Pharmaceutical composition

Abstract

The present invention provides a pharmaceutical composition comprising a polypeptide or a pharmaceutically acceptable salt thereof. The polypeptide or the pharmaceutically acceptable salt thereof comprises the N-terminus of an amino acid sequence as shown in X1-Aib-X2, wherein X1 is a natural or unnatural amino acid residue comprising an imidazole group, and X2 is a natural or unnatural amino acid residue comprising an imidazole group. The pharmaceutical composition has good chemical stability.

Description

Pharmaceutical composition TECHNICAL FIELD

[0001] The present disclosure belongs to the field of medicine, and relates to a pharmaceutical composition comprising a polypeptide or a pharmaceutically acceptable salt thereof. BACKGROUND

[0002] GLP-1 drugs have become a research hotspot in recent years, which not only has a significant effect on reducing blood sugar, but also has the effects of weight loss, blood pressure reduction, improvement of blood lipid spectrum, and heart protection, and has become one of the most ideal therapeutic drugs for type 2 diabetes, obesity and other chronic diseases.

[0003] At present, polypeptide preparations for subcutaneous injection are mostly stored in an environment of 2-8°C before use. During use, the preparations may be subjected to room temperature conditions, and therefore, research on clinical use is an indispensable part of drug development, which can guarantee the quality of drugs in use, and also provide a basis for the preparation, storage conditions and use period of the prepared or diluted drugs in the instructions / labels of the drugs.

[0004] The present disclosure provides a pharmaceutical composition comprising a polypeptide, which has good chemical stability and can ensure that the quality requirements are met during storage and use. SUMMARY

[0005] The present disclosure provides a pharmaceutical composition comprising a polypeptide or a pharmaceutically acceptable salt thereof.

[0006] In some embodiments, the pharmaceutical composition described in the present disclosure further comprises a metal ion.

[0007] In some embodiments, the pharmaceutical composition described in the present disclosure further comprises a divalent metal ion.

[0008] In some embodiments, the metal ion described in the present disclosure can be selected from a divalent zinc ion, a divalent calcium ion, a divalent magnesium ion, and a divalent iron ion.

[0009] In some embodiments, the metal ion described in the present disclosure can be selected from a divalent zinc ion, a divalent calcium ion, and a divalent magnesium ion.

[0010] In some embodiments, the metal ion described in the present disclosure can be selected from a divalent zinc ion.

[0011] In some embodiments, the metal ion described in the present disclosure can be selected from zinc acetate or zinc chloride.

[0012] In some embodiments, the metal ion described in the present disclosure can be selected from zinc acetate.

[0013] In some embodiments, the pharmaceutical composition of the present disclosure comprises a polypeptide or a pharmaceutically acceptable salt thereof and a divalent metal ion; preferably the pharmaceutical composition comprises a polypeptide or a pharmaceutically acceptable salt thereof and zinc acetate or zinc chloride.

[0014] In some embodiments, the divalent metal ion of the present disclosure is not selected from divalent iron ions.

[0015] In some embodiments, the molar ratio of the polypeptide or a pharmaceutically acceptable salt thereof and the metal ion in the pharmaceutical composition of the present disclosure is 1:5-5:1; preferably the molar ratio of the polypeptide or a pharmaceutically acceptable salt thereof and the metal ion is 1:3-3:1; preferably the molar ratio of the polypeptide or a pharmaceutically acceptable salt thereof and the metal ion is 1:2-2:1; preferably the molar ratio of the polypeptide or a pharmaceutically acceptable salt thereof and the metal ion is 1:1.

[0016] In some embodiments, the molar ratio of the polypeptide or a pharmaceutically acceptable salt thereof and the divalent zinc ion in the pharmaceutical composition of the present disclosure is 1:5-5:1; preferably the molar ratio of the polypeptide or a pharmaceutically acceptable salt thereof and the divalent zinc ion is 1:3-3:1; preferably the molar ratio of the polypeptide or a pharmaceutically acceptable salt thereof and the divalent zinc ion is 1:2-2:1; preferably the molar ratio of the polypeptide or a pharmaceutically acceptable salt thereof and the divalent zinc ion is 1:1.

[0017] In some embodiments, the concentration of the polypeptide or pharmaceutically acceptable salt thereof in the pharmaceutical composition is 0.01 mg / mL to 1000 mg / mL, e.g., 0.1 mg / mL to 500 mg / mL, 0.1 mg / mL to 400 mg / mL, 0.1 mg / mL to 300 mg / mL, 0.1 mg / mL to 200 mg / mL, 0.1 mg / mL to 100 mg / mL, 0.5 mg / mL to 200 mg / mL, 0.5 mg / mL to 150 mg / mL, 0.5 mg / mL to 100 mg / mL, 0.5 mg / mL to 50 mg / mL, 0.5 mg / mL to 25 mg / mL, 1.0 mg / mL to 100 mg / mL, 1.0 mg / mL to 90 mg / mL, 1.0 mg / mL to 80 mg / mL, 1.0 mg / mL to 70 mg / mL, 1.0 mg / mL to 60 mg / mL, 1.0 mg / mL to 50 mg / mL, 1.0 mg / mL to 40 mg / mL, 1.0 mg / mL to 30 mg / mL, 1.0 mg / mL to 20 mg / mL, 2.0 mg / mL to 50 mg / mL, 2.0 mg / mL to 40 mg / mL, 2.0 mg / mL to 30 mg / mL, 2.0 mg / mL to 20 mg / mL, 5.0 mg / mL to 1000 mg / mL, 5.0 mg / mL to 500 mg / mL, 5.0 mg / mL to 400 mg / mL, 5.0 mg / mL to 300 mg / mL, 5.0 mg / mL to 200 mg / mL, 5.0 mg / mL to 100 mg / mL, 5.0 mg / mL to 90 mg / mL, 5.0 mg / mL to 80 mg / mL, 5.0 mg / mL to 70 mg / mL, 5.0 mg / mL to 60 mg / mL, 5.0 mg / mL to 50 mg / mL, 5.0 mg / mL to 40 mg / mL, 5.0 mg / mL to 30 mg / mL, 5.0 mg / mL to 20 mg / mL, 5.0 mg / mL to 10 mg / mL, 6.0 mg / mL to 1000 mg / mL, 6.0 mg / mL to 500 mg / mL, 6.0 mg / mL to 400 mg / mL, 6.0 mg / mL to 300 mg / mL, 6.0 mg / mL to 200 mg / mL, 6.0 mg / mL to 100 mg / mL, 6.0 mg / mL to 90 mg / mL, 6.0 mg / mL to 80 mg / mL, 6.0 mg / mL to 70 mg / mL, 6.0 mg / mL to 60 mg / mL, 6.0 mg / mL to 50 mg / mL, 6.0 mg / mL to 40 mg / mL, 6.0 mg / mL to 30 mg / mL, 6.0 mg / mL to 20 mg / mL.

[0018] In some embodiments, the concentration of the polypeptide or pharmaceutically acceptable salt thereof in the pharmaceutical composition is 1.0 mg / mL, 2.0 mg / mL, 3.0 mg / mL, 4.0 mg / mL, 5.0 mg / mL, 6.0 mg / mL, 7.0 mg / mL, 8.0 mg / mL, 9.0 mg / mL, 10.0 mg / mL, 11.0 mg / mL, 12.0 mg / mL, 13.0 mg / mL, 14.0 mg / mL, 15.0 mg / mL, 16.0 mg / mL, 17.0 mg / mL, 18.0 mg / mL, 19.0 mg / mL, 20.0 mg / mL, 21.0 mg / mL, 22.0 mg / mL, 23.0 mg / mL, 24.0 mg / mL, 25.0 mg / mL, 26.0 mg / mL, 27.0 mg / mL, 28.0 mg / mL, 29.0 mg / mL, 30.0 mg / mL.

[0019] In some embodiments, the concentration of the polypeptide or pharmaceutically acceptable salt thereof in the pharmaceutical composition is 12.0 mg / mL.

[0020] In some embodiments, the concentration of the divalent zinc ion in the pharmaceutical composition is 0.001 mg / mL to 100 mg / mL, 0.001 mg / mL to 90 mg / mL, 0.001 mg / mL to 80 mg / mL, 0.001 mg / mL to 70 mg / mL, 0.001 mg / mL to 60 mg / mL, 0.01 mg / mL to 50 mg / mL, 0.01 mg / mL to 40 mg / mL, 0.01 mg / mL to 30 mg / mL, 0.01 mg / mL to 20 mg / mL, 0.01 mg / mL to 10 mg / mL, 0.01 mg / mL to 5 mg / mL, 0.01 mg / mL to 3 mg / mL, 0.1 mg / mL to 5 mg / mL, 0.1 mg / mL to 3 mg / mL, 0.1 mg / mL to 1.5 mg / mL.

[0021] In some embodiments, the concentration of zinc acetate in the pharmaceutical composition is from 0.001 mg / mL to 100 mg / mL, from 0.001 mg / mL to 90 mg / mL, from 0.001 mg / mL to 80 mg / mL, from 0.001 mg / mL to 70 mg / mL, from 0.001 mg / mL to 60 mg / mL, from 0.01 mg / mL to 50 mg / mL, from 0.01 mg / mL to 40 mg / mL, from 0.01 mg / mL to 30 mg / mL, from 0.01 mg / mL to 20 mg / mL, from 0.01 mg / mL to 10 mg / mL, from 0.01 mg / mL to 5 mg / mL, from 0.01 mg / mL to 3 mg / mL, from 0.1 mg / mL to 5 mg / mL, from 0.1 mg / mL to 3 mg / mL, from 0.1 mg / mL to 1.5 mg / mL.

[0022] In some embodiments, the concentration of zinc acetate in the pharmaceutical composition is selected from 0.25 mg / mL, 0.5 mg / mL, 1.0 mg / mL.

[0023] In some embodiments, the pharmaceutical composition described in the present disclosure further comprises a buffer.

[0024] In some embodiments, the buffer described in the present disclosure is selected from one or more of acetate buffer, histidine salt buffer, phosphate buffer, succinate buffer, Tris, HEPES, MOPS, diethanolamine buffer, borate buffer, Tricine, and citrate buffer.

[0025] In some embodiments, the buffer described in the present disclosure is selected from Tris buffer, disodium hydrogen phosphate buffer, HEPES buffer.

[0026] In some embodiments, the concentration of the buffering agent in the pharmaceutical composition is 0.5 mM to 50.0 mM, for example, 0.5 mM to 40.0 mM, 0.5 mM to 30.0 mM, 0.5 mM to 20.0 mM, 0.5 mM to 10.0 mM, 1.0 mM to 40.0 mM, 1.0 mM to 35.0 mM, 1.0 mM to 30.0 mM, 1.0 mM to 25.0 mM, 1.0 mM to 20.0 mM, 1.0 mM to 15.0 mM, 1.0 mM to 10.0 mM, 2.0 mM to 40.0 mM, 2.0 mM to 35.0 mM, 2.0 mM to 30.0 mM, 2.0 mM to 25.0 mM, 2.0 mM to 20.0 mM, 2.0 mM to 15.0 mM, 5.0 mM to 15.0 mM, 2.0 mM to 10.0 mM, or 5.0 mM to 10.0 mM.

[0027] In some embodiments, the concentration of the buffering agent in the pharmaceutical composition is 1.0 mM, 2.0 mM, 3.0 mM, 4.0 mM, 5.0 mM, 6.0 mM, 7.0 mM, 8.0 mM, 9.0 mM, or 10.0 mM.

[0028] In some embodiments, the concentration of the buffering agent in the pharmaceutical composition is 10.0 mM.

[0029] In some embodiments, the pharmaceutical composition further comprises an osmotic pressure adjusting agent. The osmotic pressure adjusting agent includes, but is not limited to, a salt (e.g., sodium chloride, phosphate, sodium citrate, boric acid, and sodium tartrate), a sugar or sugar alcohol (lactose, trehalose, sucrose, glucose, mannitol, sorbitol, xylitol), an amino acid (e.g., L-glycine, L-histidine, arginine, lysine, isoleucine, aspartic acid, tryptophan, threonine), a polyol [e.g., glycerol, 1,2-propanediol (also referred to as propylene glycol in the present disclosure), 1,3-propanediol, 1,3-butanediol], a polyethylene glycol (e.g., PEG 400), or a mixture thereof.

[0030] In some embodiments, the osmotic pressure adjusting agent is selected from one or more of propylene glycol, mannitol, sorbitol, xylitol, glycerol, lactose, trehalose, sucrose, glucose, sodium chloride, phosphate, sodium citrate, boric acid, and sodium tartrate.

[0031] In some embodiments, the osmotic pressure adjusting agent is sodium chloride, propylene glycol, mannitol, glycerol.

[0032] In some embodiments, the osmotic pressure adjusting agent is propylene glycol or mannitol.

[0033] In some embodiments, the concentration of propylene glycol in the pharmaceutical composition is 10 mg / mL to 20 mg / mL, for example, 10 mg / mL to 19 mg / mL, 10 mg / mL to 18 mg / mL, 10 mg / mL to 17 mg / mL, 10 mg / mL to 16 mg / mL, 10 mg / mL to 15 mg / mL, 11 mg / mL to 20 mg / mL, 11 mg / mL to 19 mg / mL, 11 mg / mL to 18 mg / mL, 11 mg / mL to 17 mg / mL, 11 mg / mL to 16 mg / mL, 11 mg / mL to 15 mg / mL, 12 mg / mL to 20 mg / mL, 12 mg / mL to 19 mg / mL, 12 mg / mL to 18 mg / mL, 12 mg / mL to 17 mg / mL, 12 mg / mL to 16 mg / mL, 12 mg / mL to 15 mg / mL, 13 mg / mL to 20 mg / mL, 13 mg / mL to 19 mg / mL, 13 mg / mL to 18 mg / mL, 13 mg / mL to 17 mg / mL, 13 mg / mL to 16 mg / mL, 13 mg / mL to 15 mg / mL.

[0034] In some embodiments, the concentration of propylene glycol in the pharmaceutical composition is 12 mg / mL to 16 mg / mL, for example, 14 mg / mL.

[0035] In some embodiments, the concentration of sodium chloride in the pharmaceutical composition is 1 mg / mL to 20 mg / mL, for example, 1 mg / mL to 19 mg / mL, 1 mg / mL to 18 mg / mL, 1 mg / mL to 17 mg / mL, 1 mg / mL to 16 mg / mL, 1 mg / mL to 15 mg / mL, 2 mg / mL to 18 mg / mL, 2 mg / mL to 17 mg / mL, 2 mg / mL to 16 mg / mL, 2 mg / mL to 15 mg / mL, 3 mg / mL to 18 mg / mL, 3 mg / mL to 17 mg / mL, 3 mg / mL to 16 mg / mL, 3 mg / mL to 15 mg / mL, 4 mg / mL to 14 mg / mL, 5 mg / mL to 13 mg / mL, 6 mg / mL to 12 mg / mL, 7 mg / mL to 11 mg / mL, 7 mg / mL to 10.5 mg / mL, 7 mg / mL to 10 mg / mL, 7 mg / mL to 9 mg / mL, 7 mg / mL to 9.5 mg / mL, 7 mg / mL to 9 mg / mL, 7 mg / mL to 8.5 mg / mL, 7.5 mg / mL to 11 mg / mL, 7.5 mg / mL to 10.5 mg / mL, 7.5 mg / mL to 10 mg / mL, 7.5 mg / mL to 9.5 mg / mL, 7.5 mg / mL to 9.0 mg / mL, 7.5 mg / mL to 8.5 mg / mL, 8 mg / mL to 11 mg / mL, 8 mg / mL to 10.5 mg / mL, 8 mg / mL to 10 mg / mL, 8 mg / mL to 9.5 mg / mL, 8 mg / mL to 9 mg / mL.

[0036] In some embodiments, the concentration of sodium chloride in the pharmaceutical composition is 2 mg / mL to 18 mg / mL.

[0037] In some embodiments, the concentration of sodium chloride in the pharmaceutical composition is 3 mg / mL to 15 mg / mL.

[0038] In some embodiments, the concentration of sodium chloride in the pharmaceutical composition is 7 mg / mL to 10 mg / mL, for example, 7.5 mg / mL, 7.6 mg / mL, 7.7 mg / mL, 7.8 mg / mL, 7.9 mg / mL, 8 mg / mL, 8.1 mg / mL, 8.2 mg / mL, 8.3 mg / mL, 8.4 mg / mL, 8.5 mg / mL, 8.6 mg / mL, 8.7 mg / mL, 8.8 mg / mL, 8.9 mg / mL, 9.0 mg / mL, 9.1 mg / mL, 9.28 mg / mL, 9.3 mg / mL, 9.4 mg / mL, 9.5 mg / mL, 9.6 mg / mL, 9.7 mg / mL, 9.8 mg / mL, 9.9 mg / mL, or 10 mg / mL.

[0039] In some embodiments, the concentration of sodium chloride in the pharmaceutical composition is 7.5 mg / mL to 9.5 mg / mL.

[0040] In some embodiments, the concentration of mannitol in the pharmaceutical composition is 10 mg / mL to 60 mg / mL, for example 20 mg / mL to 60 mg / mL, 20 mg / mL to 55 mg / mL, 30 mg / mL to 55 mg / mL, 35 mg / mL to 55 mg / mL, 40 mg / mL to 55 mg / mL, 40 mg / mL to 50 mg / mL.

[0041] In some embodiments, the concentration of mannitol in the pharmaceutical composition is 30 mg / ml, 31 mg / ml, 32 mg / ml, 33 mg / ml, 34 mg / ml, 35 mg / ml, 36 mg / ml, 37 mg / ml, 38 mg / ml, 39 mg / ml, 40 mg / ml, 41 mg / ml, 42 mg / ml, 43 mg / ml, 44 mg / ml, 45 mg / ml, 46 mg / ml, 47 mg / ml, 48 mg / ml, 49 mg / ml, 50 mg / ml.

[0042] In some embodiments, the concentration of mannitol in the pharmaceutical composition is 35 mg / ml to 50 mg / ml, preferably the concentration of mannitol in the pharmaceutical composition is 46 mg / ml.

[0043] In some embodiments, the pharmaceutical composition further comprises a pH adjusting agent, for example sodium hydroxide and / or hydrochloric acid.

[0044] In some embodiments, the pH of the pharmaceutical composition is 6.5 to 9.0, for example 6.6 to 9.0, 6.7 to 9.0, 6.8 to 9.0, 6.9 to 9.0, 7.0 to 9.0, 7.1 to 9.0, 7.2 to 9.0, 7.3 to 9.0, 7.4 to 9.0, 7.5 to 9.0, 7.6 to 9.0, 7.7 to 9.0, 7.8 to 9.0, 7.9 to 9.0, 8.0 to 9.0, 7.0 to 8.9, 7.0 to 8.8, 7.0 to 8.7, 7.0 to 8.6, 7.0 to 8.5, 7.0 to 8.4, 7.0 to 8.3, 7.0 to 8.2, 7.0 to 8.1, 7.0 to 8.0, 7.1 to 8.9, 7.1 to 8.8, 7.1 to 8.7, 7.1 to 8.6, 7.1 to 8.5, 7.1 to 8.4, 7.1 to 8.3, 7.1 to 8.2, 7.1 to 8.1, 7.1 to 8.0, 7.1 to 7.9, 7.1 to 7.8, or 7.1 to 7.7.

[0045] In some embodiments, the pH of the pharmaceutical composition is 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9, 8.0, 8.1, 8.2, 8.3, 8.4, or 8.5.

[0046] In some embodiments, the pH of the pharmaceutical composition is 7.0 to 8.0.

[0047] In some embodiments, the pH of the pharmaceutical composition is 7.1 to 7.7, for example 7.5 or 7.4.

[0048] In some embodiments, the pharmaceutical composition further comprises a pharmaceutically acceptable bacteriostatic agent.

[0049] In some embodiments, the pharmaceutically acceptable bacteriostatic agent includes, but is not limited to, phenol, o-cresol, m-cresol, p-cresol, methyl paraben, propyl paraben, 2-phenoxyethanol, butyl paraben, 2-phenylethanol, benzyl alcohol, ethanol, chlorobutanol, and thimerosal, bronopol, benzoic acid, imidurea, chlorhexidine, sodium dehydroacetate, chlorocresol, ethylparaben, benzalkonium chloride, or mixtures thereof.

[0050] In some embodiments, the pharmaceutically acceptable bacteriostatic agent is phenol.

[0051] In some embodiments, the concentration of the bacteriostatic agent in the pharmaceutical composition is 4.0 mg / mL to 7.0 mg / mL, for example, 4.2 mg / mL to 6.9 mg / mL, 4.4 mg / mL to 6.9 mg / mL, 4.6 mg / mL to 6.9 mg / mL, 4.8 mg / mL to 6.9 mg / mL, 5.0 mg / mL to 6.9 mg / mL, 5.1 mg / mL to 6.9 mg / mL, 5.2 mg / mL to 6.9 mg / mL, 5.3 mg / mL to 6.9 mg / mL, 5.4 mg / mL to 6.9 mg / mL, 5.5 mg / mL to 6.9 mg / mL, 4.2 mg / mL to 6.8 mg / mL, 4.4 mg / mL to 6.8 mg / mL, 4.6 mg / mL to 6.8 mg / mL, 4.8 mg / mL to 6.8 mg / mL, 5.0 mg / mL to 6.8 mg / mL, 5.1 mg / mL to 6.8 mg / mL, 5.2 mg / mL to 6.8 mg / mL, 5.3 mg / mL to 6.8 mg / mL, 5.4 mg / mL to 6.8 mg / mL, 5.5 mg / mL to 6.8 mg / mL, 4.4 mg / mL to 6.7 mg / mL, 4.6 mg / mL to 6.7 mg / mL, 4.8 mg / mL to 6.7 mg / mL, 5.0 mg / mL to 6.7 mg / mL, 5.1 mg / mL to 6.7 mg / mL, 5.2 mg / mL to 6.7 mg / mL, 5.3 mg / mL to 6.7 mg / mL, 5.4 mg / mL to 6.7 mg / mL, 5.5 mg / mL to 6.7 mg / mL, 4.4 mg / mL to 6.6 mg / mL, 4.6 mg / mL to 6.6 mg / mL, 4.8 mg / mL to 6.6 mg / mL, 5.0 mg / mL to 6.6 mg / mL, 5.1 mg / mL to 6.6 mg / mL, 5.2 mg / mL to 6.6 mg / mL, 5.3 mg / mL to 6.6 mg / mL, 5.4 mg / mL to 6.6 mg / mL, 5.5 mg / mL to 6.6 mg / mL.

[0052] In some embodiments, the concentration of the bacteriostatic agent is 4.2 mg / mL to 6.9 mg / mL.

[0053] In some embodiments, the concentration of the bacteriostatic agent is 5.5 mg / mL to 6.6 mg / mL, for example, 5.5 mg / mL.

[0054] In some embodiments, the polypeptide or pharmaceutically acceptable salt thereof of the present disclosure comprises an N-terminus of an amino acid sequence as shown in X1-Aib-X2; wherein X1 is a natural or unnatural amino acid residue comprising an imidazole group, and X2 is a natural or unnatural amino acid residue comprising an imidazole group.

[0055] In some embodiments, the polypeptide or a pharmaceutically acceptable salt thereof described herein comprises an N-terminus of an amino acid sequence as shown by R1-X1-Aib-X2; wherein R1 is hydrogen, X1 is a natural or unnatural amino acid residue comprising an imidazole group, and X2 is a natural or unnatural amino acid residue comprising an imidazole group.

[0056] In some embodiments, the polypeptide or a pharmaceutically acceptable salt thereof described herein, wherein X1 is a His residue, and / or X2 is a His residue.

[0057] In some embodiments, the polypeptide or a pharmaceutically acceptable salt thereof described herein comprises an N-terminus of an amino acid sequence as shown by His-Aib-His.

[0058] In some embodiments, the polypeptide or a pharmaceutically acceptable salt thereof described herein comprises at least one Lys, Orn, Dap, Dab, or Cys residue having a side chain-linked substituent.

[0059] In some embodiments, the polypeptide or a pharmaceutically acceptable salt thereof described herein comprises at least one Lys residue having a side chain-linked substituent.

[0060] In some embodiments, the polypeptide or a pharmaceutically acceptable salt thereof described herein, wherein the substituent comprises a structure as shown by -Z1-Z2.

[0061] wherein,

[0062] Z1 comprises 0-10 residues independently selected from the group consisting of a Gly residue, a Glu residue, a γGlu residue, -C(O)-CH2-(O-CH2-CH2)2-NH-, -C(O)-(C6H 10 )-CH2-NH-, -C(O)-CH(NH2)-(CH2)4-NH-;

[0063] Z2 is selected from a C 12-32 fatty acid or a C 12-32 alkyl group.

[0064] In some embodiments, the pharmaceutical composition described herein, wherein the polypeptide or a pharmaceutically acceptable salt thereof comprises at least one Lys residue, and the epsilon-amino group of the Lys residue is linked to any one of the following substituents:

[0065] In some embodiments, the pharmaceutical composition described herein, wherein the polypeptide or a pharmaceutically acceptable salt thereof has agonistic activity on at least one of the following receptors: GLP-1 receptor, GIP receptor, GCG receptor.

[0066] In some embodiments, the polypeptide or pharmaceutically acceptable salt thereof in the pharmaceutical composition described in the present disclosure is a triple agonist of GLP-1 receptor, GIP receptor, and GCG receptor.

[0067] In some embodiments, the polypeptide or pharmaceutically acceptable salt thereof in the pharmaceutical composition described in the present disclosure comprises a sequence as shown below:

[0068] H-Aib-HGTFTSDYSIYLEKQAA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 2)

[0069] H-Aib-HGTFTSDYSIYLEKQAA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 2)

[0070] H-Aib-HGTFTSDYSIYLEKQAA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 2)

[0071] H-Aib-HGTFTSDYSIYLEKQAA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 2)

[0072] H-Aib-HGTFTSDYSIYLEKQAA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 2)

[0073] H-Aib-HGTFTSDYSIYLEKQAA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 2)

[0074] H-Aib-HGTFTSDYSIYLEKQAA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 2)

[0075] H-Aib-HGTFTSDYSIYLEKQAA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 2)

[0076] H-Aib-HGTFTSDYSIYLEKQAA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 2)

[0077] H-Aib-HGTFTSDYSIYLEKQAA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 2)

[0078] H-Aib-HGTFTSDYSIYLEKKAA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 11)

[0079] H-Aib-HGTFTSDYSIYLEKQKA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 12)

[0080] H-Aib-HGTFTSDYSKYLEKQAA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 13)

[0081] H-Aib-HGTFTSDYSIYLEKRAA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 14)

[0082] H-Aib-HGTFTSDYSILLEKKYA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 15)

[0083] H-Aib-HGTFTSDYSYLLEKQAA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 16)

[0084] H-Aib-HGTFTSDYSILLEKQAA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 17)

[0085] H-Aib-HGTFTSDYSILLEKQAAEEFVQWLLEGGPSSGAPPPS (SEQ ID NO: 18)

[0086] H-Aib-HGTFTSDYSILLEKQAAQEFVQWLLEGGPSSGAPPPS (SEQ ID NO: 19)

[0087] H-Aib-HGTFTSDYSILLEKQAAREFVQWLLEGGPSSGAPPPS (SEQ ID NO: 20)

[0088] H-Aib-HGTFTSDYSILLEKQAAHEFVQWLLEGGPSSGAPPPS (SEQ ID NO: 21)

[0089] H-Aib-HGTFTSDLSKLKEEQRQKEFIEWLKA-dAla-GHPS-Aib-KPPPK (SEQ ID NO: 23)

[0090] H-Aib-HGTFTSDLSKLKEEQRQKEFIEWLKA-dAla-GHPS-Aib-KPPPK (SEQ ID NO: 23)

[0091] H-Aib-HGTFTSDLSKLKEEQRQKEFIEWLKA-dAla-GHPS-Aib-KPPPK (SEQ ID NO: 23)

[0092] H-Aib-HGTFTSDLSKLKEEQRQKEFIEWLKA-dAla-GHPS-Aib-KPPPK (SEQ ID NO: 23)

[0093] H-Aib-HGTFTSDLSKLKEEQRQKEFIEWLKA-dAla-GHPS-Aib-KPPPK (SEQ ID NO: 23)

[0094] H-Aib-HGTFTSDLSKLKEEQRQKEFIEWLKA-dAla-GHPS-Aib-KPPPK (SEQ ID NO: 23)

[0095] H-Aib-HGTFTSDLSKLKEEQRQKEFIEWLKA-dAla-GHPS-Aib-KPPPK (SEQ ID NO: 23)

[0096] H-Aib-HGTFTSDLSKLKEEQRQKEFIEWLKA-dAla-GHPS-Aib-KPPPK (SEQ ID NO: 23)

[0097] H-Aib-HGTFTSDLSKLKEEQRQKEFIEWLKA-dAla-GHPS-Aib-KPPPK (SEQ ID NO: 23)

[0098] Preferably, the polypeptide or pharmaceutically acceptable salt thereof has a C-terminal carboxylic acid group or an amide group, more preferably a C-terminal amide group.

[0099] In some embodiments, the pharmaceutical composition described in the present disclosure, the polypeptide or pharmaceutically acceptable salt thereof comprises any one of the following structures:

[0100] Compound 1,

[0101] Compound 2,

[0102] Compound 3,

[0103] Compound 4,

[0104] Compound 5,

[0105] Compound 6,

[0106] Compound 7,

[0107] Compound 8,

[0108] Compound 9,

[0109] Compound 10,

[0110] Compound 11,

[0111] Compound 12,

[0112] Compound 13,

[0113] Compound 14,

[0114] Compound 15,

[0115] Compound 16,

[0116] Compound 17,

[0117] Compound 18,

[0118] Compound 19,

[0119] Compound 20,

[0120] Compound 21,

[0121] Compound 22,

[0122] Compound 23,

[0123] Compound 24,

[0124] Compound 25,

[0125] Compound 26,

[0126] Compound 27,

[0127] Compound 28,

[0128] Compound 29,

[0129] Compound 30,

[0130] Compound 31,

[0131] Compound 32,

[0132] Compound 33,

[0133] Compound 34,

[0134] Compound 35,

[0135] Compound 36,

[0136] Compound 37,

[0137] Compound 38,

[0138] Compound 39,

[0139] Compound 40,

[0140] Compound 41,

[0141] Compound 42,

[0142] Compound 43,

[0143] Compound 44,

[0144] Compound 45,

[0145] Compound 46,

[0146] Compound 47,

[0147] Compound 48,

[0148] Compound 49,

[0149] Compound 50,

[0150] Compound 51,

[0151] Compound 52,

[0152] Compound 53,

[0153] In some embodiments, the compound comprises (or is) any one of the following structures:

[0154] In some embodiments, the pharmaceutical composition of the present disclosure is selected from:

[0155] 1) the foregoing polypeptide or pharmaceutically acceptable salt thereof, e.g., comprising the structure of any one of Compounds 1-53;

[0156] 2) a divalent zinc ion, e.g., zinc acetate or zinc chloride;

[0157] 3) a buffer, e.g., a phosphate buffer, e.g., sodium phosphate monobasic;

[0158] 4) an osmotic pressure adjusting agent, e.g., sodium chloride, propylene glycol, mannitol, glycerol;

[0159] 5) optionally, an antimicrobial agent, e.g., phenol.

[0160] In some embodiments, the pharmaceutical composition of the present disclosure is selected from:

[0161] 1) the foregoing polypeptide or pharmaceutically acceptable salt thereof, e.g., comprising the structure of any one of Compounds 1-53;

[0162] 2) a divalent zinc ion, e.g., zinc acetate;

[0163] 3) a buffer, e.g., sodium phosphate monobasic;

[0164] 4) an osmotic pressure adjusting agent, such as mannitol;

[0165] 5) optionally an antimicrobial agent, such as phenol.

[0166] In some embodiments, the pharmaceutical composition described in the present disclosure is selected from:

[0167] 1) the aforementioned polypeptide or pharmaceutically acceptable salt thereof, such as comprising the structure shown in any one of compounds 1-53;

[0168] 2) a divalent zinc ion, such as zinc acetate or zinc chloride;

[0169] 3) a buffer, such as a phosphate buffer, such as sodium phosphate monobasic;

[0170] 4) an osmotic pressure adjusting agent, such as sodium chloride, propylene glycol, mannitol, glycerol;

[0171] 5) optionally an antimicrobial agent, such as phenol;

[0172] 6) a pH adjusting agent, such as sodium hydroxide and / or hydrochloric acid;

[0173] 7) water for injection.

[0174] In some embodiments, the pharmaceutical composition described in the present disclosure is selected from:

[0175] 1) the aforementioned polypeptide or pharmaceutically acceptable salt thereof, such as comprising the structure shown in any one of compounds 1-53;

[0176] 2) a divalent zinc ion, such as zinc acetate;

[0177] 3) a buffer, such as sodium phosphate monobasic;

[0178] 4) an osmotic pressure adjusting agent, such as mannitol;

[0179] 5) optionally an antimicrobial agent, such as phenol;

[0180] 6) a pH adjusting agent, such as sodium hydroxide and / or hydrochloric acid;

[0181] 7) water for injection.

[0182] In some embodiments, the pharmaceutical composition described in the present disclosure is selected from:

[0183] 1) the aforementioned polypeptide or pharmaceutically acceptable salt thereof, such as comprising the structure shown in any one of compounds 1-53;

[0184] 2) a divalent zinc ion, such as zinc chloride;

[0185] 3) a buffer, such as sodium phosphate monobasic;

[0186] 4) an osmotic pressure adjusting agent, such as mannitol;

[0187] 5) Optional antimicrobial agent, such as phenol;

[0188] 6) pH adjusting agent, such as sodium hydroxide and / or hydrochloric acid;

[0189] 7) Water for injection.

[0190] In some embodiments, the polypeptide or pharmaceutically acceptable salt thereof in the pharmaceutical composition of the present disclosure comprises the structure shown in Compound 1.

[0191] In some embodiments, the polypeptide or pharmaceutically acceptable salt thereof in the pharmaceutical composition of the present disclosure is selected from the structure shown in Compound 1.

[0192] In some embodiments, the pharmaceutical composition of the present disclosure can be used for treating a patient in need of such treatment by parenteral administration. The parenteral administration route can be selected from subcutaneous injection, intramuscular injection or intravenous injection.

[0193] In some embodiments, the pharmaceutical composition of the present disclosure has a hydrolytic impurity content of no more than 5%, preferably no more than 3%, preferably no more than 2%, preferably no more than 1% after being placed under high temperature conditions for 10 days.

[0194] In some embodiments, the pharmaceutical composition of the present disclosure has a hydrolytic impurity content of no more than 10%, preferably no more than 6%, preferably no more than 5%, preferably no more than 4%, preferably no more than 3%, preferably no more than 2%, preferably no more than 1.5% after being placed under high temperature conditions for 1 month.

[0195] In some embodiments, the pharmaceutical composition of the present disclosure has a hydrolytic impurity content of no more than 5%, preferably no more than 3%, preferably no more than 2%, preferably no more than 1% of N-terminal His-Aib cleavage after being placed under high temperature conditions for 10 days.

[0196] In some embodiments, the pharmaceutical composition of the present disclosure has a hydrolytic impurity content of N-terminal His-Aib cleavage of no more than 10% when placed under high temperature conditions for 1 month, preferably no more than 6%, preferably no more than 5%, preferably no more than 4%, preferably no more than 3%, preferably no more than 2%, preferably no more than 1.5%.

[0197] In some embodiments, the pharmaceutical composition of the present disclosure has a total impurity content of no more than 15% when placed under high temperature conditions for 10 days, preferably no more than 12%, preferably no more than 10%, preferably no more than 9%, preferably no more than 8%, preferably no more than 7%, preferably no more than 6%, preferably no more than 5%, preferably no more than 4%, preferably no more than 3%.

[0198] In some embodiments, the pharmaceutical composition of the present disclosure has a total impurity content of no more than 15% when placed under high temperature conditions for 1 month, preferably no more than 12%, preferably no more than 10%, preferably no more than 9%, preferably no more than 8%, preferably no more than 7%, preferably no more than 6%, preferably no more than 5%, preferably no more than 4%, preferably no more than 3%.

[0199] In some embodiments, the high temperature condition of the present disclosure is 25°C, 30°C or 40°C; preferably the high temperature condition is 30°C or 40°C.

[0200] The present disclosure also provides a method for preparing a lyophilized formulation comprising a polypeptide or a pharmaceutically acceptable salt thereof, which comprises the step of freeze-drying the aforementioned pharmaceutical composition. In alternative embodiments, the freeze-drying comprises the steps of pre-freezing, primary drying and secondary drying, in sequence.

[0201] The present disclosure also provides a lyophilized formulation comprising a polypeptide or a pharmaceutically acceptable salt thereof, which is prepared by the aforementioned method for preparing a lyophilized formulation comprising a polypeptide or a pharmaceutically acceptable salt thereof.

[0202] The present disclosure also provides a method of reconstituting a lyophilized formulation containing a polypeptide or a pharmaceutically acceptable salt thereof, comprising the step of reconstituting the aforementioned lyophilized formulation with a solution selected from, but not limited to, water for injection, physiological saline or dextrose solution.

[0203] The present disclosure also provides a method of reconstituting a lyophilized formulation containing a polypeptide or a pharmaceutically acceptable salt thereof, comprising the step of reconstituting the aforementioned lyophilized formulation with a solution selected from, but not limited to, water for injection, physiological saline or dextrose solution.

[0204] The present disclosure further provides an article of manufacture or a kit comprising a container filled with any of the stable pharmaceutical compositions described herein. In some embodiments, the glass bottle is a Type I borosilicate glass tube manufactured as a vial for injection.

[0205] The present disclosure also provides an article of manufacture comprising a container filled with the aforementioned pharmaceutical composition or a lyophilized formulation or a reconstituted solution of a lyophilized formulation.

[0206] The present disclosure also provides a pharmaceutical composition or a lyophilized formulation or a reconstituted solution of a lyophilized formulation for use in a method of treatment and prevention of a disease or a disorder.

[0207] The present disclosure also provides the use of the aforementioned pharmaceutical composition or a lyophilized formulation or a reconstituted solution of a lyophilized formulation in the manufacture of a medicament for the treatment and / or prevention of a disease or a disorder.

[0208] The present disclosure also provides a method of treatment and prevention of a disease or a disorder, comprising administering to a patient in need thereof a therapeutically effective amount of the aforementioned pharmaceutical composition or a lyophilized formulation or a reconstituted solution of a lyophilized formulation.

[0209] The present disclosure provides the use of a pharmaceutical composition in the manufacture of a medicament for the treatment of non-insulin dependent diabetes mellitus, insulin dependent diabetes mellitus, obesity, non-alcoholic fatty liver, hepatic steatosis, diabetic retinopathy, diabetic neuropathy, diabetic nephropathy, insulin resistance, dyslipidemia associated with insulin resistance, and / or diabetic dyslipidemia.

[0210] The present disclosure provides a pharmaceutical composition for use in the treatment of non-insulin dependent diabetes mellitus / Type II diabetes mellitus, insulin dependent diabetes mellitus, obesity, non-alcoholic fatty liver, hepatic steatosis, dyslipidemia associated with insulin resistance, and / or diabetic dyslipidemia.

[0211] The present disclosure provides a method for the treatment of non-insulin dependent diabetes mellitus / Type II diabetes mellitus, insulin dependent diabetes mellitus, obesity, non-alcoholic fatty liver, hepatic steatosis, dyslipidemia associated with insulin resistance, and / or diabetic dyslipidemia, comprising administering to a subject in need thereof a pharmaceutical composition of the present disclosure.

[0212] Definitions

[0213] For the purposes of the present disclosure, certain technical and scientific terms are specifically defined below. Unless specifically defined herein, all other technical and scientific terms used in the present disclosure have the meanings commonly understood by one of ordinary skill in the art to which the present disclosure pertains.

[0214] The amino acid sequences of the present disclosure contain the standard one-letter or three-letter codes for the twenty amino acids, and all amino acid residues in the present disclosure are preferably in the L-form, unless explicitly stated otherwise. In addition, Aib is alpha amino isobutyric acid, D-Ala is D-form alanine, Orn is ornithine, Dap is 2,3-diaminopropionic acid, and Dab is 2,4-diaminobutyric acid.

[0215] The term "agonist" is defined as a substance that has an activating effect on the GLP-1 receptor or on the GIP receptor.

[0216] The term "triple agonist of the GLP-1 receptor, the GIP receptor and the GCG receptor" as used in the context of the present disclosure refers to a substance or ligand that can activate the GLP-1 receptor, the GIP receptor and the GCG receptor.

[0217] In the present disclosure, the term "treatment" includes inhibiting, slowing, stopping or reversing the progression or severity of an existing symptom or condition.

[0218] The term "alkyl" refers to saturated aliphatic hydrocarbon groups, which are straight-chain or branched-chain groups containing 1 to 20 carbon atoms, such as alkyl groups containing 1 to 8 carbon atoms, such as alkyl groups of 1 to 6 carbon atoms, such as alkyl groups of 1 to 3 carbon atoms. Non-limiting examples include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, t-butyl, sec-butyl, n-pentyl, 1,1-dimethylpropyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, 1-ethyl-2-methylpropyl, 1,1,2-trimethylpropyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 2,2-dimethylbutyl, 1,3-dimethylbutyl, 2-ethylbutyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 2,3-dimethylbutyl, n-heptyl, 2-methylhexyl, 3-methylhexyl, 4-methylhexyl, 5-methylhexyl, 2,3-dimethylpentyl, 2,4-dimethylpentyl, 2,2-dimethylpentyl, 3,3-dimethylpentyl, 2-ethylpentyl, 3-ethylpentyl, n-octyl, 2,3-dimethylhexyl, 2,4-dimethylhexyl, 2,5-dimethylhexyl, 2,2-dimethylhexyl, 3,3-dimethylhexyl, 4,4-dimethylhexyl, 2-ethylhexyl, 3-ethylhexyl, 4-ethylhexyl, 2-methyl-2-ethylpentyl, 2-methyl-3-ethylpentyl, n-nonyl, 2-methyl-2-ethylhexyl, 2-methyl-3-ethylhexyl, 2,2-diethylpentyl, n-decyl, 3,3-diethylhexyl, 2,2-diethylhexyl, and various branched isomers thereof, and the like. Alkyl groups may, for example, be lower alkyl groups containing 1 to 6 carbon atoms, non-limiting examples including methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, t-butyl, sec-butyl, n-pentyl, 1,1-dimethylpropyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, 1-ethyl-2-methylpropyl, 1,1,2-trimethylpropyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 2,2-dimethylbutyl, 1,3-dimethylbutyl, 2-ethylbutyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 2,3-dimethylbutyl, and the like. Alkyl groups can be substituted or unsubstituted, when substituted, the substituents can be substituted at any accessible attachment point, which can be one or more groups independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halo, thiol, hydroxyl, nitro, cyano, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, cycloalkoxy, heterocycloalkoxy, cycloalkylthio, heterocycloalkylthio, oxo, carboxyl, or carboxylate. Substituted alkyl groups of the present disclosure can be methyl, ethyl, isopropyl, t-butyl, haloalkyl, deuterated alkyl, alkoxy-substituted alkyl, or hydroxyl-substituted alkyl.

[0219] The term "native GLP-1" refers to a naturally occurring molecule of the glucagon family of peptides or the exendin family of peptides, wherein: the glucagon family of peptides is encoded by the preproglucagon gene and includes three small peptides with high homology, i.e., glucagon (1-29), GLP-1 (1-37), and GLP-2 (1-33); the exendin is a peptide expressed in lizards and, like GLP-1, is insulinotropic. In some embodiments, the term "native GLP-1" also refers to human GLP-1 (7-37) and human GLP-1 (7-36).

[0220] The amino acid "substitution" as described in the present disclosure refers to the substitution of one amino acid residue with a different one.

[0221] The term "fatty acid" refers to a carboxylic acid with a long aliphatic tail (chain) that can be saturated or unsaturated; a fatty acid in the present disclosure is a carboxylic acid with a C4-C30 straight chain or branched aliphatic group.

[0222] The term "peptide" as used in the present disclosure refers to a sequence of two or more amino acids. The term "peptide" encompasses the category of peptides with modified amino and carboxy termini. A "peptide" can comprise a natural amino acid sequence, or be modified. For example, where one or more amino acid residues are substituted with other amino acid residues, and / or one or more amino acid residues are deleted from the peptide, and / or one or more amino acid residues are added to the peptide, and / or one or more amino acid residues are linked to a substituent. Such modifications of amino acid residues can occur at any position in the N- and / or C-terminus and / or in the middle of the peptide. For example, an amino acid chain comprising a terminal carboxylic acid replaced with an amide group is included within the amino acid sequence designated as a natural amino acid.

[0223] The term "amino acid" is a molecule containing an amino group and a carboxylic acid group, and optionally containing one or more additional groups often referred to as side chains. Amino acids include proteinogenic (or natural) amino acids (of which there are 20 standard amino acids) as well as non-proteinogenic (or unnatural) amino acids. Proteinogenic amino acids are amino acids that are naturally incorporated into proteins. Standard amino acids are those encoded by the genetic code. Non-proteinogenic amino acids either do not occur in proteins or are not produced by standard cellular machinery (e.g., they can have undergone post-translational modification). Non-limiting examples of non-proteinogenic amino acids are Aib (alpha-aminoisobutyric acid or 2-amino isobutyric acid), norleucine (Nle), norvaline, and D-isomers of proteinogenic amino acids. D-Ala is D-form of alanine, -aL- is alpha position methylated modified leucine, aS is alpha position methylated modified serine, aK is alpha position methylated modified lysine, etc.

[0224] "Naturally occurring amino acid" refers to the 20 conventional amino acids, i.e., alanine (A), cysteine (C), aspartic acid (D), glutamic acid (E), phenylalanine (F), glycine (G), histidine (H), isoleucine (I), lysine (K), leucine (L), methionine (M), asparagine (N), proline (P), glutamine (Q), arginine (R), serine (S), threonine (T), valine (V), tryptophan (W), and tyrosine (Y).

[0225] "Non-naturally occurring amino acid" refers to an amino acid that is not naturally encoded or found in the genetic code of any organism. They can be, for example, purely synthetic compounds. Examples of non-naturally occurring amino acids include, but are not limited to, hydroxyproline, gamma-carboxyglutamate, O-phosphoserine, azetidinecarboxylic acid, 2-aminoadipic acid, 3-aminoadipic acid, beta-alanine, aminopropionic acid, 2-aminobutyric acid, 4-aminobutyric acid, 6-aminohexanoic acid, 2-aminoheptanoic acid, 2-aminoisobutyric acid, 3-aminoisobutyric acid, 2-aminoheptanedioic acid, t-butylglycine, 2,4-diaminoisobutyric acid (Dap), desmosine, 2,2'-diaminopimelic acid, 2,3-diaminopropionic acid (Dab), N-ethylglycine, N-methylglycine, N-ethylasparagine, homoproline, hydroxylysine, allo-hydroxylysine, 3-hydroxyproline, 4-hydroxyproline, isodesmosine, allo-isoleucine, N-methylalanine, N-methylglycine, N-methylisoleucine, N-methylpentylglycine, N-methylvaline, naphthalanine, norvaline, norleucine, ornithine (Orn), D-ornithine, D-arginine, p-aminophenylalanine, pentylglycine, pipecolic acid, and thioproline. In addition, C-terminal carboxyl groups, N-terminal amino groups, and / or side chain functional groups of the naturally occurring or non-naturally occurring amino acids are chemically modified.

[0226] The hydrogen atoms described in the present disclosure can be replaced by isotopes thereof (protium, deuterium, tritium), and any hydrogen atom in the compounds of the present disclosure described in the present disclosure can also be replaced by an isotope.

[0227] "Optional" or "optionally" means that the subsequently described event or circumstance can or can not occur, and thus the description includes instances where the event or circumstance occurs and instances where it does not. For example, "heterocyclic group optionally substituted with alkyl" means that alkyl can or can not be present, and the description includes instances where the heterocyclic group is substituted with alkyl and instances where the heterocyclic group is not substituted with alkyl.

[0228] "Substituted" means that one or more hydrogen atoms, preferably up to five, more preferably one to three, of the group are independently of each other replaced by a substituent. Substituents are only in their possible chemical positions, which can or cannot be possible (experimentally or theoretically) by one skilled in the art without undue effort. For example, an amino or hydroxyl group with a free hydrogen can be unstable when bound to a carbon atom with an unsaturated (e.g., olefinic) bond.

[0229] "Pharmaceutical composition" means a mixture of one or more of the compounds described herein or physiologically / pharmaceutically acceptable salts or prodrugs thereof with other chemical components, such as physiologically / pharmaceutically acceptable carriers and excipients. The purpose of a pharmaceutical composition is to facilitate administration of the active ingredient to a biological entity and to facilitate absorption of the active ingredient to thereby elicit a biological activity.

[0230] "Buffer" means a buffering agent that resists changes in pH through the action of its acid-base conjugate components. Examples of buffers that control pH in an appropriate range include acetate, succinate, citrate, phosphate, gluconate, histidine, oxalate, lactate, phosphate, citrate, tartrate, fumarate, glycylglycine, and other organic acid buffers.

[0231] "Histidine salt buffer" is a buffer that includes a histidine ion. Examples of histidine salt buffers include histidine-hydrochloric acid, histidine-acetic acid, histidine-phosphoric acid, histidine-sulfuric acid, and the like, such as a histidine-acetic acid buffer or a histidine-hydrochloric acid buffer, a histidine-acetic acid buffer being histidine formulated with acetic acid, and a histidine-hydrochloric acid buffer being histidine formulated with hydrochloric acid.

[0232] "Citrate buffer" is a buffer that includes a citrate ion. Examples of citrate buffers include citric acid-sodium citrate, citric acid-potassium citrate, citric acid-calcium citrate, citric acid-magnesium citrate, and the like. The citrate buffer can be citric acid-sodium citrate.

[0233] "Succinate buffer" is a buffer that includes a succinate ion. Examples of succinate buffers include succinic acid-sodium succinate, succinic acid-potassium succinate, succinic acid-calcium succinate, and the like. The succinate buffer can be succinic acid-sodium succinate.

[0234] "Phosphate buffer" is a buffer that includes a phosphate ion. Examples of phosphate buffers include disodium hydrogen phosphate-sodium dihydrogen phosphate, disodium hydrogen phosphate-potassium dihydrogen phosphate, disodium hydrogen phosphate-citric acid, and the like. The phosphate buffer can be disodium hydrogen phosphate-sodium dihydrogen phosphate.

[0235] "Acetate buffer" is a buffer that includes acetate ions. Examples of acetate buffers include sodium acetate, histidine acetate, potassium acetate, calcium acetate, magnesium acetate, etc. Acetate buffers can be sodium acetate.

[0236] "Pharmaceutical composition" means a mixture containing one or more of the compounds described herein or their physiologically / pharmacologically acceptable salts or prodrugs, along with other chemical components, such as physiologically / pharmacologically acceptable carriers and excipients. The purpose of a pharmaceutical composition is to facilitate administration to a living organism, thereby promoting the absorption of the active ingredient and its biological activity. In this document, "pharmaceutical composition" and "formulation" are used interchangeably.

[0237] Unless otherwise specified, the solvent in the solution form of the pharmaceutical compositions described in this disclosure is water.

[0238] "Lyophilized formulation" refers to a formulation or pharmaceutical composition obtained by a vacuum freeze-drying step after a liquid or solution form of a pharmaceutical composition or solution preparation.

[0239] The three-letter and single-letter codes for amino acids used in this disclosure are as described in J. biol. chem, 243, p3558 (1968).

[0240] The values ​​in this disclosure are instrument measurements or calculated values ​​after instrument measurement, and are subject to a certain degree of error. Generally speaking, ±10% is within the reasonable error range. Of course, the context in which the value is used needs to be considered. For example, for the content of total impurities, the value is defined as having an error variation of no more than ±10% after measurement, and can be ±9%, ±8%, ±7%, ±6%, ±5%, ±4%, ±3%, ±2%, or ±1%, preferably ±5%. Attached Figure Description

[0241] Figure 1 shows the plasma concentration-time curves of pharmacokinetic data for prescriptions 26 and 27 in rats. Detailed Implementation

[0242] The present disclosure is further described below with reference to embodiments, but these embodiments are not intended to limit the scope of the present disclosure.

[0243] Experimental methods in the embodiments of this disclosure that do not specify specific conditions are generally performed under conventional conditions or as recommended by the raw material or product manufacturer. Reagents whose specific source is not specified are commercially available conventional reagents.

[0244] Example 1: Preparation of compound 0120

[0245] Chemical synthesis of compound 0120 was accomplished on a Prelude-X automated peptide synthesizer using fluorenylmethyloxycarbonyl (Fmoc) / tert-butyl (t-Bu) synthesis strategy. Rink-amide MBHA resin with a substitution of 0.54 mmole / g was used. Fmoc-L-Lys(Mtt)-OH was used for the attachment of the fatty acid modification site. Boc-L-His(Trt)-OH was used for the N-terminal amino acid. Fmoc group was removed using 20% 4-methylpiperidine in DMF (2 times for 8 min each) before the amino acid condensation step. All standard amino acid condensations were performed using equimolar ratio of Fmoc amino acid, HATU and 2 equivalents of 4-methylmorpholine at room temperature for 25 min with 10 equivalents of excess theoretical peptide loading. Exceptions were condensations to C-alpha-methylated amino acids, which required two or three condensations for 60 min each to ensure complete condensation. After the completion of the above peptide-resin extension, the resin peptide was washed with dichloromethane and then treated with hexafluoroisopropanol / dichloromethane mixture (30% hexafluoroisopropanol, 10 mL) for 45 min at room temperature. The reaction was drained and then treated with hexafluoroisopropanol / dichloromethane mixture (30%, 10 mL) for 45 min at room temperature. The reaction was drained and the resin was washed with DMF three times. Additional coupling / deprotection cycles to extend the lysine side chain using Fmoc / tBu solid phase synthesis strategy on the same automated peptide synthesizer involved Fmoc-AEEA-OH (2 times), Fmoc-L-Glu-OtBu (1 time) and eicosanedioic acid mono-t-butyl ester (1 time). Condensations were performed using equimolar ratio of Fmoc amino acid, HATU and 2 equivalents of 4-methylmorpholine at room temperature for 25 min with 10 equivalents of excess theoretical peptide loading. An exception was the condensation of eicosanedioic acid mono-t-butyl ester, which required at least 3 hours of condensation at room temperature to ensure complete condensation.

[0246] After the synthesis is completed, the obtained resin peptide is washed with DMF, DCM for 3 times, and then vacuum dried. Then, fresh prepared cleavage solution 10 mL (trifluoroacetic acid: triisopropylsilane: water = 90:5:5, v:v:v) is added, and the reaction is oscillated at room temperature for 3-4 hours. After the reaction is completed, the filtrate is filtered, and the resin is washed with trifluoroacetic acid for 2 times. Then, the filtrate is combined, and a large amount of methyl tert-butyl ether is added to precipitate the crude peptide solid. After centrifugation to remove the supernatant, the crude peptide with the number 0120 is obtained. The crude peptide is dissolved in a mixed solvent containing 20% acetic acid / water, filtered through a 0.22 um membrane, and then separated by a WATERS Prep150 LC reverse phase high performance liquid chromatography system. The mobile phase is A (0.1% trifluoroacetic acid, 10% acetonitrile, aqueous solution) and B (0.1% trifluoroacetic acid, 90% acetonitrile, aqueous solution). The chromatographic column is X-SELECT OBD C-18 (WATERS) reverse phase chromatographic column, and the detection wavelength of the chromatograph is set to 220 nm during the purification process, and the flow rate is 15-20 mL / min. After the product related fractions are collected and freeze-dried, the pure polypeptide of compound number 0120 is obtained. The purity and compound identity of the polypeptide pure product are determined by analytical ultra-high performance liquid chromatography and liquid chromatography / mass spectrometry, and the purity is 98.03%, and the measured molecular weight of the compound is 4854.2.

[0247] Example 2: Biological activity test of compound 0120

[0248] The purpose of this example is to detect the agonist activity of the test compound 0120 on human glucagon-like peptide-1 receptor, human glucose-dependent insulinotropic polypeptide receptor and human glucagon receptor.

[0249] 2.1 Experimental method:

[0250] Functional activity was determined in CHO-K1 clonal cell lines expressing GIPR, GLP-1R and GCGR. In a 10 pL assay volume, polypeptides were diluted (8-10 points, 10-fold dilutions) in DMEM / F12 1:1 (HyClone, SH30023.01) supplemented with 5% FBS (Fetal Bovine Serum, Gibco, 10099141), 0.1% Casein (Sigma, C4765), 0.25 mM IBMX (Selleck, S5836). SV 96-well plates (Cisbio, 66PL96025) were seeded with the respective receptor cells (5000 cells / well) and incubated with the configured polypeptides under incubation conditions of 37°C for 30 minutes. The resulting increase in intracellular cAMP was quantitatively determined using the cAMP-GS dynamic HTRF assay kit (Cisbio, 62AM4PEJ). Briefly, intracellular cAMP levels were detected by adding cAMP-d2 conjugate in cell lysis buffer followed by the addition of the antibody anti-cAMP-Eu cryptate (also in cell lysis buffer). The resulting competition assay was incubated for at least 60 minutes at room temperature and then detected using a Tecan Infinite F Plex instrument with excitation at 320 nm and emission at 665 nm and 620 nm. The measured data (emission at 665 nm / 620 nm * 10000) is inversely proportional to the amount of cAMP present and converted to cAMP content per well using a cAMP standard curve.

[0251] The amount of cAMP generated per well was converted to percent of the maximal response observed with human GLP-1 (7-36)-NH2, human GCG (1-29) or human GIP (1-42)-NH2 (purchased from sigma). Using the percent maximal response vs. added polypeptide concentration, relative EC50 values were derived by nonlinear regression analysis, fitted into a four-parameter logistic equation. See Table 1 below for specific data. LY3437943 molecule see WO2019 / 125938 Example 12, the aforementioned patent is incorporated by reference in its entirety into the present application.

[0252] Table 1

[0253] Example 3. Detection of the pharmacokinetic properties of the compounds in mice

[0254] Male C57 mice were used as test animals, and the pharmacokinetic characteristics of compound 0120 in C57 mice (plasma) were detected after a single intravenous injection of the target compound.

[0255] 3.1 Test method:

[0256] Male C57 mice, body weight about 25 g, 4-6 weeks old. After the polypeptide compound solution was prepared using 1xPBS dissolving solution, the administration was completed by intravenous injection at a dose of 200 nmol / kg, and 3 animals were administered for each compound. At 5 min, 15 min, 0.5 h, 1 h, 2 h, 4 h, 6 h, 24 h, 48 h and 72 h time points after the end of infusion, 60 μL of blood was collected and transferred to a centrifuge tube containing EDTA-K2 anticoagulant. The whole blood sample was centrifuged at 4000g for 5 min at 4°C, and the plasma was separated and stored at -80°C.

[0257] Sample pretreatment method: take 30 μL of the plasma sample, add 120 μL of 0.1% formic acid methanol solution containing 1 ng / mL of internal standard verapamil, vortex for 5 min, centrifuge at low temperature 10000 rpm for 10 min, take 70 μL of supernatant, add 70 μL of water, mix well, and analyze by LC-MS instrument.

[0258] LC-MS analysis method: (1) Chromatographic conditions: mobile phase A and B are 0.1% formic acid solution and 0.1% formic acid acetonitrile solution, respectively; gradient elution mode is used; flow rate is 0.5 mL / min; injection volume is 10 μL; chromatographic column is nanomicro Unisil C18aq (4.6 mm x 150 mm, 5 μm); column temperature: 40°C. (2) Mass spectrometry conditions: mass spectrometry uses an electrospray ion source (ESI), positive ion analysis mode, and multiple reaction monitoring (MRM) scanning mode.

[0259] Data processing uses SCIEX OS (Version 2.1.6) software to quantify blood drug concentration data, and PKSolver (Version 2.0) software to calculate pharmacokinetic parameters.

[0260] 3.2 Test results

[0261] Table 2. Comparison of pharmacokinetic parameters of administered compounds (200 nmol / kg) in mice

[0262] Example 4: Preparation process of pharmaceutical composition

[0263] The pharmaceutical composition of the present disclosure can be prepared according to the following process:

[0264] Preparation of mother liquor: take an appropriate amount of water for injection, add an appropriate amount of buffer salt solution, weigh the prescription amount of bacteriostatic agent and osmotic pressure regulator into the above solution, and stir until dissolved. Weigh the prescription amount of API-0120 molecule into the above solution, stir until dissolved, and add an appropriate amount of water for injection.

[0265] Preparation of the formulation: Take an appropriate amount of mother liquor, add the required amount of metal ion solution according to the prescription, adjust the pH with 1M NaOH solution or 0.5M HC1 solution, and add water for injection to constant volume.

[0266] Filtration and filling: filter the prescription with 0.22 μm PVDF filter membrane into 2 mL siren bottles, 1 mL per bottle.

[0267] Table 3 Formulation prescription information (unit dose)

[0268] Example 5: Stability investigation of pharmaceutical composition

[0269] The formulations 1-5 prepared in Example 4 were placed under high temperature conditions for 1 month to investigate the chemical stability. The prescription samples were detected by HPLC for related substances and by SEC for HMPW. The results are shown in Table 4.

[0270] The HPLC detection used Agilent AdvanceBio Peptide Map, 4.6*150mm, 2.7 μm as the chromatographic column, the detection wavelength was 220 nm, and the mobile phase was: mobile phase A: 50 mM sodium sulfate (0.1% perchloric acid), mobile phase B: 80% acetonitrile (0.2% perchloric acid).

[0271] The SEC detection used Sepax Zenix SEC-80 (7.8*300mm, 3 μm) as the chromatographic column, the detection wavelength was 280 nm, and the mobile phase was 50% ACN (0.1% TFA).

[0272] Table 4 Formulation 3-5 chemical stability results

[0273] The ASAP stability prediction results show that the hydrolytic impurities produced by the N-terminal His-Aib breakage are low-activation-energy sensitive, and the increase in temperature will significantly increase the chemical instability of the impurities.

[0274] From the results of the high-temperature accelerated test in Table 4, it can be seen that by adding zinc ions in the prescription, the growth of hydrolytic impurities (RRT 1.08) and aggregates (HMPW) in the compound is slowed down, and the total degradation of impurities is reduced, which meets the requirements for impurities in the product at the end of the shelf life in clinical use.

[0275] Example 6: Stability investigation of pharmaceutical composition with different metal ions

[0276] The effect of different metal ions on the stability of the pharmaceutical composition was investigated according to the preparation method and prescription amount shown in prescription 4 in Example 4 (API-0120: 12 mg / mL, disodium hydrogen phosphate buffer: 10 mM, phenol: 5.5 mg / mL, mannitol: 46 mg / mL, pH 7.5).

[0277] Table 5 Formulation prescription information (zinc ion)

[0278] Table 6 Formulation prescription information (magnesium ion)

[0279] Table 7 Formulation prescription information (calcium ion, iron ion)

[0280] The initial impurity levels of formulations 6-23 prepared using the same batch of API were the same, with total impurities of 3.15%, hydrolytic impurities of 0.62%, and HMPW of 0.11%. As can be seen from the results in Table 8, the chemical stability of the formulation prescription was more excellent when using divalent zinc ions compared to other divalent metal ions.

[0281] Table 8 Chemical stability results

[0282] Example 7: In vitro activity evaluation of zinc-containing formulations

[0283] The effect of the addition of zinc ions on the activity of the API was investigated according to the preparation method shown in prescription 4 in Example 4.

[0284] Table 9 Formulation prescription information

[0285] In vitro activity tests were performed on prescription 24 and prescription 25, and the results showed that the in vitro activity of the three target points of prescription 25 did not have a significant effect compared to prescription 24, indicating that the addition of zinc ions did not affect the activity of the terminal amino acids.

[0286] Example 8: Investigation of the PK behavior of zinc-containing formulations in rats

[0287] The difference in the PK behavior of zinc ions in rats was investigated by comparing the addition of zinc ions according to the preparation method shown in prescription 4 in Example 4. Six to seven-week-old male SD rats were used as test animals, and the target formulations were injected subcutaneously for a single dose, and the pharmacokinetic characteristics of compound 0120 in SD rats (plasma) were detected.

[0288] Table 10

[0289] 8.1 Test method:

[0290] Male SD rats, body weight 200-250 g, 6-7 weeks old. According to the dose of 1060 nmol / kg, the subcutaneous injection was completed, and 3 animals were administered for each compound. At 5 min, 15 min, 0.5 h, 1 h, 2 h, 4 h, 6 h, 24 h, 48 h, 72 h, 96 h and 168 h after administration, 500 μL of blood was collected and transferred to a centrifuge tube containing EDTA-K2 anticoagulant. The whole blood sample was centrifuged at 4000g for 5 min at 4°C, and the plasma was separated and stored at -80°C.

[0291] Sample pretreatment method: take 30 μL of plasma sample, add 120 μL of 0.1% formic acid methanol solution containing 1 ng / mL internal standard verapamil, vortex for 5 min, centrifuge at low temperature 10000 rpm for 10 min, take 70 μL of supernatant, add 70 μL of water, mix well, and analyze by LC-MS instrument.

[0292] LC-MS analysis method: (1) Chromatographic conditions: mobile phase A and B are 0.1% formic acid solution and 0.1% formic acid acetonitrile solution respectively; gradient elution mode is used; flow rate is 0.5 mL / min; injection volume is 10 μL; chromatographic column is nanomicro Unisil C18aq (4.6 mm x 150 mm, 5 μm); column temperature: 40°C. (2) Mass spectrometry conditions: mass spectrometry uses electrospray ion source (ESI), positive ion analysis mode, and multiple reaction monitoring (MRM) scanning mode.

[0293] Data processing uses SCIEX OS (Version 2.1.6) software to quantitate blood drug concentration data.

[0294] The results show that the overall PK behavior of the zinc-containing preparation has no significant difference.

[0295] Example 9: Characterization of the high-level structure of 0120

[0296] Referring to the preparation method shown in prescription 4 in Example 4, the effect of the preparation containing different proportions of zinc ions on the high-level structure of API was investigated.

[0297] Table 11 Formulation prescription information

[0298] Since mannitol and phenol have a large interference in the far ultraviolet region during circular dichroism test, mannitol and phenol and other excipients are not added in the prescription for high-level structure characterization. The circular dichroism characterization of 0120 zinc-containing (10 mM disodium hydrogen phosphate, pH 7.5) / zinc-free (10 mM disodium hydrogen phosphate, pH 7.5) preparation was carried out, and the results are shown in Table 12. The addition of zinc ions does not significantly change the secondary structure under the condition of pH 7.5.

[0299] Table 12 Aggregate detection in different zinc ratio formulations

[0300] Example 10: SAR441255 molecular formulation prescription stability investigation

[0301] Referring to the preparation method shown in prescription 4 in Example 4, the effect of formulations containing different proportions of zinc ions on the chemical stability of API-SAR441255 formulations was investigated. Four groups of different proportions of zinc ions were set (group 1 without adding zinc ions, group 2 API: zinc ion (molar ratio) is 2:1, group 3 API: zinc ion (molar ratio) is 1:1, group 4 API: zinc ion (molar ratio) is 1:2. ) were investigated respectively. The prescription sample was detected by HPLC for related substances, by SEC for HMPW, and SAR441255 structure:

[0302] Table 13 Chemical stability results

[0303] Without zinc prescription accelerated (25 degrees) for 3 months, the total impurities and hydrolytic impurities increased significantly, and the addition of zinc ions in the formulation can significantly reduce the generation of hydrolytic impurities and polymers.

Claims

1. A pharmaceutical composition comprising a polypeptide or a pharmaceutically acceptable salt thereof and a divalent metal ion, The polypeptide comprises an N terminus of an amino acid sequence as shown in X1-Aib-X2; wherein, X1 is a natural or unnatural amino acid residue comprising an imidazole group, and X2 is a natural or unnatural amino acid residue comprising an imidazole group.

2. The pharmaceutical composition according to claim 1, wherein, the polypeptide comprises an N-terminal end of an amino acid sequence as shown in R1-X1-Aib-X2; wherein R1 is hydrogen, X1 is a natural or unnatural amino acid residue comprising an imidazole group, and X2 is a natural or unnatural amino acid residue comprising an imidazole group.

3. The pharmaceutical composition according to claim 1 or 2, wherein, X1 is a His residue, and / or X2 is a His residue; preferably, the polypeptide comprises an N-terminal end of an amino acid sequence as shown in His-Aib-His.

4. The pharmaceutical composition according to any one of claims 1-3, wherein, the polypeptide or the pharmaceutically acceptable salt thereof comprises at least one Lys, Orn, Dap, Dab or Cys residue having a side chain attached substituent; preferably, the polypeptide or the pharmaceutically acceptable salt thereof comprises at least one Lys residue having a side chain attached substituent; preferably, the substituent comprises a structure as shown in -Z1-Z2; wherein, Z1comprises 0-10 residues independently of one another selected from the group consisting of Gly residues, Glu residues, γGlu residues, -C(O)-CH2-(O-CH2-CH2)2-NH-, -C(O)-(C6H 10 -CH2-NH-, -C(O)-CH(NH2)-(CH2)4-NH-; Z2is selected from C 12-32 a fatty acid or C 12-32 alkyl.

5. The pharmaceutical composition according to any one of claims 1-4, wherein, The polypeptide or pharmaceutically acceptable salt thereof comprises at least one Lys residue, and the epsilon-amino group of the Lys residue is linked to any one of the following substituents:

6. The pharmaceutical composition according to any one of claims 1-5, wherein, the polypeptide or the pharmaceutically acceptable salt thereof has agonistic activity on at least one of the following receptors: GLP-1 receptor, GIP receptor, GCG receptor; preferably, the polypeptide or the pharmaceutically acceptable salt thereof is a triple agonist of GLP-1 receptor, GIP receptor and GCG receptor.

7. The pharmaceutical composition according to any one of claims 1-6, wherein, the polypeptide or the pharmaceutically acceptable salt thereof comprises a sequence as shown in: H-Aib-HGTFTSDYSIYLEKQAA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 2) H-Aib-HGTFTSDYSIYLEKQAAQEFVQWLLEGGPSSGAPPPS (SEQ ID NO: 3) H-Aib-HGTFTSDYSIYLEKQYA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 4) H-Aib-HGTFTSDYSYYLEKQAA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 5) H-Aib-HGTFTSDYSIYLEKQAAEEFVQWLLEGGPSSGAPPPS (SEQ ID NO: 6) H-Aib-HGTFTSDYSIYLDKQAA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 7) H-Aib-HGTFTSDYSYYLEKQYA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 8) H-Aib-HGTFTSDYSIYLEKKYA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 9) ​ ​ ​ ​ ​ ​ ​ ​ H-Aib-HGTFTSDYSYYLEKKAA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 10) H-Aib-HGTFTSDYSIYLEKKAA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 11) H-Aib-HGTFTSDYSIYLEKQKA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 12) H-Aib-HGTFTSDYSKYLEKQAA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 13) H-Aib-HGTFTSDYSIYLEKRAA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 14) H-Aib-HGTFTSDYSILLEKKYA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 15) H-Aib-HGTFTSDYSYLLEKQAA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 16) H-Aib-HGTFTSDYSILLEKQAA-Aib-EFVQWLLEGGPSSGAPPPS (SEQ ID NO: 17) H-Aib-HGTFTSDYSILLEKQAAEEFVQWLLEGGPSSGAPPPS (SEQ ID NO: 18) H-Aib-HGTFTSDYSILLEKQAAQEFVQWLLEGGPSSGAPPPS (SEQ ID NO: 19) H-Aib-HGTFTSDYSILLEKQAAREFVQWLLEGGPSSGAPPPS (SEQ ID NO: 20) H-Aib-HGTFTSDYSILLEKQAAHEFVQWLLEGGPSSGAPPPS (SEQ ID NO: 21) H-Aib-HGTFTSDLSKLKEEQRQKEFIEWLKAGGHPS-Aib-KPPPK (SEQ ID NO: 22) H-Aib-HGTFTSDLSKLKEEQRQKEFIEWLKA-dAla-GHPS-Aib-KPPPK (SEQ ID NO: 23) H-Aib-HGTFTSDLSKLKEEQRQKEFIEWLKAGGPPS-Aib-KPPPK (SEQ ID NO: 24) H-Aib-HGTFTSDLSKLKEEQRQKEFIEWLKA-dAla-GPPS-Aib-KPPPK (SEQ ID NO: 25) H-Aib-HGTFTSDLSKLKEEQRQ-Aib-EFIEWLKAGGPPS-Aib-KPPPK (SEQ ID NO: 28) H-Aib-HGTFTSDLSKLKEEQRQ-Aib-EFIEWLKAGGPPS-Aib-KPPPK (SEQ ID NO: 28) H-Aib-HGTFTSDLSKLKEEQRQ-Aib-EFIEWLKAGGPPS-Aib-KPPPK (SEQ ID NO: 28) H-Aib-HGTFTSDLSKLKEEQRQ-Aib-EFIEWLKAGGPPS-Aib-KPPPK (SEQ ID NO: 28) H-Aib-HGTFTSDLSKLKEEQRQ-Aib-EFIEWLKAGGPPS-Aib-KPPPK (SEQ ID NO: 28) Preferably, the polypeptide or the pharmaceutically acceptable salt thereof has a C-terminal carboxylic acid group or an amide group, more preferably a C-terminal amide group.

8. The pharmaceutical composition according to any one of claims 1-7, wherein, The polypeptide or the pharmaceutically acceptable salt thereof comprises any one of the following structures: Compound 1, Compound 2, Compound 3, Compound 4, Compound 5, Compound 6, Compound 7, Compound 8, Compound 9, Compound 10, Compound 11, Compound 12, Compound 13, Compound 14, Compound 15, Compound 16, Compound 17, Compound 18, Compound 19, Compound 20, Compound 21, Compound 22, Compound 23, Compound 24, Compound 25, Compound 26, Compound 27, Compound 28, Compound 29, Compound 30, Compound 31, Compound 32, Compound 33, Compound 34, Compound 35, Compound 36, Compound 37, Compound 38, Compound 39, Compound 40, Compound 41, Compound 42, Compound 43, Compound 44, Compound 45, Compound 46, Compound 47, Compound 48, Compound 49, Compound 50, Compound 51, Compound 52, Compound 53, 9. The pharmaceutical composition according to any one of claims 1-8, wherein, The divalent metal ion is selected from a divalent zinc ion, a divalent calcium ion, a divalent magnesium ion, a divalent iron ion; Preferably, the metal ion is selected from a divalent zinc ion; More preferably, the metal ion is selected from zinc acetate or zinc chloride; More preferably, the metal ion is zinc acetate.

10. The pharmaceutical composition according to any one of claims 1-9, wherein, The molar ratio of the polypeptide or the pharmaceutically acceptable salt thereof to the divalent metal ion is 1:5 to 5:1; Preferably, the molar ratio of the polypeptide or the pharmaceutically acceptable salt thereof to the divalent metal ion is 1:3 to 3:1; Preferably, the molar ratio of the polypeptide or the pharmaceutically acceptable salt thereof to the divalent metal ion is 1:2 to 2:1; Preferably, the molar ratio of the polypeptide or the pharmaceutically acceptable salt thereof to the divalent metal ion is 1:

1.

11. The pharmaceutical composition according to any one of claims 1-11, wherein, The concentration of the polypeptide or the pharmaceutically acceptable salt thereof is 0.1 mg / mL to 500 mg / mL; Preferably, the concentration of the polypeptide or the pharmaceutically acceptable salt thereof is 0.1 mg / mL to 200 mg / mL; More preferably, 1.0 mg / mL to 100 mg / mL.

12. The pharmaceutical composition according to any one of claims 1-12, wherein, The concentration of the divalent zinc ion is 0.01 mg / mL to 50 mg / mL, Preferably, the concentration of the divalent zinc ion is 0.01 mg / mL to 30 mg / mL, More preferably, the concentration of the divalent zinc ion is 0.01 mg / mL to 10 mg / mL.

13. The pharmaceutical composition according to any one of claims 1-12, wherein, The pharmaceutical composition further comprises a buffer; Preferably, the buffer is selected from one or more of acetate buffer, histidine buffer, phosphate buffer, succinate buffer, Tris, HEPES, MOPS, diethanolamine buffer, borate buffer, Tricine, and citrate buffer; More preferably, the buffer is selected from Tris, disodium hydrogen phosphate buffer, HEPES.

14. The pharmaceutical composition of claim 13, wherein, The concentration of the buffer in the pharmaceutical composition is 0.5 mM to 50.0 mM; Preferably, the concentration of the buffer is 0.5 mM to 35.0 mM; Preferably, the concentration of the buffer is 0.5 mM to 25.0 mM; Preferably, the concentration of the buffer is 2 mM to 10.0 mM.

15. The pharmaceutical composition according to any one of claims 1-14, wherein, The pharmaceutical composition further comprises an osmotic pressure regulator; Preferably, the osmotic pressure regulator is selected from one or more of propylene glycol, mannitol, sorbitol, xylitol, glycerol, lactose, trehalose, sucrose, glucose, sodium chloride, phosphate, sodium citrate, boric acid and sodium tartrate; More preferably, the osmotic pressure regulator is sodium chloride, propylene glycol, mannitol, glycerol; More preferably, the osmotic pressure regulator is propylene glycol or mannitol.

16. The pharmaceutical composition of claim 15, wherein, The concentration of the propylene glycol is 10 mg / mL to 20 mg / mL, preferably 12 mg / mL to 16 mg / mL, more preferably 14 mg / mL; Or the concentration of the mannitol in the pharmaceutical composition is 30 mg / mL to 70 mg / mL, preferably 30 mg / mL to 50 mg / mL, more preferably 46 mg / mL; Or the concentration of the sodium chloride in the pharmaceutical composition is 2 mg / mL to 18 mg / mL, preferably 7 mg / mL to 10 mg / mL, more preferably 9 mg / mL.

17. The pharmaceutical composition according to any one of claims 1-16, wherein, The pharmaceutical composition further comprises a pH regulator; preferably the pH regulator is selected from sodium hydroxide and / or hydrochloric acid.

18. The pharmaceutical composition according to any one of claims 1-17, wherein, The pH of the pharmaceutical composition is 6.5 to 9.0, preferably the pH is 7.0 to 8.0; more preferably 7.1 to 7.7; most preferably 7.5 or 7.

4.

19. The pharmaceutical composition according to any one of claims 1-18, wherein the pharmaceutical composition is selected from: 1) the polypeptide or pharmaceutically acceptable salt thereof according to any one of claims 1-9; 2) divalent zinc ions, such as zinc acetate or zinc chloride; 3) a buffer, such as a phosphate buffer, for example sodium dihydrogen phosphate; 4) an osmotic pressure regulator, such as sodium chloride, propylene glycol, mannitol, glycerol.

20. The pharmaceutical composition of any one of claims 1-19, wherein, The pharmaceutical composition further comprises a bacteriostatic agent; Preferably, the bacteriostatic agent is selected from phenol, o-cresol, m-cresol, p-cresol, methyl paraben, propyl paraben, 2-phenoxyethanol, butyl paraben, 2-phenylethanol, benzyl alcohol, ethanol, chlorobutanol, and thiomersal, bronopol, benzoic acid, imidurea, chlorhexidine, sodium dehydroacetate, chlorocresol, ethyl paraben, benzalkonium chloride or mixtures thereof; More preferably, the bacteriostatic agent is selected from phenol.

21. The pharmaceutical composition of claim 22, wherein, The concentration of the bacteriostatic agent in the pharmaceutical composition is 4.0 mg / mL-7.0 mg / mL; Preferably, 4.4 mg / mL-6.8 mg / mL; More preferably, 5.5 mg / mL to 6.6 mg / mL; Most preferably, 5.5 mg / mL.

22. A method of preparing the pharmaceutical composition according to any one of claims 1-21, comprising the step of dissolving the polypeptide or pharmaceutically acceptable salt thereof.

23. A lyophilized formulation which, upon reconstitution, forms a pharmaceutical composition according to any one of claims 1 to 21, or which is obtained by lyophilizing a pharmaceutical composition according to any one of claims 1 to 21.

24. A reconstituted solution, wherein the reconstituted solution is prepared by reconstituting a lyophilized formulation according to claim 23.

25. An article of manufacture comprising a container housing a pharmaceutical composition according to any one of claims 1 to 21, a lyophilized formulation according to claim 23, or a reconstituted solution according to claim 24.

26. Use of a pharmaceutical composition according to any one of claims 1 to 21, a lyophilized formulation according to claim 23, a reconstituted solution according to claim 24, or an article of manufacture according to claim 25, for the manufacture of a medicament for the treatment of non-insulin dependent diabetes mellitus, insulin dependent diabetes mellitus, obesity, non-alcoholic fatty liver disease, hepatic steatosis, diabetic retinopathy, diabetic neuropathy, diabetic nephropathy, insulin resistance, dyslipidemia associated with insulin resistance, and / or diabetic dyslipidemia.

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