog thereof as described in any one of claims 1 to 3 in the preparation of a medicament for lowering blood sugar, lowering blood lipids and / or losing weight, or a medicament for preventing, treating and / or improving diabetes, obesity, obesity-related diseases or metabolic disorders.

6. A method for lowering blood sugar, lowering blood lipids, and / or losing weight, or a method for preventing, treating, and / or improving diabetes, obesity, obesity-related diseases, or metabolic abnormalities, characterized in that, Administer, take or use any of the following: (I) The polypeptide as described in any one of claims 1-3, or a pharmaceutically acceptable salt, solvate, hydrate, complex, chelate, derivative, variant, or analogue thereof; or (II) The drug as described in claim 4.

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