WO2018113340A1 - 多肽p11及其用途 - Google Patents

多肽p11及其用途 Download PDF

Info

Publication number
WO2018113340A1
WO2018113340A1 PCT/CN2017/100154 CN2017100154W WO2018113340A1 WO 2018113340 A1 WO2018113340 A1 WO 2018113340A1 CN 2017100154 W CN2017100154 W CN 2017100154W WO 2018113340 A1 WO2018113340 A1 WO 2018113340A1
Authority
WO
WIPO (PCT)
Prior art keywords
polypeptide
group
glp
diabetes
pharmaceutical composition
Prior art date
Application number
PCT/CN2017/100154
Other languages
English (en)
French (fr)
Inventor
金亮
高华山
赵茜
张艳峰
潘怡
邢芸
申育萌
Original Assignee
中国药科大学
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Priority claimed from CN201611222521.7A external-priority patent/CN106519016A/zh
Priority claimed from CN201710494709.5A external-priority patent/CN107281471A/zh
Application filed by 中国药科大学 filed Critical 中国药科大学
Publication of WO2018113340A1 publication Critical patent/WO2018113340A1/zh

Links

Images

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/22—Hormones
    • A61K38/26—Glucagons
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P3/00—Drugs for disorders of the metabolism
    • A61P3/08—Drugs for disorders of the metabolism for glucose homeostasis
    • A61P3/10—Drugs for disorders of the metabolism for glucose homeostasis for hyperglycaemia, e.g. antidiabetics
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/575—Hormones
    • C07K14/605—Glucagons

Definitions

  • the invention relates to a polypeptide and a use thereof, in particular to a polypeptide P11, the use thereof comprises: the use for preparing a medicament for preventing or treating diabetes, and the use for preparing a slimming medicament, and belongs to the technical field of biochemical pharmacy.
  • diabetes is a metabolic disorder characterized by chronic hyperglycemia. It is currently believed that genetic and environmental interactions are the main causes, but the pathogenesis is not fully understood. Diabetes can be divided into 4 types: type 1 diabetes, type 2 diabetes, gestational diabetes and other types of diabetes.
  • Type 1 diabetes mellitus T1DM is an autoimmune disease. The loss of islet ⁇ cells is caused by the loss of tolerance to autoantibodies. The body cannot produce enough insulin, and insulin is absolutely absent, causing blood sugar to rise. Therefore, patients should be injected with insulin for life. According to data released by the International Diabetes Federation (IDF) in 2009, there are currently approximately 30 million patients with type 1 diabetes worldwide.
  • type 1 diabetes such as type 1 diabetes vaccine, islet transplantation, gene immunotherapy, and deciphering of disease-causing genes, providing new ideas and methods for the treatment of type 1 diabetes.
  • the main method for treating type 1 diabetes is insulin injection, but the injection of insulin is likely to cause side effects such as hypoglycemia.
  • Type 2 diabetes mellitus also known as non-insulin-dependent diabetes, is due to low beta cell function, relative insulin deficiency and insulin resistance.
  • the prevalence of type 2 diabetes has gradually increased. According to the World Health Organization, there will be 300 million patients with type 2 diabetes worldwide in 2030.
  • the main treatments for type 2 diabetes are oral hypoglycemic agents and insulin. Although these drugs are effective in lowering blood sugar and improving patient symptoms, they still suffer from defects such as hypoglycemia and weight gain. In addition, with the prolongation of the course of diabetes, these drugs can not prevent the deterioration of ⁇ cells in patients.
  • GLP-1 glucagon-like peptide-1
  • Incretin incretin
  • GLP-1 has two biologically active forms, GLP-1 (7-37) and GLP-1 (7-36) amide, which are mainly present in the blood circulation in the active form of GLP-1 (7-36) amide.
  • GLP-1 (7-37) GLP-1 (7-36) amide.
  • GLP-1 Natural human GLP-1 is poorly stable, easily degraded by dipeptidyl peptidase IV (DPPIV), and rapidly cleared by the kidneys, with a half-life of t1/2 ⁇ 2 min. GLP-1 cleaves the N-terminal His-Ala by DPPIV to generate GLP-1 (9-36) amino polypeptide.
  • the initial study concluded that the GLP-1 (9-36) amino polypeptide has no biological activity and is GLP-1.
  • the receptor (GLP-1R) also has some antagonism.
  • the GLP-1 (9-36) amino polypeptide has been found to have an insulin-like action.
  • GLP-1 The physiological role of GLP-1 mainly includes three parts. One is to exert physiological effects by binding to GLP-1R, including: 1 glucose-dependent insulin secretion; 2 inhibition of glucagon secretion; 3 delayed gastric emptying, lowering Appetite, reduce diet, control body weight; 4 inhibit ⁇ cell apoptosis, promote ⁇ cell proliferation and differentiation. Second, GLP-1 is degraded by DPPIV to produce GLP-1 (9-36) amino polypeptide, which can inhibit the expression of hepatic gluconeogenesis and fatty acid synthase, inhibit gluconeogenesis and liver fat synthesis, and play insulin-like action.
  • GLP-1(9-36) amino polypeptide is cleaved into GLP-1(32-36) amino 5 peptide by endopeptidase (NEP24.11), which increases energy expenditure and inhibits weight gain in obese mice. .
  • GLP-1 or GLP-1 analogues can help reduce blood sugar in type 1 diabetes, reduce the incidence of hypoglycemia, reduce glycated hemoglobin, reduce the amount of insulin, but there are side effects of weight loss.
  • orlistat as a gastrointestinal lipase inhibitor, partially inhibits gastric lipase, pancreatic lipase and carboxyl ester lipase, hindering the absorption of fatty acids and glyceryl monostearate by gastrointestinal mucosal cells, thereby Reduce the absorption of fat by 30%, increase the excretion of feces, and achieve the purpose of weight loss.
  • Orly's adverse reactions are mainly manifested by adverse reactions to the gastrointestinal tract, such as diarrhea, flatulence, urgency, and urinary incontinence. These adverse reactions need to be attenuated by lowering the fat content of the diet.
  • Lorcaserin increases appetite and increases satiety by selectively activating the 5-HT2C receptor in the hypothalamus.
  • Phentermine and topiramate (qsymia) sustained release capsules are marketed as novel compound weight loss drugs for appetite suppression.
  • Anthraquinone has an appetite-inhibiting activity in naltrexone hydrochloride and bupropion hydrochloride compound sustained-release tablets (contrave), which stimulates hypothalamic opioid-melanocyte cortisol (POMC) release neurons (Pro-cortisol) is related.
  • POMC hypothalamic opioid-melanocyte cortisol
  • ⁇ -melanocyte stimulating hormone ⁇ -MSH
  • ⁇ -MSH ⁇ -melanocyte stimulating hormone
  • Liraglutide is a human glucagon-like peptide-1 (GLP-1) analog
  • GLP-1 is a physiological regulator of appetite and caloric intake
  • GLP-1 receptor is present in Several brain regions involved in appetite regulation.
  • Liraglutide retains the biological activity of GLP-1, but its half-life is significantly longer than that of native GLP-1. Liraglutide reduces body weight by reducing calorie intake but does not increase energy consumption by 24 hours.
  • Liraglutide Serious adverse reactions reported during the treatment with liraglutide include pancreatitis, gallbladder disease, renal dysfunction, and suicidal tendencies. Liraglutide also speeds up heart rate, and should stop taking the drug if the patient's resting heart rate continues to increase. These weight-loss drugs are expensive, and in addition to orlistat, the other four diet pills will suppress appetite, and orlistat also needs to reduce its adverse reactions by reducing the fat content of the diet, so research new diet pills Still urgency.
  • the inventors developed the polypeptide P11 in the study of diabetes, and further found in the subsequent studies that the polypeptide P11 also has a function of treating obesity.
  • the technical problem to be solved by the present invention is to provide a polypeptide, that is, a polypeptide P11, which can treat diabetes and obesity, and further provides various uses of the polypeptide.
  • HGEGTFTSDVSSYLEGQAAKEFIAWLVKGRGP HGEGTFTSDVSSYLEGQAAKEFIAWLVKGRGP.
  • the pharmaceutical composition comprises the polypeptide itself or a pharmaceutically acceptable salt thereof.
  • the pharmaceutical composition comprises a pharmaceutical carrier and/or a pharmaceutically active substance.
  • GLP-1 HAEGTFTSDVSSYLEGQAAKEFIAWLVKGRG
  • the present invention constructs a polypeptide P11 by amino acid substitution and addition of one amino acid at the end.
  • P11 peptide can effectively exert glucose-dependent insulin secretion (IPTGG), reduce blood glucose in STZ diabetic mice, inhibit feeding in STZ diabetic mice, and control weight loss in STZ diabetic mice.
  • the islet morphology increased the islet area (immuno staining, and cumulative area) of STZ diabetic mice, and increased the C-peptide level of STZ diabetic mice, which was significantly better than Essex in reducing blood sugar and controlling body weight.
  • Peptide (Exendin-4) and GLP-1 are examples of Peptide (Exendin-4) and GLP-1.
  • polypeptide P11 has been experimentally confirmed to be effective for obesity, which not only develops new uses for peptide P11, but also finds new therapeutic agents for obesity.
  • Fig. 1 is a graph showing the results of sugar-dependent insulinotropic secretion (IPTGG) of polypeptide P11 in Example 1 of the present invention.
  • Figure 2 is a graph showing the effect of polypeptide P11 on cumulative eating in STZ diabetic model mice in Example 2 of the present invention.
  • Fig. 3 is a graph showing the effect of polypeptide P11 on blood glucose of STZ diabetic model mice in Example 2 of the present invention.
  • Figure 4 is a graph showing the effect of polypeptide P11 on the body weight of STZ diabetic model mice in Example 2 of the present invention.
  • Figure 5 is a graph showing the effect of polypeptide P11 on triglyceride and free fatty acids in STZ diabetic model mice in Example 2 of the present invention.
  • Fig. 6 is a graph showing the effect of polypeptide P11 on islet morphology (HE staining) of STZ diabetic model mice in Example 2 of the present invention.
  • Figure 7 is a graph showing the effect of polypeptide P11 on islet area (immunostaining and cumulative area) of STZ diabetic model mice in Example 2 of the present invention.
  • Figure 8 is a graph showing the effect of polypeptide P11 on the level of C-peptide of STZ diabetic model mice in Example 2 of the present invention.
  • 9 and 10 are respectively a line diagram and a bar graph of a change in body weight in Example 3 of the present invention.
  • Figure 11 is a schematic illustration of the amount of food consumed in Example 3 of the present invention.
  • Figure 12 is a graph showing the fat content in Example 3 of the present invention.
  • Figure 13 is a bar graph of fat content in Example 3 of the present invention.
  • Vehicle or model represents the solvent control group
  • Progly represents the P11 intervention group
  • Exendin-4 represents the Exendin-4 intervention group. * indicates the significance compared with the solvent control group, *P ⁇ 0.05, **P ⁇ 0.01, *** P ⁇ 0.001.
  • c57 mice Forty male 8-year-old c57 mice (purchased from the Center for Comparative Medical Sciences, Yangzhou University, license number: SCXK (Su) 2012-0004). They were randomly divided into 4 groups, 10 in each group, which were solvent control group (negative control group), Exendin-4 group (positive control group), GLP-1 group (positive control group), and P11 group (experimental group).
  • the peritoneal glucose tolerance process was as follows: c57 mice were fasted for 12 hours, fasting blood glucose was measured, peptide or solvent was injected, glucose (1.5 g/kg) was injected for 30 minutes, and blood glucose was measured at 15, 30, 60, and 120 minutes after injecting sugar, respectively. And calculate the area under the curve separately.
  • P11 had a glucose-dependent insulin-promoting function, and was significantly different from the solvent control group and the GLP-1 group, and had the same effect as the Exendin-4 group.
  • the dose was 25 nmol/kg twice a day, the solvent was physiological saline, 0.1 ml was injected subcutaneously, and the drug was administered at a fixed time point, AM 9:00-10:00, PM 8:00-9:00.
  • the negative control group was injected subcutaneously with 0.1 ml of physiological saline.
  • mice were fasted for 8 hours before administration, AM was fasted at 8:00, PM 4:00, fasting blood glucose was detected in the tail vein, and a blood glucose meter (OHA-231 type of Omron blood glucose meter) was used for blood glucose administration for 0 weeks. Then, after 1, 2, 3, 4, and 5 weeks of administration, fasting blood glucose was measured, and the fasting time and detection time were the same as 0 weeks. As shown in Fig. 3, the P11 group can significantly reduce the fasting of STZ diabetic model mice. Blood glucose was significantly better than the GLP-1 and Exendin-4 groups.
  • the body weight of the mice was measured while blood glucose was detected and recorded.
  • the body weight before administration was 0 weeks of body weight, and then administered 1, respectively. 2, 3, 4, 5 weeks, the body weight was measured, and the results are shown in Fig. 4.
  • the P11 group significantly inhibited the weight loss of STZ diabetic mice, which was significantly better than the GLP1 and Exendin-4 groups.
  • the P11 group was able to improve the ⁇ body weight change in STZ diabetic model mice.
  • mice Five weeks after the administration, the mice were sacrificed, and the pancreas was taken and fixed with formalin. After HE staining, the islet morphology was observed, and the results are shown in Fig. 6. In the solvent control group, the islet injury was severe, and the intact islet structure was almost invisible. The islet morphology of GLP-1 group, Exendin-4 group and P11 group was significantly improved. It was observed that the cells were lightly colored and arranged in a group-like islet structure. It is indicated that GLP-1 group, Exendin-4 group and P11 group can improve islet morphology.
  • mice Five weeks after the administration, the mice were sacrificed, pancreas was taken, formalin was fixed, immunostaining (insulin, ki67), and pancreatic ⁇ -cell area was calculated, and the results are shown in Fig. 7.
  • Immunostaining images showed that there were few islet ⁇ cells in the solvent control group (STZ injury induced ⁇ -cell death), while the islet ⁇ cells in the GLP-1 group, Exendin-4 group and P11 group increased, and the total islet ⁇ -cell area was calculated. Results There was significant difference between P11 group and solvent control group, p ⁇ 0.01. There was also significant difference between GLP-1 group and Exendin-4 group and solvent control group, p ⁇ 0.05. It indicated that the P11 group could significantly increase the islet ⁇ -cell volume and increase the islet ⁇ -cell area, and the results were better than GLP1 group and Exendin-4 group.
  • 6-week-old C57 mice were purchased from Yangzhou University, fed with high-fat diet, and obese model was established.
  • Mice with a body weight greater than 35 g were randomly divided into three groups: solvent control group, Exentin-4 group, and P11. The group was divided into 12 groups; the drug was administered for 3 weeks, the body weight, the food intake were measured once a week, and the fat content was measured after 3 weeks.
  • the solvent control group was injected with 0.1 ml of normal saline; the Exentin-4 group and the P11 group were injected subcutaneously with 50 nmol/kg/b, respectively.
  • the amount of shot is 0.1ml; once in the morning and evening.
  • Detection index The amount of food intake and body weight were measured once a week, and the body fat content of MRI was measured before sacrifice.
  • Progly not only has a stronger weight loss effect than Exentin-4 (the prototype peptide of liraglutide), but also differs from the published and reported GLP-1 analogues (such as Exentin-4) or its modification.
  • Polypeptides and the like have an appetite suppressing effect, and while reducing body weight and fat content, not only do not suppress appetite but may slightly increase appetite.

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Diabetes (AREA)
  • General Health & Medical Sciences (AREA)
  • Endocrinology (AREA)
  • Organic Chemistry (AREA)
  • Medicinal Chemistry (AREA)
  • Animal Behavior & Ethology (AREA)
  • Veterinary Medicine (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Zoology (AREA)
  • Public Health (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Engineering & Computer Science (AREA)
  • Biochemistry (AREA)
  • Genetics & Genomics (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Toxicology (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Obesity (AREA)
  • Hematology (AREA)
  • Biophysics (AREA)
  • Emergency Medicine (AREA)
  • Molecular Biology (AREA)
  • General Chemical & Material Sciences (AREA)
  • Immunology (AREA)
  • Epidemiology (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
  • Peptides Or Proteins (AREA)

Abstract

提供一种多肽P11及其用途。该多肽的氨基酸序列为：HGEGTFTSDVSSYLEGQAAKEFIAWLVKGRGP。该多肽的用途为：用于制备治疗或预防糖尿病药物或药物组合物的用途，用于制备减肥药物或药物组合物的用途。所述多肽能够有效地发挥糖依赖性促胰岛素分泌作用；针对STZ糖尿病模型小鼠，能降低其血糖，抑制其进食，控制体重减轻，使其保持良好的胰岛形态，增加其胰岛面积，增加其C肽水平，且降糖、控制体重作用明显优于艾塞那肽(Exendin-4)和GLP-1；并对肥胖症具有疗效。

Description

多肽P11及其用途 技术领域
本发明涉及一种多肽及其用途，具体而言涉及多肽P11，其用途包括：用于制备预防或治疗糖尿病药物的用途、以及用于制备减肥药物的用途，属于生化药学技术领域。
背景技术
据发明人了解，糖尿病是一种以慢性高血糖为特征的代谢紊乱综合征，目前认为遗传和环境的相互作用是主要诱因，但是其发病机理尚未完全明确。糖尿病可分为4型：1型糖尿病、2型糖尿病、妊娠期糖尿病和其他类型糖尿病。1型糖尿病(Type 1 diabetes mellitus,T1DM)是一种自身免疫性疾病，由于对自身抗体耐受性的丧失使胰岛β细胞损伤，机体无法产生足够的胰岛素，胰岛素绝对缺乏，引起血糖升高，故患者一旦发病需终生注射胰岛素。根据国际糖尿病联合会(IDF)于2009年发布的数据，全世界目前约有3000万1型糖尿病患者。近年，针对1型糖尿病的治疗取得了许多新的进展，如1型糖尿病疫苗、胰岛移植、基因免疫治疗以及致病基因的破译等方面的研究为1型糖尿病的治疗提供了新的思路和方法。目前治疗1型糖尿病的主要方法还是注射胰岛素，但胰岛素的注射容易引起低血糖等副作用。2型糖尿病(Type 2 diabetes mellitus,T2DM)，又被称之为非胰岛素依赖糖尿病，缘于β细胞功能低下，胰岛素相对缺乏和胰岛素抵抗。近年来，2型糖尿病的患病率逐渐增加，据世界卫生组织预测，2030年，全球将有3亿2型糖尿病患者。目前，针对2型糖尿病的治疗药物主要是口服降糖药和胰岛素。虽然该类药物能有效降低血糖，并改善患者症状，但仍然存在低血糖和体重增加等缺陷。此外，随着糖尿病病程的延长，该类药物均不能阻止患者β细胞的恶化。
在针对糖尿病的研究成果中，胰高血糖素样肽-1(glucagon-like peptide-1,GLP-1)是由胰高血糖素原基因编码，后经加工修饰得到的30个氨基酸的肠降血糖素(incretin)；主要由小肠黏膜的L细胞合成并分泌，在胰岛的α细胞、部分大脑区神经元也有表达。GLP-1具有两种生物活性形式，GLP-1(7-37)和GLP-1(7-36)酰胺，血液循环中主要以GLP-1(7-36)酰胺的活性形式存在，两者具有共同的生物学活性。
天然人GLP-1的稳定性较差，易被二肽基肽酶Ⅳ(DPPⅣ)降解，并快速被肾脏清除，其半衰期t1/2≤2min。GLP-1经DPPⅣ切割掉N末端His-Ala，生成GLP-1(9-36)氨基多肽，最初研究认为，GLP-1(9-36)氨基多肽不具有生物学活性，而且对GLP-1受体(GLP-1R)还具有一定的拮抗作用，近些年发现GLP-1(9-36)氨基多肽具有类胰岛素的作用。
GLP-1的生理作用主要包括三个部分，一是通过与GLP-1R结合，发挥生理学效应，包括：①葡萄糖依赖性促胰岛素分泌；②抑制胰高血糖素分泌；③延迟胃排空，降低食欲，减少饮食,控制体重；④抑制β细胞凋亡，促进β细胞增值和分化。二是，GLP-1经过DPPⅣ降解，生成GLP-1(9-36)氨基多肽，能够抑制肝脏糖异生酶和脂肪酸合酶的表达，抑制糖异生和肝脏脂肪合成，发挥类胰岛素作用。三是GLP-1(9-36)氨基多肽经过肽链内切酶(NEP24.11)切割成GLP-1(32-36)氨基5肽，在肥胖型小鼠上增加能量消耗，抑制体重增长。
研究发现，胰岛素与GLP-1或GLP-1类似物联合用药有助于降低I型糖尿病的血糖，减少低血糖的发生，降低糖化血红蛋白，减少胰岛素的用量，但是存在体重减轻的副作用。
肥胖症在全球范围内已成为一种严重威胁健康的流行病，无论是发达国家还是发展中国家均面临着肥胖症加速发展的困扰，随之带来巨大的医疗费用和社会负担。据统计，目前全球已有15亿超重和5亿肥胖患者。对于这些BMI指数高的人群来说，肥胖会引发其他一些疾病发病率的上升，其中最为显著的是心血管疾病、糖尿病和癌症。
关于肥胖的医学治疗手段主要有外科手术和药物治疗两种，通过减肥手术减少胃的大小、增加饱胀感、减少食物摄取的量，从而达到减肥的效果，但是减肥手术有严重的手术和代谢并发症的风险，而且手术非常昂贵，因此药物治疗是更主要的选择。
目前，有5种减肥药在临床中使用，其中包括1999年批准的奥利司他(orlistat)胶囊、2012年批准的氯卡色林(lorcaserin)和苯丁胺和托吡酯(qsymia)缓释剂胶囊、以及2014年批准的盐酸纳曲酮与盐酸安非他酮复方缓释片(contrave)及利拉鲁肽(liraglutide)注射液。其中，奥利司他(orlistat)作为胃肠道脂酶抑制剂，部分抑制胃脂肪酶、胰脂肪酶和羧基酯脂肪酶，阻碍胃肠道黏膜细胞吸收脂肪酸和单硬脂酸甘油酯，从而减少30％摄入脂肪的吸收，增加粪便的排泄，达到减重的目的。奥利司的不良反应主要表现为对胃肠道不良反应，如腹泻、肠胃气胀、便急和尿失禁，这些不良反应需要通过降低膳食中的脂肪含量来减弱。氯卡色林(lorcaserin)是通过选择性地激活下丘脑中的5-HT2C受体使食欲减退，增加饱腹感。苯丁胺和托吡酯(qsymia)缓释剂胶囊是作为食欲抑制类新型复方减肥药物上市。盐酸纳曲酮与盐酸安非他酮复方缓释片(contrave)中安非他酮具有抑制食欲的活性，这种活性与刺激下丘脑的阿片-促黑素细胞皮质素原(POMC)释放神经元(促皮质素原)相关。这些神经元释放α-黑素细胞刺激素(α-MSH)，减少食物的摄入，同时增加了能量的消耗；在服用安非他酮的同时服用纳曲酮增强了食欲抑制活性，加强了减肥疗效。利拉鲁肽(liraglutide)是一种人胰高血糖素样肽-1(GLP-1)类似物，GLP-1是一种食欲和热量摄取的生理调节剂，而GLP-1受体存在于涉及食欲调节的几个脑区。利拉鲁肽保留了GLP-1的生物活性，但其半衰期较天然GLP-1明显延长。利拉鲁肽是通过减低热量摄入，但并不增加24h能量消耗来降低体重。据报告，利拉鲁肽治疗时的严重不良反应包括胰腺炎、胆囊疾病、肾功能障碍和自杀倾向。利拉鲁肽还会加快心率，若患者的静息心率持续加快则应停止用药。以上这些减肥药物价格昂贵，而且除了奥利司他外，其他四种减肥药都会抑制食欲，而奥利司他也需要通过降低膳食中的脂肪含量来减弱它的不良反应，因此研究新型减肥药仍具有迫切性。
发明人在针对糖尿病的研究中研制出了多肽P11，并在后续研究中进一步发现多肽P11还具有治疗肥胖症的功能。
发明内容
本发明所要解决的技术问题是：针对现有技术存在的问题，提供一种多肽，即多肽P11，能治疗糖尿病和肥胖症；并在此基础上，进一步提供该多肽的多种用途。
本发明的技术方案如下：
一种多肽，其特征是，该多肽的氨基酸序列为：
HGEGTFTSDVSSYLEGQAAKEFIAWLVKGRGP。
上述多肽用于制备治疗或预防1型糖尿病药物或药物组合物的用途。
上述多肽用于制备治疗或预防2型糖尿病药物或药物组合物的用途。
上述多肽用于制备减肥药物或药物组合物的用途。
上述用途中，所述药物组合物包括该多肽自身或其药用盐。
上述用途中，所述药物组合物包括药物载体和/或药物活性物质。
发明人在实践研究中，将GLP-1的野生型进行改造：第2位的氨基酸由Ala替换成Gly，在GLP-1的末尾添加Pro，序列如下：
GLP-1：HAEGTFTSDVSSYLEGQAAKEFIAWLVKGRG
P11：H
Figure PCTCN2017100154-appb-000001
EGTFTSDVSSYLEGQAAKEFIAWLVKGRG
Figure PCTCN2017100154-appb-000002
。
本发明通过氨基酸替换和末尾添加一个氨基酸，构建了一种多肽P11。实验结果证明P11肽能够有效地发挥糖依赖性促胰岛素分泌作用(IPTGG)，降低STZ糖尿病模型小鼠的血糖，抑制STZ糖尿病模型小鼠的进食，控制STZ糖尿病模型小鼠的体重减轻，保持良好的胰岛形态(HE染色)，增加STZ糖尿病模型小鼠的胰岛面积(免疫染色，并累计面积)，增加STZ糖尿病模型小鼠的C肽水平，其降糖、控制体重作用明显优于艾塞那肽(Exendin-4)和GLP-1。
在进一步地实践研究中，发明人为筛选出能用于制备减肥药物的多肽，尝试了若干已研发出的多肽，并发现先前获得的多肽P11符合要求。多肽P11经实验确证对肥胖症具有疗效，这不仅为多肽P11开发出新的用途，也为肥胖症找到了新的治疗药剂。
附图说明
图1为本发明实施例1中多肽P11的糖依赖性的促胰岛素分泌作用(IPTGG)结果图。
图2为本发明实施例2中多肽P11对STZ糖尿病模型小鼠累计进食的影响结果图。
图3为本发明实施例2中多肽P11对STZ糖尿病模型小鼠血糖的影响结果图。
图4为本发明实施例2中多肽P11对STZ糖尿病模型小鼠体重的影响结果图。
图5为本发明实施例2中多肽P11对STZ糖尿病模型小鼠甘油三酯和游离脂肪酸的影响结果图。
图6为本发明实施例2中多肽P11对STZ糖尿病模型小鼠胰岛形态(HE染色)的影响结果图。
图7为本发明实施例2中多肽P11对STZ糖尿病模型小鼠胰岛面积(免疫染色，并累计面积)的影响结果图。
图8为本发明实施例2中多肽P11对STZ糖尿病模型小鼠的C肽水平的影响结果图。
图9、图10分别为本发明实施例3中的体重变化折线图和柱形图。
图11为本发明实施例3中的进食量示意图。
图12为本发明实施例3中的脂肪含量图。
图13为本发明实施例3中的脂肪含量柱形图。
以上各图中，Vehicle或model表示溶剂对照组，Progly表示P11干预组，Exendin-4表示Exendin-4干预组。用*表示与溶剂对照组比较的显著性，*P≤0.05，**P≤0.01，***P≤0.001。
具体实施方式
下面参照附图并结合实施例对本发明作进一步详细描述。但是本发明不限于所给出的例子。
实施例1：多肽P11的糖依赖性的促胰岛素分泌作用(IPTGG)
40只8周龄的雄性c57小鼠(购自扬州大学实比较医学中心，许可证号：SCXK(苏)2012-0004)。随机分为4组，每组10只，分别为溶剂对照组(阴性对照组)、Exendin-4组(阳性对照组)、GLP-1组(阳性对照组)、P11组(实验组)。腹腔糖耐量过程如下：c57小鼠空腹12小时，测空腹血糖，分别注射肽或者溶剂，30分钟注射葡萄糖(1.5g/kg)，之后分别在注射糖之后15、30、60、120分钟检测血糖，并分别计算曲线下面积。结果如图1所示，P11具有葡萄糖依赖性的促胰岛素分泌功能，与溶剂对照组和GLP-1组均具有显著性差异，与Exendin-4组效果一样。
实施例2：多肽P11对STZ糖尿病模型小鼠的药效学评价
1、STZ糖尿病模型小鼠造模，及分组
80只6周龄的雄性c57小鼠(购自扬州大学实比较医学中心，许可证号：SCXK(苏)2012-0004)，适应性饲养1周。实验前空腹12小时，按50mg/kg的剂量腹腔注射STZ溶液(pH＝5.2的柠檬酸缓冲液)，连续注射5天，注射后1周、2周分别检测空腹血糖，血糖值均超过11.0mmol/L，为造模成功。将成模小鼠随机分成4组，每组12只，分别为溶剂对照组(阴性对照组)、Exendin-4组(阳性对照组)、GLP-1组(阳性对照组)、P11组(实验组)。给药剂量为25nmol/kg，每天2次，溶剂为生理盐水，皮下注射0.1ml，固定时间点给药，AM 9：00—10:00，PM 8:00—9:00。阴性对照组，皮下注射0.1ml生理盐水。
2、对STZ糖尿病模型小鼠累计进食的影响
从给药开始，每组分别记录初始添食量，隔1天或2天检测剩食量，并继续添加新的鼠粮，并记录，依次类推，到5周给药结束，记录小鼠的总进食量，并绘制累计进食曲线，结果如图2所示，P11组同Exendin-4阳性对照组具有相同的抑制进食的效果，明显少于溶剂组和GLP-1组。
3、对STZ糖尿病模型小鼠血糖的影响
给药前小鼠空腹8小时，AM 8:00开始禁食，PM 4:00，尾静脉检测空腹血糖，检测用血糖检测仪(欧姆龙血糖仪HEA-231型)，作为给药0周血糖。之后分别在给药1、2、3、4、5周，检测空腹血糖，禁食时间和检测时间同0周方法，结果如图3所示，P11组能够显著降低STZ糖尿病模型小鼠的空腹血糖，明显优于GLP-1组和Exendin-4组。
4、对STZ糖尿病模型小鼠体重的影响
检测血糖的同时检测小鼠体重，并记录。给药前的体重为0周体重，之后分别在给药1、 2、3、4、5周，检测体重，结果如图4所示。P11组能够显著抑制STZ糖尿病模型小鼠的体重降低，明显优于GLP1组和Exendin-4组。P11组能够改善STZ糖尿病模型小鼠的Δ体重变化。
5、对STZ糖尿病模型小鼠甘油三酯和游离脂肪酸的影响
给药5周后，眼眶取血，静置收集血清，试剂盒分别检测甘油三酯(TG)和游离脂肪酸(FFA)水平，结果如图5所示。结果显示，血清甘油三酯水平，P11组与溶剂对照组比较，p≤0.001，存在极明显差异；Exendin-4组与溶剂对照组比较存在明显差异，p≤0.01；GLP-1与溶剂对照组比较，无明显差异性；血清游离脂肪酸水平，P11组与溶剂对照组比较存在明显差异，p≤0.01；GLP-1组、Exendin-4组与溶剂对照组比较，无明显差异性。结果说明，多肽P11处理后能够明显降低STZ糖尿病模型小鼠的甘油三酯和游离脂肪酸水平，优于GLP-1组、Exendin-4组。
6、对STZ糖尿病模型小鼠胰岛形态(HE染色)的影响
给药5周后，处死小鼠，取胰腺，用福尔马林固定，经HE染色后，观察胰岛形态，结果如图6所示。溶剂对照组胰岛损伤严重，几乎看不到完整的胰岛结构，GLP-1组、Exendin-4组和P11组胰岛形态得到明显改善，可以观察到细胞着色浅、排列呈团索状的胰岛结构。说明GLP-1组、Exendin-4组和P11组均能改善胰岛形态。
7、对STZ糖尿病模型小鼠胰岛面积(免疫染色，并累计面积)的影响
给药5周后，处死小鼠，取胰腺，福尔马林固定，免疫染色(insulin、ki67)，并计算胰腺β-细胞面积，结果如图7所示。免疫染色图片显示，溶剂对照组胰岛β细胞很少(STZ损伤诱导β-细胞死亡)，而GLP-1组、Exendin-4组和P11组胰岛β细胞增多，经过胰岛β-细胞面积累计计算，结果P11组与溶剂对照组存在明显差异，p≤0.01，GLP-1组、Exendin-4组与溶剂对照组比较，也存明显差异，p≤0.05。说明P11组能够明显增加胰岛β-细胞量，增加胰岛β-细胞面积，并且结果优于GLP1组、Exendin-4组。
8、对STZ糖尿病模型小鼠的C肽水平的影响
给药5周后，眼眶取血，静置收集血清，试剂盒检测C肽水平，结果如图8所示。结果显示，P11组与溶剂对照组存在明显差异，p≤0.05；GLP-1组、Exendin-4组与溶剂对照组比较，无明显差异，说明P11处理后能够明显降低STZ糖尿病模型小鼠的C肽水平，优于GLP-1组、Exendin-4组。
实施例3：多肽P11对肥胖模型小鼠的影响
实验分组：
自扬州大学购入6周龄C57小鼠，用高脂饲料喂养，造肥胖模型，选取体重大于35g的成模小鼠，随机分组，共分成三组，溶剂对照组、Exentin-4组、P11组每组12只；给药3周，每周测量一次体重、进食量、3周后测量脂肪含量。
给药方式：
溶剂对照组注射0.1ml生理盐水；Exentin-4组和P11组分别皮下注射50nmol/kg/只，注 射量0.1ml；早晚各一次。
检测指标：每周检测一次进食量和体重，处死前检测MRI体脂含量。
实验数据
体重：
结果分析：从图9和图10中可以看出Exentin-4和P11肽组体重减少比空白组多，且P11肽组减肥效果比Exentin-4组好。
进食量：
结果分析：如图11所示，Exentin-4组进食量比空白组少，P11组进食量比空白组略多。
脂肪含量：
结果分析：如图12、图13所示，Exentin-4组和P11组脂肪含量与空白组比显著降低，而且P11肽组比Exentin-4组的效果更明显。
由此可知，Progly不仅比Exentin-4(利拉鲁肽的原型肽)具有更强的减肥效果，而且不同于已上市和已报道的GLP-1类似物(如Exentin-4)或其改构多肽等具有抑制食欲功效，其在减少体重和脂肪含量的同时，不仅没有抑制食欲反而可以轻微的增进食欲。
以上实验结果表明，多肽P11对肥胖症具有疗效。
Figure PCTCN2017100154-appb-000003

Claims (6)

  1. 一种多肽，其特征是，该多肽的氨基酸序列为：
    HGEGTFTSDVSSYLEGQAAKEFIAWLVKGRGP。
  2. 权利要求1述及多肽用于制备治疗或预防1型糖尿病药物或药物组合物的用途。
  3. 权利要求1述及多肽用于制备治疗或预防2型糖尿病药物或药物组合物的用途。
  4. 权利要求1述及多肽用于制备减肥药物或药物组合物的用途。
  5. 根据权利要求2或3或4述及的用途，其特征是，所述药物组合物包括该多肽自身或其药用盐。
  6. 根据权利要求2或3或4述及的用途，其特征是，所述药物组合物包括药物载体和/或药物活性物质。
PCT/CN2017/100154 2016-12-20 2017-09-01 多肽p11及其用途 WO2018113340A1 (zh)

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
CN201611222521.7A CN106519016A (zh) 2016-12-20 2016-12-20 降糖调脂肽——Progly肽的设计及其用途
CN201611222521.7 2016-12-20
CN201710494709.5 2017-06-26
CN201710494709.5A CN107281471A (zh) 2017-06-26 2017-06-26 一种改构的胰高血糖素样肽－1及其修饰物的应用

Publications (1)

Publication Number Publication Date
WO2018113340A1 true WO2018113340A1 (zh) 2018-06-28

Family

ID=62624476

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/CN2017/100154 WO2018113340A1 (zh) 2016-12-20 2017-09-01 多肽p11及其用途

Country Status (1)

Country Link
WO (1) WO2018113340A1 (zh)

Citations (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1322137A (zh) * 1998-09-24 2001-11-14 伊莱利利公司 Glp－1或类似物在中风治疗中的用途
CN1746188A (zh) * 2004-09-06 2006-03-15 上海华谊生物技术有限公司 Glp－1类似物
CN101367873A (zh) * 2008-10-08 2009-02-18 南开大学 一种改构的胰高血糖素样肽-1的类似物和修饰物及其应用
US20110236431A1 (en) * 2008-09-12 2011-09-29 Biocompatibles Uk Ltd. Treatment of acute myocardial infarction (ami) using encapsulated cells encoding and secreting glp-1 peptides or analogs thereof
WO2014202727A1 (en) * 2013-06-20 2014-12-24 Novo Nordisk A/S Glp-1 derivatives and uses thereof
WO2015014228A1 (zh) * 2013-08-01 2015-02-05 江苏泰康生物医药有限公司 Glp-1类似物融合蛋白，及其制备方法和用途
WO2016083499A1 (en) * 2014-11-27 2016-06-02 Novo Nordisk A/S Glp-1 derivatives and uses thereof
WO2016120378A1 (en) * 2015-01-29 2016-08-04 Novo Nordisk A/S Tablets comprising glp-1 agonist and enteric coating
CN106519016A (zh) * 2016-12-20 2017-03-22 中国药科大学 降糖调脂肽——Progly肽的设计及其用途

Patent Citations (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1322137A (zh) * 1998-09-24 2001-11-14 伊莱利利公司 Glp－1或类似物在中风治疗中的用途
CN1746188A (zh) * 2004-09-06 2006-03-15 上海华谊生物技术有限公司 Glp－1类似物
US20110236431A1 (en) * 2008-09-12 2011-09-29 Biocompatibles Uk Ltd. Treatment of acute myocardial infarction (ami) using encapsulated cells encoding and secreting glp-1 peptides or analogs thereof
CN101367873A (zh) * 2008-10-08 2009-02-18 南开大学 一种改构的胰高血糖素样肽-1的类似物和修饰物及其应用
WO2014202727A1 (en) * 2013-06-20 2014-12-24 Novo Nordisk A/S Glp-1 derivatives and uses thereof
WO2015014228A1 (zh) * 2013-08-01 2015-02-05 江苏泰康生物医药有限公司 Glp-1类似物融合蛋白，及其制备方法和用途
WO2016083499A1 (en) * 2014-11-27 2016-06-02 Novo Nordisk A/S Glp-1 derivatives and uses thereof
WO2016120378A1 (en) * 2015-01-29 2016-08-04 Novo Nordisk A/S Tablets comprising glp-1 agonist and enteric coating
CN106519016A (zh) * 2016-12-20 2017-03-22 中国药科大学 降糖调脂肽——Progly肽的设计及其用途

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
KJEMS, L.L. ET AL.: "The Influence of GLP-1 on Glucose-Stimulated Insulin Secretion Effects on beta- Cell Sensitivity in Type 2 and Nondiabetic Subjects", DIABETES, vol. 52, 28 February 2003 (2003-02-28), pages 380 - 386, XP055277761 *

Similar Documents

Publication Publication Date Title
AU2015276203B2 (en) Exendin-4 derivatives as selective glucagon receptor agonists
CN110172083B (zh) 具有抗肥胖及抗糖尿病功效的肽及其的用途
CN106279400A (zh) P8降糖肽的设计及其用途
TW201803895A (zh) 新穎之調酸素衍生物及含有該衍生物之用於治療肥胖之醫藥組成物
CN103800301B (zh) 治疗糖尿病的脉冲给药组合物及其制备方法
Gupta et al. Glucagon like peptide-1 and its receptor agonists: Their roles in management of Type 2 diabetes mellitus
CN104159598A (zh) 治疗糖尿病的方法和组合物
Brandt et al. Are peptide conjugates the golden therapy against obesity?
Kounatidis et al. Therapeutic potential of GLP-2 analogs in gastrointestinal disorders: current knowledge, nutritional aspects, and future perspectives
CN111138552A (zh) 一种降脂多肽及其制药应用
CN106519016A (zh) 降糖调脂肽——Progly肽的设计及其用途
Reboredo et al. Oral administration of zein-based nanoparticles reduces glycemia and improves glucose tolerance in rats
Patil et al. Role of insulin in management of type 2 diabetes mellitus
WO2018113340A1 (zh) 多肽p11及其用途
CN112079931B (zh) 长效glp-1类似物多肽及其降糖调脂用途
WO2018045872A1 (zh) 一种多肽及其用途
CN103566354B (zh) 含有glp‑1类似物的衍生物或其可药用盐的药物组合物
Lebovitz Therapeutic options in development for management of diabetes: pharmacologic agents and new technologies
Kinalska et al. The influence of incretin mimetics on cardiovascular risk factors in diabetes
Conlon et al. Recent advances in peptide-based therapy for Type 2 diabetes and obesity
CN113476591B (zh) 一种乳源多肽衍生物在制备糖尿病防治药物、保健品以及食品添加物中的应用
Nikkar et al. Review of the mechanism and function of glucagon-like peptide 1 (GLP-1) and GLP-1 receptor agonists in the improvement of type 2 diabetes
Forst The role of glucagon-like petide-1 receptor agonists in the treatment of type 2 diabetes
Meier The role of incretin-based therapies in the management of type 2 diabetes mellitus: perspectives on the past, present and future
Reboredo-Fuentes et al. Oral administration of zein-based nanoparticles reduces glycemia and improves glucose tolerance in rats

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 17883230

Country of ref document: EP

Kind code of ref document: A1

NENP Non-entry into the national phase

Ref country code: DE

122 Ep: pct application non-entry in european phase

Ref document number: 17883230

Country of ref document: EP

Kind code of ref document: A1