WO2012113286A1 - 一种glp-1类似物、制备方法及其应用 - Google Patents

一种glp-1类似物、制备方法及其应用 Download PDF

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WO2012113286A1
WO2012113286A1 PCT/CN2012/070958 CN2012070958W WO2012113286A1 WO 2012113286 A1 WO2012113286 A1 WO 2012113286A1 CN 2012070958 W CN2012070958 W CN 2012070958W WO 2012113286 A1 WO2012113286 A1 WO 2012113286A1
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Prior art keywords
seq
haegt
ssyle gqaak
ggggg
lvkgr
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PCT/CN2012/070958
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English (en)
French (fr)
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龚珉
付刚
徐为人
汤立达
任晓文
刘鹏
王玉丽
吴疆
邹美香
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天津药物研究院
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Application filed by 天津药物研究院 filed Critical 天津药物研究院
Priority to EP12749806.1A priority Critical patent/EP2894163A4/en
Priority to US14/405,060 priority patent/US20150274799A1/en
Publication of WO2012113286A1 publication Critical patent/WO2012113286A1/zh

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    • 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
    • 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
    • A61P3/04—Anorexiants; Antiobesity agents
    • 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
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides

Definitions

  • GLP-1 analogue preparation method and application thereof
  • the present invention relates to the field of diabetes-related drugs, and in particular, to a GLP-1 analogue having an in vivo half-life of extended glucagon-like peptide-1 (GLP-1).
  • GLP-1 extended glucagon-like peptide-1
  • the present invention also relates to a process for the preparation of the GLP-1 analogue and its use in the preparation of a medicament for diabetes. Background technique
  • GLP-1 glycopeptide-1, hereinafter referred to as: GLP-1
  • GLP-1 glycopeptide-1, hereinafter referred to as: GLP-1
  • GLP-1 (7-37) OH and GLP-1 (7-36) NH2 (Mojsov S, J Clin Invest. 1987 Feb; 79(2): 616-9). GLP-1 significantly reduces blood sugar after eating, stimulates insulin production, and also has a certain weight loss effect, and does not cause hypoglycemia (Drucker DJ, Diabetes. 1998 Feb; 47(2): 159-69 ). Recent studies have also shown that GLP-1 has pancreatic regeneration (Drucker D J, 2003 Dec; 144(12): 5145-8). In addition, because GLP-1 is a fully humanized polypeptide, it is a safety advantage as a clinical drug. However, the serum half-life of GLP-1 (7-37) is only 3-5 minutes, and multiple injections per day are very inconvenient in clinical use.
  • Fmoc strategy refers to a method of synthesizing a polypeptide by sequentially condensing an amino terminal Fmoc-protected amino acid in the presence of a coupling reagent using a polymer resin as a solid substrate.
  • Fmoc solid phase peptide synthesis a practical approach, 2000, Oxford University Press.
  • One object of the present invention is to provide a short-lived GLP-1 analog in vivo, which requires To be limited in daily injection administration, an analog of GLP-1 with a longer half-life is provided.
  • Another object of the present invention is to provide a use of a GLP-1 analogue for the preparation of a medicament for treating diabetes, and the use of the GLP-1 analogue for the preparation of a medicament for the treatment and/or prevention of obesity.
  • Still another object of the present invention is to provide a pharmaceutical composition
  • a pharmaceutical composition comprising a GLP-1 analogue as an active ingredient, which comprises the above-mentioned GLP-1 analogue.
  • the pharmaceutical composition further comprises one or more pharmaceutically acceptable excipients, and the pharmaceutical composition is preferably an injection, further preferably a lyophilized powder or a solution for injection.
  • the invention provides a GLP-1 analogue having the following general formula A:
  • X 15 is aspartic acid or cysteine
  • X 22 is glycine or cysteine
  • X 3 is valine or cysteine
  • X 1() , Xi5, X 2 2, X 3 .
  • X 33 is cysteine
  • both cysteines contained in the formula A form a disulfide bond.
  • the GLP-1 analogue has the following formula I: 7HAEX 10 T FTSX15V SSYLE X 22 QAAK EFIX 30 W LX 33 KGR G 37 niXiC n 2 X 2 Formula I,
  • the GLP-1 analog of the formula I is as follows:
  • SEQ ID NO 82 7 HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G 37 GGGGG GGGGG C GGGGG GGGGG;
  • SEQ ID NO 83 7 HAEGT FTSDV SSYLE CQAAK EFIAW LVKGR G 37 GGGGG C GGGGG GGGGG; or
  • SEQ ID NO 84 7 HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G 37 GGG C GGGGG GGGGGGGGGGGGG.
  • the GLP-1 analogue has the following formula:
  • the 11 corpses 5 ⁇ 15, n 2 3 ⁇ 9;
  • the GLP-1 analogue is:
  • SEQ ID NO 2 7 HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G 37 GGGGG GGGGGC GGG;
  • SEQ ID NO 4 7 HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G 37 GGGGG C GGGGG G;
  • SEQ ID NO 7 7 HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G 37 GGGGG GGGGGC GGGGG GGGG;
  • G 37 GGGGG GGGGGGGGGGC GGGGG G;
  • SEQ ID NO 10 7 HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G 37 AAAAA C AAA;
  • SEQ ID NO 12 7 HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G 37 AAAAA AAAAAAAAAAC AAA;
  • SEQ ID NO 18 7 HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G 37 AAAAA AAAAAAAAAAC AAAAA AAAA;
  • GLP-1 analog having the general formula A may further have the following formula III:
  • ⁇ 3 ⁇ 5
  • ⁇ 2 10 ⁇ 20.
  • the GLP-1 analogue is:
  • SEQ ID NO 30 7 HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G 37 GC GGGGG GGGGGGGGGGGGGGGG;
  • SEQ ID NO 32 7 HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G 37 GG C GGGGG GGGGGGGGGG;
  • G 37 GG C GGGGG GGGGGGGGGGGGGGG;
  • SEQ ID NO 34 7 HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G 37 GGGGG C GGGGG GGGGG;
  • SEQ ID NO 36 7 HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G 37 GGGGG C GGGGG GGGGGGGGGGGGG;
  • SEQ ID NO 37 7 HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G 37 AC AAAAA AAAAA;
  • SEQ ID NO 39 7 HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G 37 AC AAAAA AAAAAAAAAAAAAAAAAAAA;
  • SEQ ID NO 40 7 HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G 37 AA C AAAAA AAAAA;
  • SEQ ID NO 41 7 HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G 37 AA C AAAAA AAAAAAAAAA; SEQ ID NO 42: 'HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G 37 AA C AAAAA AAAAAAAAAAAAA;
  • SEQ ID NO 45 7 HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G 37 AAAAA C AAAAA AAAAAAAAAAAAAAA;
  • SEQ ID NO 46 7 HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G 37 VC VVVVV VVVV;
  • the GLP-1 analog having the formula A may further have the following formula IV: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G 37 C n 2 X 2 Formula IV
  • X is glycine, alanine or valine
  • X 2 is glycine, alanine or valine
  • the GLP-1 analogue is:
  • SEQ ID NO 56 7 HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G 37 GC GGGGG GGGGGGGGGG;
  • SEQ ID NO 57 7 HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G 37 GC GGGGG GGGGGGGGGGGGG;
  • SEQ ID NO 59 7 HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G 37 GG C GGGGG GGGGGGGGGG;
  • SEQ ID NO 60 7 HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G 37 GG C GGGGG GGGGGGGGGGGGG;
  • SEQ ID NO 61 7 HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G 37 GGGGG C GGGGG GGGGG;
  • SEQ ID NO 62 7 HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G 37 GGGGG C GGGGG GGGGGGGGGG;
  • G 37 GGGGG C GGGGG GGGGGGGGGGGGGGG;
  • SEQ ID NO 64 7 HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G 37 AC AAAAA AAAAA;
  • SEQ ID NO 65 7 HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G 37 AC AAAAA AAAAAAAAAA;
  • SEQ ID NO 66 7 HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G 37 AC AAAAA AAAAA AAAAAAAAAA;
  • SEQ ID NO 69 7 HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G' AA C AAAAA AAAAAAAAAAAAAAA;
  • SEQ ID NO 70 7 HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G 37 AAAAA C AAAAA AAAAA;
  • SEQ ID NO 71 7 HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G 37 AAAAA C AAAAA AAAAAAAAAA;
  • SEQ ID NO 72 7 HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G 37 AAAAA C AAAAA AAAAAAAAAAAAAAA;
  • the invention provides a method of making the above-described GLP-1 analog, which comprises solid phase polypeptide synthesis of the Fmoc strategy.
  • the present invention provides the use of the above-mentioned GLP-1 analogue for the preparation of a medicament for the treatment of diabetes, obesity and diabetes, obesity-related diseases, and prevention of obesity.
  • the present invention provides a pharmaceutical composition
  • a pharmaceutical composition comprising the GLP-1 analogue according to any one of claims 1-8 and one or more pharmaceutically acceptable excipients;
  • the pharmaceutical composition is an injection, and more preferably a lyophilized powder or a solution for injection.
  • the formula of the GLP-1 analogue is as follows:
  • amino acid formula based on the mutation of the amino acid at position 15, 30 and 33 to cysteine may preferably be:
  • the GLP-1 -containing analog of the present invention may be combined with one or more pharmaceutically acceptable excipients to form a pharmaceutical composition
  • a pharmaceutical composition comprising: a water-soluble filler, a pH adjuster, a stabilizer, water for injection, permeation Pressure regulator and so on.
  • the water-soluble filler adjuvant of the present invention is one or more selected from the group consisting of mannitol, low molecular dextran, sorbitol, polyethylene glycol, glucose, lactose, galactose and the like.
  • the pH adjusting agent is one or one selected from the group consisting of non-volatile acids such as capric acid, phosphoric acid, lactic acid, tartaric acid, hydrochloric acid, and potassium hydroxide, sodium hydroxide or potassium hydroxide or ammonium hydroxide, sodium carbonate. Or physiologically acceptable organic or inorganic acids and bases and salts such as potassium carbonate or ammonium carbonate, sodium hydrogencarbonate or potassium hydrogencarbonate or ammonium hydrogencarbonate.
  • non-volatile acids such as capric acid, phosphoric acid, lactic acid, tartaric acid, hydrochloric acid, and potassium hydroxide, sodium hydroxide or potassium hydroxide or ammonium hydroxide, sodium carbonate.
  • physiologically acceptable organic or inorganic acids and bases and salts such as potassium carbonate or ammonium carbonate, sodium hydrogencarbonate or potassium hydrogencarbonate or ammonium hydrogencarbonate.
  • the stabilizer is one or one selected from the group consisting of: EDTA-2Na, sodium thiosulfate, sodium metabisulfite, sodium sulfite, dipotassium hydrogen phosphate, sodium hydrogencarbonate, sodium carbonate, arginine, glutamic acid, polyethylene Alcohol 6000, polyethylene glycol 4000, sodium lauryl sulfate or trihydroxydecylaminodecane.
  • Preferred are sodium metabisulfite, dipotassium hydrogen phosphate, arginine, polyethylene glycol 6000, and trishydroxyalkylaminodecane.
  • the osmotic pressure adjusting agent is one or a combination of two of sodium chloride and potassium chloride.
  • the pharmaceutical composition of the present invention can be administered by intramuscular, intravenous or subcutaneous injection, and the preferred dosage form is a lyophilized powder or a solution for injection.
  • Preparation method of freeze-dried injection Take appropriate amount of GLP-1 analogue solution, add water-soluble filler, stabilizer, osmotic pressure regulator, etc., add appropriate amount of water for injection, adjust pH to 4-8 to dissolve, dilute with water to At the appropriate concentration, 0.1-0.5% activated carbon is added, stirred at 0-10 °C for 10-20 minutes, decarburized, filtered by microporous membrane filtration, and the filtrate is packed, and freeze-dried to obtain white loose block. Extract, sealed, each specification contains GLP-1 analogues in 5 g to 1 mg between.
  • Preparation method of injection Take GLP-1 analogue solution or lyophilized powder, add water-soluble filler, stabilizer, osmotic pressure regulator, etc., add appropriate amount of water for injection, adjust pH to 4-8 to dissolve, Dilute to the appropriate concentration with water, add 0.1-0.5% activated carbon, stir at 0 - 10 °C for 10-20 minutes, decarburize, filter and sterilize by microporous membrane filtration, the filtrate is divided and sealed, and each specification is obtained. Contains a GLP-1 analog between 5 g and 1 mg.
  • the GLP-1 analogue of the present invention can be used in the preparation of a therapeutic drug for diabetes.
  • the composition of the present invention can be administered in the form of intravenous, intramuscular or subcutaneous injection. While the dosage varies depending on the subject to be treated, the mode of administration, the symptoms and other factors, the GLP-1 analogues of the invention are effective over a relatively wide range of dosages. In the treatment of adults, the dose ranges from 5 g/person to 1 mg/person, once a day or once every few days.
  • the actual dose should be determined by the doctor according to the relevant circumstances, including the physical condition of the subject, the route of administration, age, weight, individual response of the patient to the drug, the severity of the patient's symptoms, etc., so the above dose range is The scope of the invention is not limited in any way.
  • the GLP-1 analog of the present invention overcomes the problem of short half-life of GLP-1, and the half-life of the GLP-1 analog provided in the body can reach 15 to 75 hours or more, compared with the half-life (half-life) of GLP-1 administered alone. Only 3-5 minutes) significantly extended, greatly facilitating its clinical promotion and application.
  • Figure 1 is a hypoglycemic test containing GLP-1 analogue (SEQ ID NO: 1-27) in Example 2; wherein, Figure 1-A shows the hypoglycemic test of GLP-1 analogue (SEQ ID NO: -9) , wherein the ten data bars corresponding to each time point represent SEQ1-9 and the control from left to right; Figure 1-B shows the hypoglycemic test of the GLP-1 analogue (SEQ ID NO: 10-18), wherein Ten data bars at each time point represent SEQ10-18 and control from left to right; Figure 1-C shows hypoglycemic experiments of GLP-1 analogs (SEQ ID NO 19-27), corresponding to each The ten data bars at the time point represent SEQ 19-27 and the control from left to right;
  • Figure 2 is a hypoglycemic assay containing the GLP-1 analogue (SEQ ID NO 28-54) in Example 3; wherein, Figure 2-A shows the hypoglycemic assay of the GLP-1 analog (SEQ ID NO 28-36) , wherein the ten data bars corresponding to each time point represent SEQ28-36 and the control from left to right; Figure 2-B shows the hypoglycemic test of the GLP-1 analog (SEQ ID NO 37-45), wherein Ten data bars at each time point represent SEQ37-45 and control from left to right; Figure 2-C shows hypoglycemic experiments of GLP-1 analogs (SEQ ID NO 46-54), corresponding to each The ten data bars at the time point represent SEQ 46-54 and the control from left to right;
  • Figure 3 is a hypoglycemic assay containing the GLP-1 analogue (SEQ ID NO 55-81) in Example 4; wherein, Figure 3-A shows the hypoglycemic assay of the GLP-1 analog (SEQ ID NO 55-64) Wherein eleven data bars corresponding to each time point represent SEQ 55-64 and control from left to right; Figure 3-B shows hypoglycemic experiments of GLP-1 analogs (SEQ ID NO 65-74), wherein The eleven data bars corresponding to each time point represent SEQ 65-74 and the control from left to right; Figure 3-C shows the hypoglycemic test of the GLP-1 analog (SEQ ID NO 75-81), which corresponds to The eight data bars at each time point represent SEQ 75-81 and the control from left to right;
  • Figure 4 is a hypoglycemic assay containing GLP-1 analogues (SEQ ID NO 82-84) in Example 5, wherein four data bars corresponding to each time point represent SEQ 82-84 and control, respectively;
  • Figure 5 is a graph depicting the stability of GLP-1 analogs in human serum in Example 6;
  • Figure 6 is a graph depicting the stability of GLP-1 analogs in human serum as described in Example 7.
  • Figure 7 is a graph depicting the stability of GLP-1 analogs in human serum as described in Example 8. detailed description
  • Example 1 Solid phase synthesis of polypeptide
  • the synthesis of the polypeptide of the present invention was carried out using a solid phase polypeptide synthesis method of the Fmoc strategy using a CS 336X type instrument manufactured by CSBio. The method of synthesis is carried out according to the manufacturer's instructions.
  • the prepared polypeptide was purified using a HPLC C18 semi-preparative column, and the mobile phase was acetonitrile.
  • the polypeptide lyophilized powder is obtained by desalting and freeze-drying.
  • the polypeptides encompassed by this patent all contain disulfide bonds, ammonium bicarbonate or other reducing agents used to form disulfide bonds in the polypeptide.
  • Example 2 Hypoglycemic function associated with GLP-1 analogues (Formula II)
  • polypeptides used are as follows: SEQ ID NO 1: 'HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G J GGGGG C GGG
  • mice 200 ⁇ /only, 6/group, purchased from Shanghai Experimental Animal Center of Chinese Academy of Sciences
  • 400 micrograms of glucose was then injected into each mouse. 2 hours after glucose injection,
  • mice The blood glucose of the mice was measured at 24 hours, 48 hours, 72 hours, and 96 hours (Note: the same dose of glucose was again given two hours before each blood glucose measurement), and the results are shown in Fig. 1, in which the control (blank) in this example In order to subcutaneously inject saline into mice, the amount of glucose was the same as in the other test groups.
  • Example 3 Hypoglycemic function associated with GLP-1 analogues (Formula III).
  • polypeptides used are as follows: SEQ ID NO 28: 'HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G' GC GGGGG GGGGG
  • G 37 GGGGG C GGGGG GGGGGGGGGGGG
  • G 37 GGGGG C GGGGGGGGGGGGGGGGGGG
  • SEQ ID NO 38 7 HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G 37 AC AAAAA AAAAAAAAAA
  • AAAAA C AAAAA AAAAA SEQ ID NO 44: 'HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G 37 AAAAA C AAAAA AAAAAAAAAA
  • the polypeptide was dissolved in 1 ml of physiological saline to prepare a polypeptide solution, and the polypeptide solution was subcutaneously injected into mice (200 ⁇ l/only, 6/group, purchased from Shanghai Experimental Animal Center of Chinese Academy of Sciences), 30 minutes after administration. 400 micrograms of glucose was injected into each mouse. 2 hours after glucose injection,
  • mice The blood glucose of the mice was measured at 24 hours, 48 hours, 72 hours, and 96 hours (Note: the same dose of glucose was given again two hours before each blood glucose measurement), and the results are shown in Fig. 2, in which the control in this example (blank) In order to subcutaneously inject saline into mice, the amount of glucose was the same as in the other test groups.
  • Example 4 Hypoglycemic function associated with GLP-1 analogues (Formula IV)
  • polypeptides used are as follows: SEQ ID NO 55: 'HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G' GC GGGGG GGGGG
  • SEQ ID NO 58 7 HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G 37 GG C GGGGG GGGGG
  • G 37 GGGGG C GGGGG GGGGGGGGGGGG
  • G 37 GGGGG C GGGGGGGGGGGGGGGGGGG
  • SEQ ID NO 68 7 HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G 37 AA C AAAAA AAAAAAAAAA
  • AAAAA C AAAAA AAAAA SEQ ID NO 71: 'HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G 37 AAAAA C AAAAA AAAAAAAAAA
  • the polypeptide was dissolved in 1 ml of physiological saline to prepare a polypeptide solution, and the polypeptide solution was subcutaneously injected into mice (200 ⁇ l/only, 6/group, purchased from Shanghai Experimental Animal Center of Chinese Academy of Sciences), 30 minutes after administration. 400 micrograms of glucose was injected into each mouse. 2 hours after glucose injection,
  • mice The blood glucose of the mice was measured at 24 hours, 48 hours, 72 hours, and 96 hours (Note: the same dose of glucose was again given two hours before each blood glucose measurement), and the results are shown in Fig. 3, in which the control (blank) in this example In order to subcutaneously inject saline into mice, the amount of glucose was the same as in the other test groups.
  • Example 5 GLP-1 analogue based on mutation of amino acids at other sites to cysteine
  • polypeptides used are as follows: SEQ ID NO 82: 'HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR GGGGG GGGGG C GGGGG GGGGG
  • SEQ ID NO 83 7 HAEGT FTSDV SSYLE CQAAK EFIAW LVKGR G 37 GGGGG C GGGGG GGGGG SEQ ID NO 84: 7 HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G 37 GGG C GGGGG GGGGGGGGGGGGG
  • mice 200 ⁇ /only, 6/group, purchased from Shanghai Experimental Animal Center of Chinese Academy of Sciences
  • 400 mg of glucose was injected per 30 minutes after administration. Only mice.
  • the blood glucose of the mice was measured at 2 hours, 24 hours, 48 hours, 72 hours, and 96 hours after the glucose injection, and the results are shown in Fig. 4, wherein: the control (blank) in this example was physiological saline, and the amount was 1 ml.
  • Example 6 Determination of stability of GLP-1 analogues in human serum
  • polypeptides used are as follows:
  • the above two polypeptides and the GLP-1 standard (purchased from Shanghai Shenggong Co., Ltd., the sequence of which is: HAEGTFTSDVSSYLEGQAAKEFIAWLVKGRG) were dissolved in 0.5 ml of physiological saline, fully dissolved, and added to 1 ml of serum to mark the serum.
  • NO 58 experimental group O.lmg peptide is SEQ ID NO 58 in 0.5 ml physiological saline
  • NO 59 experimental group O The .lmg sequence is the peptide of SEQ ID NO 59 dissolved in 0.5 ml of physiological saline).
  • polypeptides used are as follows:
  • the above polypeptide and GLP-1 standard (purchased from Shanghai Shenggong Company, the sequence of which is: SEQ ID NO 85, HAEGTFTSDVSSYLEGQAAKEFIAWLVKGRG) each dissolved in 0.5 ml of physiological saline, fully dissolved and added to the serum, respectively
  • the serum was labeled as a blank group (O.lmg GLP-1 was dissolved in 0.5 ml of normal saline), N0.3 experimental group (O.lmg of the peptide of SEQ ID NO 3 was dissolved in 0.5 ml of physiological saline) and NO 25 experiment.
  • Group (O.lmg sequence is the peptide of SEQ ID NO 25 dissolved in 0.5 ml of physiological saline).
  • polypeptides used are as follows:
  • SEQ ID NO 28 7 HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G 37 GC GGGGG GGGGG;
  • the above polypeptide and GLP-1 standard (purchased from Shanghai Shenggong Company, the sequence of which is: SEQ ID NO 85, HAEGTFTSDVSSYLEGQAAKEFIAWLVKGRG) each dissolved in 0.5 ml of physiological saline, fully dissolved and added to the serum, respectively Serum was labeled as a blank group (O.lmg GLP-1 dissolved in 0.5 ml normal saline), N0.28 experimental group (O.lmg sequence was SEQ ID NO The peptide of 28 was dissolved in 0.5 ml of physiological saline) and the NO 29 experimental group (O.lmg of the peptide of SEQ ID NO 29 was dissolved in 0.5 ml of physiological saline).

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Description

一种 GLP-1类似物、 制备方法及其应用 技术领域
本发明涉及糖尿病相关的药物领域， 具体而言， 本发明涉及一种具有延 长的胰高血糖素样肽 -1 ( GLP-1 )类的体内半衰期的 GLP-1类似物。 本发明还 涉及该 GLP- 1类似物的制备方法以及其在制备糖尿病药物中的应用。 背景技术
本发明涉及的 GLP-1 ( glucagon-likepeptide-1 , 以下筒称： GLP-1)是主要 由小肠 L细胞分泌的一种 37个氨基酸组成的多肽， 其活性形式为
GLP-l(7-37)OH和 GLP-1(7-36)NH2 (Mojsov S, J Clin Invest. 1987 Feb; 79(2): 616-9)。 GLP-1明显减低人用餐后的血糖， 能刺激胰岛素的产生， 同时还能 起到一定减肥效应，并且不会引起低血糖症（ Drucker D J, Diabetes. 1998 Feb; 47(2): 159-69)。 近期研究还表明 GLP-1有胰腺再生作用 （ Drucker D J, 2003Dec;144(12):5145-8)。 此外， 因为 GLP-1为完全人源化多肽， 作为临床 药物在安全上具有较大优势。 然而， GLP-1 ( 7-37 )的血清半衰期仅仅为 3-5 分钟， 每天多次注射给药在临床使用中非常不方便。
目前已经有不少研究采用 GLP-1类似物融合蛋白技术解决 GLP-1类似 物在体内的存留时间 （CN90101167.3、 CN200710018734.2、
CN200410054397.9、 CN01820232.2、 CN200380110152.7、 CN200510039265.3、 CN200610127237.1、 CN200910009642.7 ) , 然而， 现有的技术与临床理想的 目标还有很大距离，普遍都还没有达到临床标准,最近 Novo Norisk生产的利 拉鲁肽为 GLP-1类似物，其对 GLP-1进行了棕榈酸的修饰，并于 2009年于美 国上市。 但是利拉鲁肽也存在半衰期短的问题， 其剂型仍需每日注射。
因此， 目前存在对解决 GLP-1体内半衰期短的方法的需求。 发明内容
除非另外指出， 本文所述的 "Fmoc策略"是指以聚合物树脂作为固相反 应基质，将氨基端 Fmoc保护的氨基酸在偶联试剂的存在下依次缩合，从而合 成多肽的合成方法。 其具体方法参见 Fmoc solid phase peptide synthesis: a practical approach, 2000, Oxford University Press.
本发明的一个目的是针对临床上 GLP-1类似物在体内存留时间较短， 需 要每天注射给药的缺限， 提供一种半衰期较长的 GLP-1的类似物。
本发明的另一个目的是提供一种 GLP- 1类似物在制备治疗糖尿病的药物 中的应用， 以及该 GLP-1类似物在制备治疗和 /或预防肥胖症的药物中的应 用。
本发明的又一个目的是提供一种以 GLP-1类似物作为有效成分的药用 组合物， 其包含上述 GLP-1类似物。 其中， 所述的药物组合物还包含一种或 多种药学上可接受的辅料， 并且所述药物组合物优选为注射剂， 进一步优选 为冻干粉针或溶液注射剂。
用于实现上述目的的技术方案如下：
一方面， 本发明提供一种 GLP-1类似物， 所述 GLP-1类似物具有下列 通式 A:
7HAEX10T FTSX15V SSYLE X22QAAK EFIX30W LX33KGR G^ ^C n2X2 通式 A,
其中， 。为甘氨酸或半胱氨酸， X15为天冬氨酸或半胱氨酸， X22为甘氨 酸或半胱氨酸， X3。为丙氨酸或半胱氨酸， 33为缬氨酸或半胱氨酸，并且 X1()、 Xi5、 X22、 X3。和 X33中至少有一个为半胱氨酸； iiiXi表示有 个 ηι=1~30, 且 为甘氨酸、 丙氨酸或缬氨酸； n2X2表示有 n2个 X2, n2=0~30, 且 X2为甘氨 酸、 丙氨酸或缬氨酸； 以及通式 A中含有的两个半胱氨酸均形成二硫键。
优选地， 所述的 GLP-1类似物所述 GLP-1类似物具有下列通式 I： 7HAEX10T FTSX15V SSYLE X22QAAK EFIX30W LX33KGR G37niXiC n2X2 通式 I,
其中 1^=3 - 30, =3 ~ 30。
所述具有通式 I的 GLP-1类似物如：
SEQ ID NO 82: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37 GGGGG GGGGG C GGGGG GGGGG;
SEQ ID NO 83: 7HAEGT FTSDV SSYLE CQAAK EFIAW LVKGR G37 GGGGG C GGGGG GGGGG; 或
SEQ ID NO 84: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37 GGG C GGGGG GGGGGGGGGGGGGGG。
优选地， 所述 GLP-1类似物具有下列通式 Π:
7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37niXiC n2X2
通式 Π, 1^=1 - 20, n2=0 ~ 25, 并且其中 X为甘氨酸、 丙氨酸或缬氨酸， X2为甘 氨酸、 丙氨酸或缬氨酸；
优选地， ^=3 ~ 20, n2=3 ~ 25；
进一步优选地， 所述 11尸5 ~ 15, n2=3 ~ 9;
进一步优选地， 所述 GLP-1类似物为：
SEQ ID NO 1： 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37GGGGG C GGG;
SEQ ID NO 2: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37GGGGG GGGGGC GGG;
SEQ ID NO 3: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR
G37GGGGG GGGGGGGGGGC GGG;
SEQ ID NO 4: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37GGGGG C GGGGG G;
SEQ ID NO 5: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37GGGGG C GGGGG GGGG;
SEQ ID NO 6: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37GGGGG GGGGGC GGGGG G;
SEQ ID NO 7: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37GGGGG GGGGGC GGGGG GGGG;
SEQ ID NO 8: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR
G37GGGGG GGGGGGGGGGC GGGGG G;
SEQ ID NO 9: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37GGGGG GGGGGGGGGGC GGGGG GGGG;
SEQ ID NO 10: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37AAAAA C AAA;
SEQ ID NO 11: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37AAAAA AAAAAC AAA;
SEQ ID NO 12: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37AAAAA AAAAAAAAAAC AAA;
SEQ ID NO 13: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR
G37AAAAA C AAAAA A;
SEQ ID NO 14: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G' AAAAA C AAAAA AAAA;
SEQ ID NO 15: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37AAAAA AAAAAC AAAAA A;
SEQ ID NO 16: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37AAAAA AAAAAC AAAAA AAAA;
SEQ ID NO 17: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37AAAAA AAAAAAAAAAC AAAAA A;
SEQ ID NO 18: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37AAAAA AAAAAAAAAAC AAAAA AAAA;
SEQ ID NO 19: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR
G37VVVVV C VVV;
SEQ ID NO 20: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37VVVVV VVVVVC VVV;
SEQ ID NO 21 : 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37VVVVV VVVVVVVVVVC VVV;
SEQ ID NO 22: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37VVVVV C VVVVV V;
SEQ ID NO 23: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37VVVVV C VVVVV WW;
SEQ ID NO 24: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR
G37VVVVV VVVVVC VVVVV V;
SEQ ID NO 25: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37VVVVV VVVVVC VVVVV WW;
SEQ ID NO 26: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37VVVVV VVVVVVVVVVC VVVVV V; 或
SEQ ID NO 27: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37VVVVV VVVVVVVVVVC VVVVV VVVV。
此外， 具有通式 A的 GLP-1类似物还可以具有下列通式 III：
7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37 C n2X2 通式 III,
其中， ηι=1 ~ 10, n2=0 ~ 25 , 为甘氨酸、 丙氨酸或缬氨酸， X2为甘氨 酸、 丙氨酸或缬氨酸； 优选地， Π!=3 - 10, η2=3 ~ 25；
或者 - 5, η2=10 ~ 20;
更优选地， ^=3 ~ 5, η2=10 ~ 20。
优选地， 所述 GLP- 1类似物为：
SEQ ID NO 28: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37G
C GGGGG GGGGG;
SEQ ID NO 29: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37G C GGGGG GGGGGGGGGG;
SEQ ID NO 30: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37G C GGGGG GGGGGGGGGGGGGGGGGGGG；
SEQ ID NO 31: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37GG C GGGGG GGGGG;
SEQ ID NO 32: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37GG C GGGGG GGGGGGGGGG;
SEQ ID NO 33: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR
G37GG C GGGGG GGGGGGGGGGGGGGG;
SEQ ID NO 34: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37GGGGG C GGGGG GGGGG;
SEQ ID NO 35: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37GGGGG C GGGGG GGGGGGGGGG;
SEQ ID NO 36: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37GGGGG C GGGGG GGGGGGGGGGGGGGG;
SEQ ID NO 37: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37A C AAAAA AAAAA;
SEQ ID NO 38: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37A
C AAAAA AAAAAAAAAA;
SEQ ID NO 39: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37A C AAAAA A A A A A A A A A A A A A A A A A A A A；
SEQ ID NO 40: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37AA C AAAAA AAAAA；
SEQ ID NO 41: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37AA C AAAAA AAAAAAAAAA; SEQ ID NO 42: 'HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37AA C AAAAA AAAAAAAAAAAAAAA;
SEQ ID NO 43: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37AAAAA C AAAAA AAAAA;
SEQ ID NO 44: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR
G37AAAAA C AAAAA AAAAAAAAAA;
SEQ ID NO 45: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37AAAAA C AAAAA AAAAAAAAAAAAAAA;
SEQ ID NO 46: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37V C VVVVV VVVVV;
SEQ ID NO 47: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37V C VVVVV VVVVVVVVVV;
SEQ ID NO 48: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37V C VVVVV VVV VVVVVVVVVV VVVVVVV;
SEQ ID NO 49: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR
G37VV C VVVVV VVVVV;
SEQ ID NO 50: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37VV C VVVVV VVVVVVVVVV;
SEQ ID NO 51 : 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37VV C VVVVV VVVVVVVVVVVVVVV;
SEQ ID NO 52: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37VVVVV C VVVVV VVVVV;
SEQ ID NO 53: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37VVVVV C VVVVV VVVVVVVVVV; 或
SEQ ID NO 54: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR
G37VVVVV C VVVVV VVVVVVVVVVVVVVV。
另外， 所述具有通式 A的 GLP-1类似物还可以具有下列通式 IV: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37 C n2X2 通式 IV
其中， 1^=1 - 10, n2=0 ~ 30, X为甘氨酸、 丙氨酸或缬氨酸， X2为甘氨 酸、 丙氨酸或缬氨酸；
优选地 11尸3 ~ 10, n2=3 ~ 30; 或者， 11!=1 - 5, n2=10 ~ 20;
更优选地， 11!=3 ~ 5, n2=10 ~ 20。
优选地， 所述 GLP- 1类似物为：
SEQ ID NO 55: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37G C GGGGG GGGGG;
SEQ ID NO 56: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37G C GGGGG GGGGGGGGGG;
SEQ ID NO 57: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37G C GGGGG GGGGGGGGGGGGGGG；
SEQ ID NO 58: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR
G37GG C GGGGG GGGGG;
SEQ ID NO 59: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37GG C GGGGG GGGGGGGGGG;
SEQ ID NO 60: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37GG C GGGGG GGGGGGGGGGGGGGG;
SEQ ID NO 61: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37GGGGG C GGGGG GGGGG;
SEQ ID NO 62: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37GGGGG C GGGGG GGGGGGGGGG;
SEQ ID NO 63： 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR
G37GGGGG C GGGGG GGGGGGGGGGGGGGG;
SEQ ID NO 64: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37A C AAAAA AAAAA;
SEQ ID NO 65: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37A C AAAAA AAAAAAAAAA；
SEQ ID NO 66: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37A C AAAAA AAAAA AAAAAAAAAA;
SEQ ID NO 67: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37AA C AAAAA AAAAA;
SEQ ID NO 68: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR
G37AA C AAAAA AAAAAAAAAA;
SEQ ID NO 69: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G' AA C AAAAA AAAAAAAAAAAAAAA;
SEQ ID NO 70: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37AAAAA C AAAAA AAAAA;
SEQ ID NO 71: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37AAAAA C AAAAA AAAAAAAAAA;
SEQ ID NO 72: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37AAAAA C AAAAA AAAAAAAAAAAAAAA;
SEQ ID NO 73: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37V C VVVVV VVVVV;
SEQ ID NO 74: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37V
C VVVVV VVVVVVVVVV;
SEQ ID NO 75: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37V C VVVVV VVVVV VVVVVVVVVV;
SEQ ID NO 76: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37VV C VVVVV VVVVV；
SEQ ID NO 77: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37VV C VVVVV VVVVVVVVVV;
SEQ ID NO 78: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37VV C VVVVV VVVVVVVVVVVVVVV;
SEQ ID NO 79: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR
G37VVVVV C VVVVV VVVVV;
SEQ ID NO 80: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37VVVVV C VVVVV VVVVVVVVVV; 或
SEQ ID NO 81: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37VVVV V C VVVVV VVVVVVVVVVVVVVV。 另一方面，本发明提供上述 GLP-1类似物的制备方法，所述制备方法包 括 Fmoc策略的固相多肽合成。
又一方面，本发明提供上述 GLP-1类似物在制备治疗糖尿病，肥胖症和 糖尿病、 肥胖症相关疾病以及预防肥胖症的药物中的应用。
并且， 本发明还提供一种药物组合物， 所述药物组合物包含如权利要求 1-8中任一项所述的 GLP-1类似物和一种或多种药学上可接受的辅料； 优选地， 所述药物组合物为注射剂， 进一步优选为冻干粉针或溶液注射 剂。
以下是本发明的详细描述：
( 1 ) GLP- 1类似物
本发明所述的 GLP- 1类似物的通式如下：
通式 A:
7HAEX10T FTSX15V SSYLE X22QAAK EFIX30W LX33KGR G37niXiC n2X2 7HAEGT FTSDV SSYLE GQAAK EFIAW LVKGR G37为人源 GLP-1序列，上 述通式 A的 GLP- 1类似物为人工序列， 分别在第 10、 15、 22、 30或 33位氨 基酸上进行了修饰， 由半胱氨酸取代了原有的氨基酸。 并且在上述的氨基酸 通式中 可为 Gly, Ala, Val;通式中的 X2可为 Gly, Ala, Val。最后通式中的（ 及 n2 )指代在多肽的 C末端含有！^及 个疏水氨基酸（ Gly, Ala或 Val ) , n^l ~ 30 , n2=0 ~ 30。 在上述的多肽通式中， 两个半胱氨酸均形成二石克键。
优选地， GLP-1类似物的通式如下：
7HAEX10T FTSX15V SSYLE X22QAAK EFIX30W LX33KGR
Figure imgf000010_0001
n2X2 通式 I ,
其中 1^=3 - 30 , =3 ~ 30。
基于 15位、 30位和 33位氨基酸突变为半胱氨酸的氨基酸通式可优选为： 通式 Π:
7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37niXiC n2X2, n^l ~ 20; n2=0 ~ 25。 优选地， 11 3 ~ 20; n2=3 ~ 25。 其中 1为0^, Ala或 Val; X2为 Gly, Ala或 Val。
通式 ΠΙ:
7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37 C n2X2, n^l ~ 10; n2=0 ~ 25。 优选地， 11尸3 ~ 10; n2=3 ~ 25。 其中 1为0^, Ala或 Val; X2为 Gly, Ala或 Val。
通式 IV:
7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37 C n2X2, n^l ~ 10; n2=0 ~ 30。 优选地， 11尸3 ~ 10; n2=3 ~ 30。 其中 1为01^ Ala或 Val; X2为 Gly, Ala或 Val。
上述的多肽通式 II可进一步优化为：
7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37 C n2X2, ^=5 ~ 15; n2=3 ~ 9。 其中 1为0^, Ala或 Val; X2为 Gly, Ala或 Val。
上述的多肽通式 III可优化为：
7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37 C n2X2, n^l ~ 5; n2=10 ~ 20。 优选地， 11尸3 ~ 5; n2=10 ~ 20。 其中 1为0^, Ala或 Val; X2为 Gly, Ala或 Val。
上述的多肽通式 IV可优化为：
7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37 C n2X2, n^l ~ 5; n2=10 ~ 20。 优选地， 11尸3 ~ 5; n2=10 ~ 20。 其中 1为0^, Ala或 Val; X2为 Gly, Ala或 Val。
( 2 )本发明的药用组合物
本发明的含有 GLP-1类似物可以与一种或多种药学上可接受的辅料共 同制成药用组合物， 这些辅料包括： 水溶性填充剂、 PH调节剂、 稳定剂、 注射用水、 渗透压调节剂等等。
本发明所述的水溶性填充剂辅料为选自以下的一种或一种： 甘露醇、 低 分子右旋糖苷、 山梨醇、 聚乙二醇、 葡萄糖、 乳糖、 半乳糖等。
PH调节剂为选自以下的一种或一种： 枸橼酸、 磷酸、 乳酸、 酒石酸、 盐酸等非挥发性的酸以及氢氧化钾、 氢氧化钠或氢氧化钾或氢氧化铵、 碳酸 钠或碳酸钾或碳酸铵盐、碳酸氢钠或碳酸氢钾或碳酸氢铵盐等生理可接受的 有机或无机酸和碱及盐等。
稳定剂为选自以下的一种或一种： EDTA-2Na、 硫代硫酸钠、 焦亚硫酸 钠、 亚硫酸钠、 磷酸氢二钾、 碳酸氢钠、 碳酸钠、 精氨酸、 谷氨酸、 聚乙二 醇 6000、 聚乙二醇 4000、 十二烷基硫酸钠或三羟曱基氨基曱烷等。 优选焦 亚硫酸钠、 磷酸氢二钾、 精氨酸、 聚乙二醇 6000、 三羟曱基氨基曱烷。
渗透压调节剂是氯化钠、 氯化钾的一种或两种的组合。
( 3 )注射剂的制备方法
本发明的药用组合物可以通过肌肉、 静脉内、 皮下注射途经进行给药， 优选的剂型为冻干粉或溶液注射剂。
冷冻干燥注射剂的制备方法： 取 GLP-1类似物溶液适量，加入水溶性填 充剂、 稳定剂、 渗透压调节剂等， 加入注射用水适量， 调节 PH值至 4-8使 其溶解，加水稀释至适当浓度，加入 0.1-0.5%活性炭，在 0 - 10°C下搅拌 10-20 分钟， 脱碳， 采用微孔滤膜过滤除菌， 滤液进行分装， 采用冷冻干燥法， 制 得白色疏松块状物， 封口即得， 每个规格含有 GLP-1类似物在 5 g到 lmg 之间。
注射液的制备方法： 取 GLP-1类似物溶液或冻干粉适量，加入水溶性填 充剂、 稳定剂、 渗透压调节剂等， 加入注射用水适量， 调节 PH值至 4-8使 其溶解，加水稀释至适当浓度，加入 0.1-0.5%活性炭，在 0 - 10°C下搅拌 10-20 分钟， 脱碳， 采用微孔滤膜过滤除菌， 滤液进行分装， 封口即得， 每个规格 含有 GLP-1类似物在 5 g到 lmg之间。
( 4 )药用组合物的用途
本发明的 GLP-1类似物可以用在制备糖尿病治疗药物方面。具体地，本 发明的组合物可以静脉、肌肉或皮下注射剂形式给药。虽然剂量依治疗对象、 给药方式、症状及其它因素而改变，本发明的 GLP-1类似物在相当宽的剂量 范围内是有效的。 在成人的治疗中， 剂量范围在 5 g/人 -lmg/人， 每日一次 或每几日一次给药。 实际剂量应该由医生根据有关的情况来决定， 这些情况 包括被治疗者的身体状态、给药途径、年龄、体重、 患者对药物的个体反应， 患者症状的严重程度等等， 因此上述剂量范围并不是以任何方式限制本发明 的范围。
本发明的 GLP-1类似物克服了 GLP-1半衰期短的问题，所提供的 GLP-1 类似物在体内的半衰期可达到 15 ~ 75小时以上， 较单独给药的 GLP-1的半 衰期（半衰期仅为 3-5分钟） 明显延长， 极大地便利了其临床推广和应用。 附图说明
以下， 结合附图来详细说明本发明的实施例， 其中：
图 1为实施例 2中含 GLP-1类似物（ SEQ ID NO 1—27 )的降血糖实验； 其中， 图 1-A表示 GLP-1类似物 （SEQ ID N0 1— 9 ) 的降血糖实验， 其中 对应于每个时间点的十个数据条从左到右依次代表 SEQ1-9及对照； 图 1-B 表示 GLP- 1类似物 （ SEQ ID NO 10—18 ) 的降血糖实验， 其中对应于每个 时间点的十个数据条从左到右依次代表 SEQ10-18及对照；图 1-C表示 GLP-1 类似物（ SEQ ID NO 19—27 )的降血糖实验， 其中对应于每个时间点的十个 数据条从左到右依次代表 SEQ 19-27及对照；
图 2为实施例 3中含 GLP-1类似物 （ SEQ ID NO 28—54 ) 的降血糖实 验； 其中， 图 2-A表示 GLP-1类似物（ SEQ ID NO 28—36 )的降血糖实验， 其中对应于每个时间点的十个数据条从左到右依次代表 SEQ28-36及对照； 图 2-B表示 GLP-1类似物 （ SEQ ID NO 37—45 ) 的降血糖实验， 其中对应 于每个时间点的十个数据条从左到右依次代表 SEQ37-45及对照； 图 2-C表 示 GLP-1类似物 （ SEQ ID NO 46—54 ) 的降血糖实验， 其中对应于每个时 间点的十个数据条从左到右依次代表 SEQ46-54及对照；
图 3为实施例 4中含 GLP-1类似物 （ SEQ ID NO 55—81 ) 的降血糖实 验； 其中， 图 3-A表示 GLP-1类似物（ SEQ ID NO 55—64 )的降血糖实验， 其中对应于每个时间点的十一个数据条从左到右依次代表 SEQ55-64及对 照； 图 3-B表示 GLP-1类似物 （ SEQ ID NO 65—74 ) 的降血糖实验， 其中 对应于每个时间点的十一个数据条从左到右依次代表 SEQ65-74及对照； 图 3-C表示 GLP-1类似物（ SEQ ID NO 75—81 )的降血糖实验， 其中对应于每 个时间点的八个数据条从左到右依次代表 SEQ75-81及对照；
图 4为实施例 5中含 GLP-1类似物 （ SEQ ID NO 82—84 ) 的降血糖实 验， 其中对应于每个时间点的四个数据条依次代表 SEQ82-84及对照；
图 5为实施例 6中描述了 GLP-1类似物在人血清中的稳定性的图； 图 6为实施例 7中描述了 GLP-1类似物在人血清中的稳定性的图。
图 7为实施例 8中描述了 GLP-1类似物在人血清中的稳定性的图。 具体实施方式
下面结合具体实施方式对本发明进行进一步的详细描述， 给出的实施 例仅为了阐明本发明， 而不是为了限制本发明的范围。
在以下的实施例中， 未详细描述的各种过程和方法是本领域中公知的 常规方法。 实施例 1：多肽的固相合成
使用 Fmoc策略的固相多肽合成方法，使用 CSBio公司生产的 CS 336X 型仪器进行本发明的多肽的合成。 合成的方法按照生产商的仪器说明书进 行。
将制得的多肽使用 HPLC C18半制备柱进行纯化， 流动相为乙腈。 经 脱盐及冷冻干燥得到多肽冻干粉。 本专利包含的多肽均含有二硫键， 碳酸 氢铵或其他还原剂用来形成多肽中的二硫键。 实施例 2: GLP-1类似物（通式 II )相关的降血糖功能
本实施例中， 采用的多肽如下： SEQ ID NO 1: 'HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR GJ GGGGG C GGG
SEQ ID NO 2: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37GGGGG GGGGGC GGG SEQ ID NO 3: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37GGGGG GGGGGGGGGGC GGG
SEQ ID NO 4: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37GGGGG C GGGGG G
SEQ ID NO 5: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37GGGGG C GGGGG GGGG
SEQ ID NO 6: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37GGGGG GGGGGC GGGGG G
SEQ ID NO 7: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37GGGGG GGGGGC GGGGG GGGG SEQ ID NO 8: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37GGGGG GGGGGGGGGGC GGGGG G
SEQ ID NO 9: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37GGGGG GGGGGGGGGGC GGGGG GGGG
SEQ ID NO 10: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR
G37AAAAA C AAA
SEQ ID NO 11: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR
G37AAAAA AAAAAC AAA
SEQ ID NO 12: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR
G37AAAAA AAAAAAAAAAC AAA
SEQ ID NO 13: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR
G37AAAAA C AAAAA A
SEQ ID NO 14: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR
G37AAAAA C AAAAA AAAA
SEQ ID NO 15: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR
G37AAAAA AAAAAC AAAAA A
SEQ ID NO 16: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR
G37AAAAA AAAAAC AAAAA AAAA SEQ ID NO 17: 'HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37AAAAA AAAAAAAAAAC AAAAA A
SEQ ID NO 18: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR
G37AAAAA AAAAAAAAAAC AAAAA AAAA
SEQ ID NO 19: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR
G37VVVVV C VVV
SEQ ID NO 20: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR
G37VVVVV VVVVVC VVV
SEQ ID NO 21: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR
G37VVVVV VVVVVVVVVVC VVV
SEQ ID NO 22: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR
G37VVVVV C VVVVV V
SEQ ID NO 23: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR
G37VVVVV C VVVVV WW
SEQ ID NO 24: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR
G37VVVVV VVVVVC VVVVV V
SEQ ID NO 25: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR
G37VVVVV VVVVVC VVVVV WW
SEQ ID NO 26: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR
G37VVVVV VVVVVVVVVVC VVVVV V
SEQ ID NO 27: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR
G37VVVVV VVVVVVVVVVC VVVVV WW
将上述多肽各 1 mg溶于 1 ml生理盐水制成多肽溶液， 将此多肽溶液皮 下注射于小鼠（200 μΐ/只， 6只 /组， 购自中科院上海实验动物中心） ， 给药 30分钟后将 400微克葡萄糖注射于每只小鼠。 分别于葡萄糖注射后 2小时、
24小时、 48小时、 72小时及 96小时测定小鼠的血糖（注： 每次测血糖前两 个小时再次给相同剂量的葡萄糖），结果见图 1 ,其中本实施例中的对照（空 白） 为向小鼠皮下注射生理盐水， 用量 lml, 葡萄糖的注射与其他试验组相 同。 实施例 3: GLP-1类似物（通式 III )相关的降血糖功能。
本实施例中， 采用的多肽如下： SEQ ID NO 28: 'HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G' G C GGGGG GGGGG
SEQ ID NO 29: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37G C GGGGG GGGGGGGGGG SEQ ID NO 30: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37G C GGGGG GGGGGGGGGGGGGGGGGGGG
SEQ ID NO 31: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37GG C GGGGG GGGGG
SEQ ID NO 32: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37GG C GGGGG GGGGGGGGGG
SEQ ID NO 33: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37GG C GGGGG GGGGGGGGGGGGGGG
SEQ ID NO 34: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR
G37GGGGG C GGGGG GGGGG
SEQ ID NO 35: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR
G37GGGGG C GGGGG GGGGGGGGGG
SEQ ID NO 36: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR
G37GGGGG C GGGGG GGGGGGGGGGGGGGG
SEQ ID NO 37: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37A C AAAAA AAAAA
SEQ ID NO 38: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37A C AAAAA AAAAAAAAAA
SEQ ID NO 39: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37A C AAAAA AAAAAAAAAAAAAAAAAAAA SEQ ID NO 40: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37AA C AAAAA AAAAA
SEQ ID NO 41: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37AA C AAAAA AAAAAAAAAA
SEQ ID NO 42: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37AA C AAAAA AAAAAAAAAAAAAAA
SEQ ID NO 43: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR
G37AAAAA C AAAAA AAAAA SEQ ID NO 44: 'HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37AAAAA C AAAAA AAAAAAAAAA
SEQ ID NO 45: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR
G37AAAAA C AAAAA AAAAAAAAAAAAAAA
SEQ ID NO 46: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37V C vvvvv vvvvv
SEQ ID NO 47: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37V C VVVVV VVVVVVVVVV
SEQ ID NO 48: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37V C VVVVV VVVVVV VVVVVVVVVV vvvv
SEQ ID NO 49: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37VV C VVVVV VVVVV
SEQ ID NO 50: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37VV C VVVVV VVVVVVVVVV SEQ ID NO 51: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37VV C
VVVVV vvvvvvvvvvvvvvv
SEQ ID NO 52: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR
G37VVVVV C VVVVV VVVVV
SEQ ID NO 53: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR
G37VVVVV C VVVVV VVVVVVVVVV
SEQ ID NO 54: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR
G37VVVVV C VVVVV VVVVVVVVVVVVVVV
将上述多肽各 1 mg溶于 1 ml生理盐水制成多肽溶液， 将此多肽溶液皮 下注射于小鼠 （200μ1/只， 6只 /组， 购自中科院上海实验动物中心） ， 给药 30分钟后将 400微克葡萄糖注射于每只小鼠。分别于葡萄糖注射后 2小时、
24小时、 48小时、 72小时及 96小时测定小鼠的血糖（注： 每次测血糖前两 个小时再次给相同剂量的葡萄糖），结果见图 2,其中本实施例中的对照（空 白） 为向小鼠皮下注射生理盐水， 用量 lml, 葡萄糖的注射与其他试验组相 同。 实施例 4: GLP-1类似物（通式 IV )相关的降血糖功能
本实施例中， 采用的多肽如下： SEQ ID NO 55: 'HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G' G C GGGGG GGGGG
SEQ ID NO 56: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37G C GGGGG GGGGGGGGGG SEQ ID NO 57: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37G C GGGGG GGGGGGGGGGGGGGG
SEQ ID NO 58: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37GG C GGGGG GGGGG
SEQ ID NO 59: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37GG C GGGGG GGGGGGGGGG
SEQ ID NO 60: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37GG C GGGGG GGGGGGGGGGGGGGG
SEQ ID NO 61: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR
G37GGGGG C GGGGG GGGGG
SEQ ID NO 62: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR
G37GGGGG C GGGGG GGGGGGGGGG
SEQ ID NO 63: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR
G37GGGGG C GGGGG GGGGGGGGGGGGGGG
SEQ ID NO 64: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37A C AAAAA AAAAA
SEQ ID NO 65: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37A C AAAAA AAAAAAAAAA
SEQ ID NO 66: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37A C AAAAA AAAAAAAAAAAAAAA SEQ ID NO 67: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37AA C AAAAA AAAAA
SEQ ID NO 68: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37AA C AAAAA AAAAAAAAAA
SEQ ID NO 69: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37AA C AAAAA AAAAAAAAAAAAAAA
SEQ ID NO 70: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR
G37AAAAA C AAAAA AAAAA SEQ ID NO 71: 'HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37AAAAA C AAAAA AAAAAAAAAA
SEQ ID NO 72: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR
G37AAAAA C AAAAA AAAAAAAAAAAAAAA
SEQ ID NO 73： 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37V C vvvvv vvvvv
SEQ ID NO 74: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37V C VVVVV VVVVVVVVVV
SEQ ID NO 75: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37V C VVVVV vvvvvvvvvvvvvvv
SEQ ID NO 76: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37VV C VVVVV VVVVV
SEQ ID NO 77: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37VV C VVVVV VVVVVVVVVV SEQ ID NO 78: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37VV C
VVVVV vvvvvvvvvvvvvvv
SEQ ID NO 79: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR
G37VVVVV C VVVVV VVVVV
SEQ ID NO 80: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR
G37VVVVV C VVVVV VVVVVVVVVV
SEQ ID NO 81: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR
G37VVVVV C VVVVV VVVVVVVVVVVVVVV
将上述多肽各 1 mg溶于 1 ml生理盐水制成多肽溶液， 将此多肽溶液皮 下注射于小鼠 （200μ1/只， 6只 /组， 购自中科院上海实验动物中心） ， 给药 30分钟后将 400微克葡萄糖注射于每只小鼠。分别于葡萄糖注射后 2小时、
24小时、 48小时、 72小时及 96小时测定小鼠的血糖（注： 每次测血糖前两 个小时再次给相同剂量的葡萄糖），结果见图 3 ,其中本实施例中的对照（空 白） 为向小鼠皮下注射生理盐水， 用量 lml, 葡萄糖的注射与其他试验组相 同。 实施例 5: 基于其他位点的氨基酸突变为半胱氨酸的 GLP-1类似物
本实施例中， 采用的多肽如下： SEQ ID NO 82: 'HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR GGGGG GGGGG C GGGGG GGGGG
SEQ ID NO 83: 7HAEGT FTSDV SSYLE CQAAK EFIAW LVKGR G37 GGGGG C GGGGG GGGGG SEQ ID NO 84: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37 GGG C GGGGG GGGGGGGGGGGGGGG
将上述多肽各 1 mg溶于 1 ml生理盐水， 皮下注射于小鼠（200 μΐ/只， 6 只 /组， 购自中科院上海实验动物中心）， 给药 30分钟后将 400 mg葡萄糖注 射于每只小鼠。 分别于葡萄糖注射后 2小时、 24小时、 48小时、 72小时及 96小时测定小鼠的血糖， 结果见图 4, 其中： 本实施例中的对照（空白）为 生理盐水， 用量 lml。 实施例 6: GLP-1类似物在人血清中的稳定性测定
本实施例中， 采用的多肽如下：
SEQ ID NO 58: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR
G37GG C GGGGG GGGGG
SEQ ID NO 59: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37GG C GGGGG GGGGGGGGGG
用真空采血针（ BD Biosciences, Franklin Lakes, NJ )取志愿者血样三 份， 而后立即于离心机上 13000rpm离心 20分钟， 取上层血清备用。
将上述两条多肽及 GLP-1标准品（购自上海生工公司， 其序列为： HAEGTFTSDVSSYLEGQAAKEFIAWLVKGRG ) 各 O.lmg溶于 0.5 ml生理 盐水中， 充分溶解后分别加入到 lml血清中， 将血清标记为空白组（O.lmg GLP-1溶于 0.5 ml生理盐水中）、 NO 58实验组（ O.lmg序列为 SEQ ID NO 58的肽溶于 0.5 ml生理盐水中）和 NO 59实验组（ O.lmg序列为 SEQ ID NO 59的肽溶于 0.5 ml生理盐水中）。 于 37°C孵育 0, 0.1, 0.3, 2.5, 5, 10, 15及 24 h后，分别取 50 μΐ血清混合物， 用 GLP-1 酶联免疫检测试剂盒（购自 ALPCO Immunoassays )对各组血清进行检测， 于 SpectraMax M5 酶标仪上读取吸光 值， 算出不同血清样品中 GLP-1类似物在不同时间点的浓度， 结果见图 5。 其中圓圏代表空白组， 倒三角代表 NO 58实验组， 正三角代表 NO 59实验 组。 结果表明 No 58及 No 59实验组的半衰期>2411。 实施例 7: GLP-1类似物在人血清中的稳定性测定
本实施例中， 采用的多肽如下：
SEQ ID NO 3: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR
G37GGGGG GGGGGGGGGGC GGG
SEQ ID NO 25: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR
G37VVVVV VVVVVC VVVVV WW
用真空采血针（ BD Biosciences, Franklin Lakes, NJ )取志愿者血样三 份， 而后立即于离心机上 13000rpm离心 20分钟， 取上层血清备用。
将上述多肽及 GLP-1标准品（购自上海生工公司，其序列为： SEQ ID NO 85, HAEGTFTSDVSSYLEGQAAKEFIAWLVKGRG) 各 O.lmg溶于 0.5 ml 生理盐水中， 充分溶解后分别加入到血清中， 将血清标记为空白组（O.lmg GLP-1溶于 0.5 ml生理盐水中 )、 N0.3实验组（ O.lmg序列为 SEQ ID NO 3 的肽溶于 0.5 ml生理盐水中）和 NO 25实验组（ O.lmg序列为 SEQ ID NO 25 的肽溶于 0.5 ml生理盐水中 ) 。 于 37°C孵育 0, 0.5, 1, 4, 8, 12, 24, 48, 72及 96 h后， 分别取 50 μΐ血清混合物， 用 GLP-1 酶联免疫检测试剂盒（购自 ALPCO Immunoassays )对各组血清进行检测， 于 SpectraMax M5 酶标仪上 读取吸光值， 算出不同血清样品中 GLP-1类似物在不同时间点的浓度， 结果 见图 6。 其中倒三角代表代表空白组， 空心圓圏 N0 3实验组， 实心圓圏代 表 N025实验组。 结果表明 No3及 No25实验组的半衰期为 48h 实施例 8: GLP-1类似物在人血清中的稳定性测定
本实施例中， 采用的多肽如下：
SEQ ID NO 28: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37G C GGGGG GGGGG;
SEQ ID NO 29: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37G
C GGGGG GGGGGGGGGG;
用真空采血针（ BD Biosciences, Franklin Lakes, NJ )取志愿者血样三 份， 而后立即于离心机上 13000rpm离心 20分钟， 取上层血清备用。
将上述多肽及 GLP-1标准品（购自上海生工公司，其序列为： SEQ ID NO 85, HAEGTFTSDVSSYLEGQAAKEFIAWLVKGRG )各 O.lmg溶于 0.5 ml 生理盐水中， 充分溶解后分别加入到血清中， 将血清标记为空白组（O.lmg GLP-1溶于 0.5 ml生理盐水中 )、 N0.28实验组（ O.lmg序列为 SEQ ID NO 28的肽溶于 0.5 ml生理盐水中）和 NO 29实验组（ O.lmg序列为 SEQ ID NO 29的肽溶于 0.5 ml生理盐水中 ) 。 于 37°C孵育 0, 0.5, 1, 4, 8, 12, 24及 48h 后，分别取 50 μΐ血清混合物， 用 GLP-1 酶联免疫检测试剂盒（购自 ALPCO Immunoassays )对各组血清进行检测， 于 SpectraMax M5 酶标仪上读取吸光 值， 算出不同血清样品中 GLP-1类似物在不同时间点的浓度， 结果见图 7。 其中倒三角代表代表空白组， 空心圓圏 N0 28实验组，实心圓圏代表 N029 实验组。 结果表明 No28及 No29实验组的半衰期为 24h。

Claims

权 利 要 求
1、 一种 GLP-1类似物， 所述 GLP-1类似物具有下列通式 A:
7HAEX10T FTSX15V SSYLE X22QAAK EFIX30W LX33KGR G37niXiC n2X2 通式 A,
其中， 。为甘氨酸或半胱氨酸， X15为天冬氨酸或半胱氨酸， X22为甘氨 酸或半胱氨酸， X3。为丙氨酸或半胱氨酸， X33为缬氨酸或半胱氨酸，并且 X1()、 χ15、 χ22、 χ3。和 χ33中至少有一个为半胱氨酸；
i^Xi表示有 个 ^ ηι=1~30 , 且 为甘氨酸、 丙氨酸或缬氨酸； n2X2表示有 n2个 X2, n2=0~30 , 且 X2为甘氨酸、 丙氨酸或缬氨酸； 以及 通式 A中含有的两个半胱氨酸均形成二硫键。
2、 如权利要求 1所述的 GLP-1类似物， 其特征在于， 所述 GLP-1类似 物具有下列通式 I：
7HAEX10T FTSX15V SSYLE X22QAAK EFIX30W LX33KGR G^ ^C n2X2 通式 I,
其中 1^=3 - 30, n2=3 - 30;
优选地， 所述 GLP- 1类似物为：
SEQ ID NO 82: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37 GGGGG GGGGG C GGGGG GGGGG;
SEQ ID NO 83: 7HAEGT FTSDV SSYLE CQAAK EFIAW LVKGR G37 GGGGG C GGGGG GGGGG; 或
SEQ ID NO 84: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37 GGG C GGGGG GGGGGGGGGGGGGGG。
3、 如权利要求 1所述的 GLP-1类似物， 其特征在于， 所述 GLP-1类似 物具有下列通式 Π:
7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37niXiC n2X2
通式 Π,
1^=1 - 20, n2=0 ~ 25 , 并且其中 X为甘氨酸、 丙氨酸或缬氨酸， X2为甘 氨酸、 丙氨酸或缬氨酸；
优选地， 11!=3 ~ 20, n2=3 ~ 25； 进一步优选地， 所述 11尸5 ~ 15, n2=3 ~ 9。
4、 如权利要求 3所述的 GLP-1类似物， 其特征在于， 所述 GLP-1类似 物为：
SEQ ID NO 1： 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR
G37GGGGG C GGG;
SEQ ID NO 2: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37GGGGG GGGGGC GGG;
SEQ ID NO 3: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37GGGGG GGGGGGGGGGC GGG;
SEQ ID NO 4: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37GGGGG C GGGGG G;
SEQ ID NO 5: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37GGGGG C GGGGG GGGG;
SEQ ID NO 6: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR
G37GGGGG GGGGGC GGGGG G;
SEQ ID NO 7: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37GGGGG GGGGGC GGGGG GGGG;
SEQ ID NO 8: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37GGGGG GGGGGGGGGGC GGGGG G;
SEQ ID NO 9: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37GGGGG GGGGGGGGGGC GGGGG GGGG;
SEQ ID NO 10: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37AAAAA C AAA;
SEQ ID NO 11: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR
G37AAAAA AAAAAC AAA;
SEQ ID NO 12: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37AAAAA AAAAAAAAAAC AAA;
SEQ ID NO 13: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37AAAAA C AAAAA A;
SEQ ID NO 14: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37AAAAA C AAAAA AAAA; SEQ ID NO 15: 'HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37AAAAA AAAAAC AAAAA A;
SEQ ID NO 16: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37AAAAA AAAAAC AAAAA AAAA;
SEQ ID NO 17: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37AAAAA AAAAAAAAAAC AAAAA A;
SEQ ID NO 18: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37AAAAA AAAAAAAAAAC AAAAA AAAA;
SEQ ID NO 19: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37VVVVV C VVV;
SEQ ID NO 20: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37VVVVV VVVVVC VVV;
SEQ ID NO 21 : 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37VVVVV VVVVVVVVVVC VVV;
SEQ ID NO 22: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37VVVVV C VVVVV V;
SEQ ID NO 23: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37VVVVV C VVVVV WW;
SEQ ID NO 24: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37VVVVV VVVVVC VVVVV V;
SEQ ID NO 25: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37VVVVV VVVVVC VVVVV WW;
SEQ ID NO 26: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37VVVVV VVVVVVVVVVC VVVVV V; 或
SEQ ID NO 27: 7HAEGT FTSCV SSYLE GQAAK EFIAW LVKGR G37VVVVV VVVVVVVVVVC VVVVV VVVV。
5、 如权利要求 1所述的 GLP-1类似物， 其特征在于， 所述 GLP-1类似 物具有下列通式 III：
7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37 C n2X2 通式 ΠΙ,
其中， ηι=1 ~ 10, n2=0 ~ 25 , 为甘氨酸、 丙氨酸或缬氨酸， X2为甘氨 酸、 丙氨酸或缬氨酸； 优选地， 1^=3 - 10, n2=3 ~ 25；
或者， 11!=1 - 5, n2=10 ~ 20;
进一步优选地， ^=3 ~ 5, n2=10 ~ 20。
6、 如权利要求 5所述的 GLP-1类似物， 其特征在于， 所述 GLP-1类似 物为：
SEQ ID NO 28: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37G C GGGGG GGGGG;
SEQ ID NO 29: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37G C GGGGG GGGGGGGGGG；
SEQ ID NO 30: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37G C GGGGG GGGGGGGGGGGGGGGGGGGG；
SEQ ID NO 31: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37GG C GGGGG GGGGG;
SEQ ID NO 32: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR
G37GG C GGGGG GGGGGGGGGG;
SEQ ID NO 33: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37GG C GGGGG GGGGGGGGGGGGGGG;
SEQ ID NO 34: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37GGGGG C GGGGG GGGGG；
SEQ ID NO 35: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37GGGGG C GGGGG GGGGGGGGGG;
SEQ ID NO 36: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37GGGGG C GGGGG GGGGGGGGGGGGGGG;
SEQ ID NO 37: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37A
C AAAAA AAAAA;
SEQ ID NO 38: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37A C AAAAA AAAAAAAAAA;
SEQ ID NO 39: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37A C AAAAA AAAAAAAAAAAAAAAAAAAA；
SEQ ID NO 40: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37AA C AAAAA AAAAA; SEQ ID NO 41 : 'HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37AA C AAAAA AAAAAAAAAA;
SEQ ID NO 42: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37AA C AAAAA AAAAAAAAAAAAAAA;
SEQ ID NO 43： 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR
G37AAAAA C AAAAA AAAAA;
SEQ ID NO 44: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37AAAAA C AAAAA AAAAAAAAAA;
SEQ ID NO 45: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37AAAAA C AAAAA AAAAAAAAAAAAAAA;
SEQ ID NO 46: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37V C VVVVV VVVVV;
SEQ ID NO 47: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37V C VVVVV VVVVVVVVVV;
SEQ ID NO 48: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37V
C VVVVV VVV VVVVVVVVVV vvvvvvv;
SEQ ID NO 49: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37VV C VVVVV VVVVV;
SEQ ID NO 50: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37VV C VVVVV VVVVVVVVVV;
SEQ ID NO 51 : 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37VV C VVVVV VVVVVVVVVVVVVVV;
SEQ ID NO 52: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37VVVVV C VVVVV VVVVV;
SEQ ID NO 53： 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR
G37VVVVV C VVVVV VVVVVVVVVV; 或
SEQ ID NO 54: 7HAEGT FTSDV SSYLE GQAAK EFICW LVKGR G37VVVVV C VVVVV VVVVVVVVVVVVVVV。
7、 如权利要求 1所述的 GLP-1类似物， 其特征在于， 所述 GLP-1类似 物具有下列通式 IV：
7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37 C n2X2 通式 IV
其中， ηι=1 ~ 10, η2=0 ~ 30, 为甘氨酸、 丙氨酸或缬氨酸， X2为甘氨 酸、 丙氨酸或缬氨酸； 优选地， 1^=3 - 10, n2=3 - 30;
或者， 11!=1 - 5, n2=10 ~ 20;
进一步优选地 11尸3 ~ 5, n2=10 ~ 20o
8、 如权利要求 7所述的 GLP-1类似物， 其特征在于， 所述 GLP-1类似 物为：
SEQ ID NO 55: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37G C GGGGG GGGGG;
SEQ ID NO 56: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37G C GGGGG GGGGGGGGGG;
SEQ ID NO 57: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37G C GGGGG GGGGGGGGGGGGGGG；
SEQ ID NO 58: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR
G37GG C GGGGG GGGGG;
SEQ ID NO 59: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37GG C GGGGG GGGGGGGGGG;
SEQ ID NO 60: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37GG C GGGGG GGGGGGGGGGGGGGG;
SEQ ID NO 61: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37GGGGG C GGGGG GGGGG;
SEQ ID NO 62: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37GGGGG C GGGGG GGGGGGGGGG;
SEQ ID NO 63： 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR
G37GGGGG C GGGGG GGGGGGGGGGGGGGG;
SEQ ID NO 64: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37A C AAAAA AAAAA;
SEQ ID NO 65: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37A C AAAAA AAAAAAAAAA；
SEQ ID NO 66: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37A C AAAAA AAAAA AAAAAAAAAA; SEQ ID NO 67: 'HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37AA C AAAAA AAAAA;
SEQ ID NO 68: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37AA C AAAAA AAAAAAAAAA;
SEQ ID NO 69: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR
G37AA C AAAAA AAAAAAAAAAAAAAA;
SEQ ID NO 70: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37AAAAA C AAAAA AAAAA;
SEQ ID NO 71: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37AAAAA C AAAAA AAAAAAAAAA;
SEQ ID NO 72: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37AAAAA C AAAAA AAAAAAAAAAAAAAA;
SEQ ID NO 73: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37V C VVVVV VVVVV;
SEQ ID NO 74: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37V
C VVVVV VVVVVVVVVV;
SEQ ID NO 75: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37V C VVVVV VVVVV VVVVVVVVVV;
SEQ ID NO 76: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37VV C VVVVV VVVVV；
SEQ ID NO 77: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37VV C VVVVV VVVVVVVVVV;
SEQ ID NO 78: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37VV C VVVVV VVVVVVVVVVVVVVV;
SEQ ID NO 79: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR
G37VVVVV C VVVVV VVVVV;
SEQ ID NO 80: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37VVVVV C VVVVV VVVVVVVVVV; 或
SEQ ID NO 81: 7HAEGT FTSDV SSYLE GQAAK EFICW LCKGR G37VVVVV C VVVVV VVVVVVVVVVVVVVV。
9、 如权利要求 1-8中任一项所述的 GLP-1类似物的制备方法， 所述制 备方法包括 Fmoc策略的固相多肽合成。
10、 如权利要求 1-8中任一项所述的 GLP-1类似物在制备治疗糖尿病、 肥胖症和 /或糖尿病、 肥胖症相关疾病的药物中的应用。
11、如权利要求 1-8中任一项所述的 GLP-1类似物在制备预防肥胖症的 药物中的应用。
12、 一种药物组合物， 所述药物组合物包含如权利要求 1 - 8中任一项所 述的 GLP-1类似物和一种或多种药学上可接受的辅料；
优选地， 所述药物组合物为注射剂， 进一步优选为冻干粉针或溶液注射 剂。
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CN102718858B (zh) * 2011-03-29 2014-07-02 天津药物研究院 胰高血糖素样肽-1类似物单体、二聚体及其制备方法与应用
CN103059127B (zh) * 2013-01-07 2014-12-17 天津嘉宏科技有限公司 Glp-1类似物及其制备方法与应用
CN103044540A (zh) * 2013-01-07 2013-04-17 天津嘉宏科技有限公司 一种glp-1类似物、制备方法及其应用
US9707273B2 (en) * 2013-03-01 2017-07-18 Fundació Hospital Universitari Vall D'hebron—Institut De Recerca Peptides for use in the topical treatment of retinal neurodegenerative diseases, in particular in early stages of diabetic retinopathy and other retinal diseases in which neurodegeneration plays an essential role
DK2976325T3 (en) 2013-03-21 2017-06-06 Sanofi Aventis Deutschland SYNTHESIS OF PEPTIDE PRODUCTS CONTAINING CYCLIC IMID
CA2907454C (en) 2013-03-21 2021-05-04 Sanofi-Aventis Deutschland Gmbh Synthesis of hydantoin containing peptide products
CN104262481B (zh) * 2013-08-09 2018-02-09 天津药物研究院有限公司 一种侧链修饰的长效glp‑1类似物的制备方法及其应用
CN103787938B (zh) * 2014-01-29 2016-08-17 清华大学 化合物及其制备方法和应用
CN106554409B (zh) * 2015-09-30 2020-04-07 天津药物研究院有限公司 一种长效胰高血糖素样肽-1类似物及其应用
CN112898406B (zh) * 2019-10-12 2023-11-10 深圳纳福生物医药有限公司 不同构型的glp-1类似肽修饰二聚体及其制备方法在治疗ii型糖尿病中的应用

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1088835A (zh) * 1992-03-19 1994-07-06 诺沃挪第克公司 肽的应用
CN1106698A (zh) * 1993-04-07 1995-08-16 美国辉瑞有限公司 肽的延长释放
CN1242707A (zh) * 1996-11-05 2000-01-26 伊莱利利公司 外周给予的glp－1类似物和衍生物在控制肥胖中的用途

Family Cites Families (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
BR112012028704A2 (pt) * 2010-05-13 2019-09-24 Univ Indiana Res & Tech Corp composto de glugagon da superfamília de peptídeos exibindo atividade do receptor com hormônio nuclear, pró fármaco, dímero ou multímero, composição farmacêutica que o compreendem e método de administração do mesmo.

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1088835A (zh) * 1992-03-19 1994-07-06 诺沃挪第克公司 肽的应用
CN1106698A (zh) * 1993-04-07 1995-08-16 美国辉瑞有限公司 肽的延长释放
CN1242707A (zh) * 1996-11-05 2000-01-26 伊莱利利公司 外周给予的glp－1类似物和衍生物在控制肥胖中的用途

Non-Patent Citations (5)

* Cited by examiner, † Cited by third party
Title
"Fmoc solid phase peptide synthesis: a practical approach", 2000, OXFORD UNIVERSITY PRESS
DRUCKER D J, DIABETES, vol. 47, no. 2, February 1998 (1998-02-01), pages 159 - 69
LI, Y. ET AL.: "GLP-1 analogs containing disulfide bond exhibited prolonged half-life in vivo than GLP-1.", PEPTIDES., vol. 32, 14 April 2011 (2011-04-14), pages 1303 - 1312, XP055179602 *
MOJSOV S, J CLIN INVEST., vol. 79, no. 2, February 1987 (1987-02-01), pages 616 - 9
See also references of EP2894163A4 *

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