GIP AND GLP-1 DUAL AGONIST POLYPEPTIDE COMPOUND, PHARMACEUTICALLY ACCEPTABLE SALT AND APPLICATION THEREOF

Abstract

A GIP and GLP-1 dual agonist polypeptide compound, having an amino acid sequence as follows:

TABLE-US-00001 (SEQ ID NO: 1) YXaa.sup.1EGTFTSDYSIXaa.sup.2LDKIAQXaa.sup.3AFVQWLIAGGPSSGAPPPS,
and a C-terminal amino acid of polypeptide compound being amidated as a C-terminal primary amide.

Claims

1. A GIP and GLP-1 dual agonist polypeptide compound, having an amino acid sequence as follows: YXaa.sup.1EGTFSDYSIXaa.sup.2LDKIAQXaa.sup.3AFVQWLIAGGPSSGAPPPS (SEQ ID NO: 1), and a C-terminal amino acid of the polypeptide compound being amidated as a C-terminal primary amide; wherein: Xaa.sup.1 is: V, Aib, A, ##STR00020##  Xaa.sup.2 is: V, Aib, A, ##STR00021##  Xaa.sup.1 and Xaa.sup.2 are the same or different;  Xaa.sup.3 is: ##STR00022##  n is an integer number ranging from 5 to 25.

2. The compound of claim 1, wherein n is an integer number 16 or 18.

3. The compound of claim 2, having one of the following amino acid sequences: TABLE-US-00029 (1) ID NO: 1  YVEGTFTSDYSIVLDKIAQK(HOOC-(CH.sub.2).sub.16-CO-γ-Glu-AEEA-AEEA)  AFVQWLIAGGPSSGAPPPS-NH.sub.2;  (2) ID NO: 2  YXEGTFTSDYSIXLDKIAQK(HOOC-(CH.sub.2).sub.16-CO-γ-Glu-AEEA-AEEA)  AFVQWLIAGGPSSGAPPPS-NH.sub.2;  (3) ID NO: 3  YX.sub.1EGTFTSDYSIX.sub.1LDKIAQK(HOOC-(CH+2).sub.16-CO-γ-Glu-AEEA-AEE  A)AFVQWLIAGGPSSGAPPPS-NH.sub.2;  (4) ID NO: 4  YAEGTFTSDYSIALDKIAQK(HOOC-(CH.sub.2).sub.16-CO-γ-Glu-AEEA-AEEA)  AFVQWLIAGGPSSGAPPPS-NH.sub.2;  (5) ID NO: 5  YAEGTFTSDYSIAibLDKIAQK(HOOC-(CH.sub.2).sub.16-CO-γ-Glu-AEEA-AEE  A)AFVQWLIAGGPSSGAPPPS-NH.sub.2;  (6) ID NO: 6  YAibEGTFTSDYSIALDKIAQK(HOOC-(CH.sub.2).sub.16-CO-γ-Glu-AEEA-AEE  A)AFVQWLIAGGPSSGAPPPS-NH.sub.2;  (7) ID NO: 7  YXEGTFTSDYSIX.sub.2LDKIAQK(HOOC-(CH.sub.2).sub.16-CO-γ-Glu-AEEA-AEEA)  AFVQWLIAGGPSSGAPPPS-NH.sub.2;  (8) ID NO: 8  YXEGTFTSDYSIX.sub.2LDKIAQK(HOOC-(CH.sub.2).sub.16-CO-γ-Glu-AEEA-AEEA)  AFVQWLIAGGPSSGAPPPS-NH.sub.2;  (9) ID NO: 9  YX.sub.1EGTFTSDYSIX.sub.2LDKIAQK(HOOC-(CH.sub.2).sub.16-CO-γ-Glu-AEEA-AEE  A)AFVQWLIAGGPSSGAPPPS-NH.sub.2;  (10) ID NO: 10  YX.sub.2EGTFTSDYSIX.sub.1LDKIAQK(HOOC-(CH.sub.2).sub.16CO-γ-Glu-AEEA-AEE  A)AFVQWLIAGGPSSGAPPPS-NH.sub.2;  (11) ID NO: 11  YAibEGTFTSDYSIAibLDKIAQK(γ-Glu-Palmitoyl)AFVQWLIAGGPSS  GAPPPS-NH.sub.2;  (12) ID NO: 12  YVEGTFTSDYSIVLDKIAQK(γ-Glu-Palmitoyl)AFVQWLIAGGPSSGA  PPPS-NH.sub.2;  (13) ID NO: 13  YXEGTFTSDYSIXLDKIAQK(γ-Glu-Palmitolyl)AFVQWLIAGGPSSGA  PPPS-NH.sub.2;  (14) ID NO: 14  YX.sub.1EGTFTSDYSIX.sub.1LDKIAQK(γ-Glu-Palmitoyl)AFVQWLIAGGPSSG  APPPS-NH.sub.2;  (15) ID NO: 15  YAEGTFTSDYSIALDKIAQK(γ-Glu-Palmitoyl)AFVQWLIAGGPSSGA  PPPS-NH.sub.2;  (16) ID NO: 16  YAEGTFTSDYSIAibLDKIAQK(γ-Glu-Palmitoyl)AFVQWLIAGGPSSG  APPPS-NH.sub.2;  (17) ID NO: 17  YAibEGTFTSDYSIALDKIAQK(γ-Glu-Palmitoyl)AFVQWLIAGGPSSG  APPPS-NH.sub.2;  (18) ID NO: 18  YXEGTFTSDYSIX.sub.2LDKIAQK(γ-Glu-Palmitoyl)AFVQVILIAGGPSSG  APPPS-NH.sub.2;  (19) ID NO: 19  YX.sub.2EGTFTSDYSIXLDKIAQK(γ-Glu-Palmitoyl)AFVQWLIAGGPSSG  APPPS-NH.sub.2;  (20) ID NO: 20  YX.sub.1EGTFTSDYSIX.sub.2LDKIAQK(γ-Glu-Palmitoyl)AFVQWLIAGGPSSG  APPPS-NH.sub.2; and  (21) ID NO: 21  YX.sub.2EGTFTSDYSIX.sub.1LDKIAQK(γ-Glu-Palmitoyl)AFVQWLIAGGPSSG  APPPS-NH.sub.2;  ##STR00023## and X.sub.2=Aib.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

[0036] FIG. 1 and

[0037] FIG. 2 are diagrams showing the effect of single administration of each test substance on random blood glucose and AUC.sub.0-24 h Glu Glu in ob/ob mice (X±s, n=7); and

[0038] FIG. 3 is a diagram showing the effect of single administration of each test substance on AUC0-24 h Glu in ob/ob mice (X±s, n=7) (data in the diagram is AUC0-24 h Glu decrease rate).

DETAILED DESCRIPTION

[0039] The disclosure is described with the following embodiments, but these embodiments do not constitute any limitation to the rights of the disclosure.

Example 1

[0040] Synthesis of polypeptide compound of ID NO: 1:

TABLE-US-00004 YVEGTFTSDYSIVLDKIAQK(HOOC-(CH.sub.2).sub.16-CO-γ-Glu-AEEA- AEEA)AFVQWLIAGGPSSGAPPPS-NH.sub.2

[0041] 1. Synthesis of Peptide Chain

[0042] 1.1 Swelling of Resin

[0043] 10 g of Fmoc-Rink Amide AM Resin (with the substitution degree of 0.35 mmol/g) was weighed and swelled for 30 min with 100 mL of DCM, suction filtration was conducted to remove DCM, swelling was conducted for 30 min with 100 mL of DMF, and the resin was washed with 100 mL of DMF and 100 mL of DCM respectively.

[0044] Synthesis of Fmoc-Ser(tBu)-Rink Amide AM Resin

[0045] Fmoc-Ser(tBu)-OH (8 mmol), HOBT (16 mmol) and DIC (16 mmol) were dissolved in 100 mL of DMF, then the solution was added into resin obtained in the previous step to react for 2 hours, after reaction ended, the reaction solution was filtered out, and the resin was washed 3 times with 100 mL of DCM and 100 mL of DMF respectively.

[0046] 1.3 Removal of Fmoc Protecting Group

[0047] A 25% piperidine/DMF (WV) solution containing 0.1 M of HoBt was added into the washed resin to remove Fmoc, and after the reaction ended, the resin was washed 3 times with 100 mL of DCM and 100 mL of DMF respectively.

[0048] 1.4 Extension of Peptide Chain

[0049] According to the sequence, the steps of deprotection and coupling were repeated to sequentially bind corresponding amino acids, and the corresponding amino acids were sequentially bound until peptide chain synthesis was completed, to obtain the peptide resin.

[0050] The 20.sup.th K could adopt a Fmoc-Lys(Alloc)-OH or Fmoc-Lys(Dde)-CH or Fmoc-Lys(iVdde)-OH or Fmoc-Lys(Mtt)-OH or Fmoc-Lys(Boc)-OH protection strategy. In this example, the Fmoc-Lys(Alloc)-OH protection strategy was adopted.

[0051] 1.5 Splicing of Side Chains

[0052] The obtained peptide resin was put into a reaction bottle, and an Alloc protecting group was removed by using Pd(PPH.sub.3).sub.4 under the condition that PhSiH.sub.3 was used as a scavenger; and a 20-site Lys side chain was synthesized by adopting an Fmoc/t-Bu strategy, or Oct(OtBu)-γ-Glu(OtBu)-AEEA-AEEA-Osu fragments were directly coupled to obtain the fully-protected peptide resin.

[0053] 1.6 Cleavage of Peptide Resin

[0054] Trifluoroacetic acid was measured to a reactor and cooled to −10° C. to 0° C., triisopropylsilane, 1,2-dithioglycol and purified water were added and stirred to be uniformly mixed. Peptide resin was slowly added, heated to 20-30° C., and a cracking reaction was conducted for 115-125 min. After the reaction ended, the resin was filtered out, the filtered resin was washed with M×8×20% ml of TFA, the filtered solution and the washing solution were completely transferred into M×8×1.2×4 ml of diethyl ether, stirred for 5-10 min, kept standing to precipitate for 15 min or more. The precipitated turbid liquid was added into a centrifugal machine, and solids were centrifuged and collected; the solids were washed with diethyl ether six times, and the amount of diethyl ether used each time was not less than 5 L. The solids were subjected to vacuum drying for 6-10 h t the temperature of 0-35° C., and crude peptide was obtained.

[0055] 1.7 Purification of Crude Peptide

[0056] Purification was conducted through preparative liquid chromatography, and the chromatographic conditions were that a C18 column (100 mm×250 mm, 10 μm) was adopted; a mobile phase A was 0.1% TFA/water (V/V), and a mobile phase B was 0.1% TFA/acetonitrile (V/V); the mobile phase gradient was 20%-60% of the mobile phase B, and the time was 60 min; the flow velocity was 200 mL/min, the detection wavelength was 214 nm, fractions with the purity larger than 98.0% were collected, and 1.36 g of samples were obtained through freeze drying after rotary evaporation and concentration.

Example 2

[0057] Synthesis of polypeptide compound of ID NO: 2:

TABLE-US-00005 YXEGTFTSDYSIXLDKIAQK(HOOC-(CH.sub.2).sub.16-CO-γ-Glu-AEEA- AEEA)AFVQWLIAGGPSSGAPPPS-NH.sub.2

##STR00008##

[0058] The synthesis procedures were the same as those in Example 1, and the collected solution was freeze-dried to obtain a purified product in a mass of 1.45 g.

Example 3

[0059] Synthesis of polypeptide compound of In NO: 3:

TABLE-US-00006 YXEGTFTSDYSIXLDKIAQK(HOOC-(CH.sub.2).sub.16-CO-γ-Glu-AEEA- AEEA)AFVQWLIAGGPSSGAPPPS-NH.sub.2

##STR00009##

[0060] The synthesis procedures were the same as those in Example 1, and the collected solution was freeze-dried to obtain a purified product in a mass of 1.51 g.

Example 4

[0061] Synthesis of polypeptide compound of ID NO: 4:

TABLE-US-00007 YAEGTFTSDYSIALDKIAQK(HOOC-(CH.sub.2).sub.16-CO-γ-Glu-AEEA- AEEA)AFVQWLIAGGPSSGAPPPS-NH.sub.2

[0062] The synthesis procedures were the same as those in Example 1, and the collected solution was freeze-dried to obtain a purified product in a mass of 1.18 g.

Example 5

[0063] Synthesis of polypeptide compound of ID NO: 5:

TABLE-US-00008 YAEGTFTSDYSIAibLDKIAQK(HOOC-(CH.sub.2).sub.16-CO-γ-Glu-AEEA- AEEA)AFVQWLIAGGPSSGAPPPS-NH.sub.2

[0064] The synthesis procedures were the same as those in Example 1, and the collected solution was freeze-dried to obtain a purified product in a mass of 1.22 g.

Example 6

[0065] Synthesis of polypeptide compound of ID NO: 6:

TABLE-US-00009 YAibEGTFTSDYSIALDKIAQK(HOOC-(CH.sub.2).sub.16-CO-γ-Glu-AEEA- AEEA)AFVQWLIAGGPSSGAPPPS-NH.sub.2

[0066] The synthesis procedures were the same as those in Example 1, and the collected solution was freeze-dried to obtain a purified product in a mass of 1.97 g.

Example 7

[0067] Synthesis of polypeptide compound of ID NO: 7:

TABLE-US-00010 YX1EGTFTSDYSIX2LDKIAQK(HOOC-(CH.sub.2).sub.16-CO-γ-Glu- AEEA-AEEA)AFVQWLIAGGPSSGAPPPS-NH.sub.2

##STR00010##

X.SUB.2.=Aib.

[0068] The synthesis procedures were the same as those in Example 1, and the collected solution was freeze-dried to obtain a purified product in a mass of 1.55 g.

Example 8

[0069] Synthesis of polypeptide compound of ID NO: 8:

TABLE-US-00011 YX.sub.1EGTFTSDYSIX.sub.2LDKIAQK(HOOC-(CH.sub.2).sub.16-CO-γ-Glu- AEEA-AEEA)AFVQWLIAGGPSSGAPPPS-NH.sub.2

X.SUB.1.=Aib;

[0070] ##STR00011##

[0071] The synthesis procedures were the same as those in Example 1, and the collected solution was freeze-dried to obtain a purified product in a mass of 1.01 g.

Example 9

[0072] Synthesis of polypeptide compound of ID NO: 9:

TABLE-US-00012 YX.sub.1EGTFTSDYSIX.sub.2LDKIAQK(HOOC-(CH.sub.2).sub.16-CO-γ-Glu- AEEA-AEEA)AFVQWLIAGGPSSGAPPPS-NH.sub.2

##STR00012##

X.SUB.2.=Aib.

[0073] The synthesis procedures were the same as those in Example 1, and the collected solution was freeze-dried to obtain a purified product in a mass of 1.64 g.

Example 10

[0074] Synthesis of polypeptide compound of ID NO: 10:

TABLE-US-00013 YX.sub.1EGTFTSDYSIX.sub.2LDKIAQK(HOOC-(CH.sub.2).sub.16-CO-γ-Glu- AEEA-AEEA)AFVQWLIAGGPSSGAPPPS-NH.sub.2

X.SUB.1.=Aib;

[0075] ##STR00013##

[0076] The synthesis procedures were the same as those in Example 1, and the collected solution was freeze-dried to obtain a purified product in a mass of 1.81 g.

Example 11

[0077] Synthesis of polypeptide compound of ID NO: 11: YAibEGTFTSDYSIAibLDKIAQK(γ-Glu-Palmitoyl)AFVQWLIAGGPSSGAPPPS-NH.sub.2

[0078] The synthesis procedures were the same as those in Example 1, and the collected solution was freeze-dried to obtain a purified product in a mass of 1.84 g.

Example 12

[0079] Synthesis of polypeptide compound of ID NO: 12:

TABLE-US-00014 YVEGTFTSDYSIVLDKIAQK(γ-Glu-Palmitoyl) AFVQWLIAGGPSSGAPPPS-NH.sub.2

[0080] The synthesis procedures were the same as those in Example 1, and the collected solution was freeze-dried to obtain a purified product in a mass of 1.12 g.

Example 13

[0081] Synthesis of polypeptide compound of ID NO: 13:

TABLE-US-00015 YXEGTFTSDYSIXLDKLAQK(γ-Glu-Palmitoyl) AFVQWLIAGGPSSGAPPPS-NH.sub.2

##STR00014##

[0082] The synthesis procedures were the same as those in Example 1, and the collected solution was freeze-dried to obtain a purified product in a mass of 1.07 g.

Example 14

[0083] Synthesis of polypeptide compound of ID NO: 14:

TABLE-US-00016 YXEGTFTSDYSIXLDKIAQK(γ-GLu-Palmitoyl) AFVQWLIAGGPSSGAPPPS-NH.sub.2

##STR00015##

[0084] The synthesis procedures were the same as those in Example 1, and the collected solution was freeze-dried to obtain a purified product in a mass of 1.89 g.

Example 15

[0085] Synthesis of polypeptide compound of ID NO: 15:

TABLE-US-00017 YAEGTFTSDYSIALDKIAQK(γ-Glu-Palmitoyl) AFVQWLIAGGPSSGAPPPS-NH.sub.2

[0086] The synthesis procedures were the same as those in Example 1, and the collected solution was freeze-dried to obtain a purified product in a mass of 1.81 g.

Example 16

[0087] Synthesis of polypeptide compound of ID NO: 16:

TABLE-US-00018 YAEGTFTSDYSIAibLDKIAQK(γ-Glu-Palmitoyl) AFVQWLIAGGPSSGAPPPS-NH.sub.2

[0088] The synthesis procedures were the same as those in Example 1, and the collected solution was freeze-dried to obtain a purified product in a mass of 1.71 g.

Example 17

[0089] Synthesis of polypeptide compound of ID NO: 17:

TABLE-US-00019 YAibEGTFTSDYSIALDKIAQK(γ-Glu-Palmitoyl) AFVQWLIAGGPSSGAPPPS-NH.sub.2

[0090] The synthesis procedures were the same as those in Example 1, and the collected solution was freeze-dried to obtain a purified product in a mass of 1.53 g.

Example 18

[0091] Synthesis of polypeptide compound of ID NO: 18:

TABLE-US-00020 YX1EGTFTSDYSIX2LDKIAQK(γ-Glu-Palmitoyl) AFVQWLIAGGPSSGAPPPS-NH.sub.2

##STR00016##

X.SUB.2.=Aib.

[0092] The synthesis procedures were the same as those in Example 1, and the collected solution was freeze-dried to obtain a purified product in a mass of 1.25 g.

Example 19

[0093] Synthesis of polypeptide compound of ID NO: 19:

TABLE-US-00021 YX.sub.1EGTFTSDYSIX.sub.2LDKIAQK(γ-Glu-Palmitoyl) AFVQWLIAGGPSSGAPPPS-NH.sub.2

X.SUB.1.=Aib;

[0094] ##STR00017##

[0095] The synthesis procedures were the same as those in Example 1, and the collected solution was freeze-dried to obtain a purified product in a mass of 0.93 g.

Example 20

[0096] Synthesis of polypeptide compound of ID NO: 20:

TABLE-US-00022 YX.sub.1EGTFTSDYSIX.sub.2LDKIAQK(γ-Glu-Palmitoyl) AFVQWLIAGGPSSGAPPPS-NH.sub.2

##STR00018##

X.SUB.2.=Aib.

[0097] The synthesis procedures were the same as those in Example 1, and the collected solution was freeze-dried to obtain a purified product in a mass of 0.96 g.

Example 21

[0098] Synthesis of polypeptide compound of ID NO: 15:

TABLE-US-00023 YX.sub.1EGTFTSDYSIX.sub.2LDKIAQK(γ-Glu-Palmitoyl) AFVQWLIAGGPSSGAPPPS-NH.sub.2

X.SUB.1.=Aib;

[0099] ##STR00019##

[0100] The synthesis procedures were the same as those in Example 1, and the collected solution was freeze-dried to obtain a purified product in a mass of 1.47 g.

Example 22

[0101] The relevant pharmacological experimental methods and results of a GIP and GLP-1 dual agonist polypeptide compound (hereinafter referred to as a polypeptide compound):

[0102] 1. GLP-1 and GIP Rceptor Agonist Activity of Polypeptide Compound

[0103] HEK293 cells were co-transfected with cDNA encoding GLP-1R or GIPR. In a test for determining the compound, the cells were inoculated in a 96-well plate 2 h in advance, the polypeptide compound was dissolved with DMSO, and diluted to different multiples with a culture medium containing 0.1% bovine serum albumin and added to the co-transfected cells. After the cells were incubated for 20 min, fluorescence values were determined through microplate reader of an ELISA kit of the Cisbo Company, a standard curve was established to convert the fluorescence values into corresponding cAMP values, and the EC.sub.50 value of the compound was calculated through nonlinear regression of Graphpad Prism 5.0 software.

TABLE-US-00024 TABLE 1 Agonist activity of polypeptide compound on GLP-1R and GIPR mGLP-1R mGIPR mGLP-1R mGIPR Peptides EC.sub.50 (nM) EC50 (nM) Peptides EC50 (nM) EC50 (nM) Semaglutide 0.0076 68 GIP \ 0.0011 GLP-1 0.00127 \ ID NO: 11 0.02427 0.00632 ID NO: 1 0.01457 0.00068 ID NO: 12 0.05613 0.00879 ID NO: 2 0.00613 0.00086 ID NO: 13 0.05396 0.02250 ID NO: 3 0.00396 0,00149 ID NO: 14 0.03109 0.05165 ID NO: 4 0.00149 0.00521 ID NO: 15 0.04290 0.05320 ID NO: 5 0.00299 0.00640 ID NO: 16 0.04250 0.00965 ID NO: 6 0.00258 0.00929 ID NO: 17 0.04541 0.00972 ID NO: 7 0.00549 0.00766 ID NO: 18 0.00902 0.00833 ID NO: 8 0.00805 0.00858 ID NO: 19 0.01189 0.00791 ID NO: 9 0.00941 0.00698 ID NO: 20 0.01587 0.00591 ID NO: 10 0.00889 0.00811 ID NO: 21 0.01783 0.00807

[0104] As shown in Table 1, the agonist activity of all polypeptide compounds to GLP-1R still kept a high degree of agonist activity, and meanwhile kept a quite high degree of agonist activity to GIPR, and modified non-natural amino acids and side chain groups did not produce great influence on the agonist activity.

[0105] 2. Blood Glucose Reducing Experiment of Polypeptide Compound

[0106] Male ob/ob mice were fed with high-fat feed in a single cage after weaning, blood glucose was predicted after 6-7 weeks, and the mice were divided into a model control group, a polypeptide compound group (selecting 9 compounds), a positive control Liraglutide group and a positive control Semaglutide group according to random weight, random blood glucose and fasting blood glucose. The mice in each test group and the positive control group received a single subcutaneous injection of test substances or positive drugs Liraglutide and Semaglutide at different doses, the mice in the model control group received a single subcutaneous injection of a PBS buffer solution. The mice in each group received random blood glucose value measurement before (0 h) administration and 2, 4, 6, 10 and 24 h after administration, and then the time for measuring the random blood glucose was adjusted and prolonged to 34, 48, 58 or 72 h after administration according to the blood glucose condition of the mice in each group. Meanwhile, the random weight of the mice in each group before (0 h) administration and 24 h after administration was weighed, and then the random weight of the mice corresponding to the random blood glucose measurement condition 48 or 72 h after administration was weighed. The blood glucose decrease rate, the blood glucose curve area (AUC.sub.0-24 h Glu) within 24 h after administration and the decrease rate were calculated according to the following formula.

[0107] Blood glucose decrease rate=(blood glucose in the model control group−blood glucose in the administration group)/blood glucose in the model control group×100%

AUC.sub.0-24 h Glu(mmol/Lh.Math.r)=(BG0−BG2)+(BG2+BG4)+(BG4+BG6)+(BG6+BG10)×2(BG10+BG24)×7

[0108] BG0, BG2, BG4, BG6, BG10 and BG24 represented the blood glucose values before administration (0 h) and 2, 4, 6, 10 and 24 h after administration respectively.

AUC.sub.0-24 h Glu(mmol/Lh.Math.r) decrease rate=(model control group AUC.sub.0-24 h Glu−administration group AUC.sub.0-24 h Glu/model control group AUC.sub.0-24 h Glu×100%

[0109] Experiments were carried out twice, at an interval of 8-10 days. There were 23 groups in each experiment, and the experiment grouping and dosage setting conditions were shown in Table 2.

TABLE-US-00025 TABLE 2 Grouping of experimental animals before administration (X ± s) Dosage Random body Random blood Fasting blood glucose Groups (μg/kg) weight (g) glucose mmol/L) (mmol/L) Model control   42.0 ± 300,8 20.23 ± 301.01 18.94 ± 300.99 GLP 100 42.5 ± 00.8 20.36 ± 300.74 18.97 ± 300.64 GLP-1 100 42.3 ± 300.7 20.30 ± 301.05 19.34 ± 301.07 Liraglutide 100 42.1 ± 01.1 20.14 ± 300.98 18.86 ± 301.28 Semiaglutide 100 42.3 ± 01.0 20.71 ± 301.14 18.83 ± 300.85 ID NO: 1 100 42.2 ± 300,5 20.31 ± 300.99 18.59 ± 300.68 ID NO: 2 100 41.3 ± 300,8 20.62 ± 301.03 18.76 ± 300.75 ID NO: 3 100 42.1 ± 300.5 20.45 ± 300.91 18.72 ± 301.02 ID NO: 4 100 42.6 ± 301.0 20.43 ± 300,68 18.82 ± 300.75 ID NO: 5 100 42.4 ± 300.9 20.57 ± 300.84 18.57 ± 301.14 ID NO: 6 100 42.6 ± 300.6 20.64 ± 300.77 19.15 ± 300.80 ID NO: 7 100 42.7 ± 01.0 20.64 ± 300.99 18.97 ± 300.69 ID NO: 8 100 42.8 ± 300.7 20.36 ± 300.87 18.84 ± 300.78 ID NO: 9 100 42.4 ± 00.8 20.24 ± 300.95 19.02 ± 300.73

[0110] As shown in FIG. 1 to FIG. 3, blood glucose reducing experiment results showed that, when the administration concentration of the polypeptide compound was 100 μg/kg, most of the blood glucose reducing effects were equivalent to those of Semaglutide and GIP, and especially the blood glucose reducing effect of ID NO: 2 was almost consistent with that of Semaglutide.

[0111] 3. Effect of Polypeptide Compound on Random Weight of Ob/Ob Mmice

[0112] As shown in Tables 3 to 5, single subcutaneous injection administration of 100 g/kg of GIP and 100 g/kg of GLP-1 and 300 g/kg of GLP-1 had no obvious influence on the random weight and the variable quantity of the ob/ob mice. After a single subcutaneous injection administration of 100 g/kg of Liraglutide and Semaglutide groups, the random weight and the variable quantity of the ob/ob mice could be significantly reduced in 24 h and 48 h following the administration. After a single subcutaneous injection administration of 300 g/kg of ID NO: 1-9, the random weight and the variable quantity of the ob/ob mice could be significantly reduced in 24 h, 48 h and 72 h following the administration. After a single subcutaneous injection administration of 100 g/kg of ID NO: 2, the random weight variable quantity of the ob/ob mice could be significantly reduced in 24 hours following the administration. After a single subcutaneous injection administration of 300 g/kg of ID NO: 9, the random weight variable quantity of the ob/ob mice could be significantly reduced in 24 h and 48 h following the administration; and the effect of ID NO: 2 was basically equivalent to that of the equal dose of Semaglutide. Therefore, the single subcutaneous injection administration of the GIP and GLP-1 dual agonist compound could significantly reduce the random blood glucose of the type 2 diabetes ob/ob mice, wherein the effect of ID NO: 2 was comparable to that of the equal dose of Semaglutide. After long-term administration, all polypeptide compounds showed a better weight control effect.

TABLE-US-00026 TABLE 3 Effect of single administration of each test substance on random blood glucose and AUC.sub.0-24 h Glu in ob/ob mice (X ± s, n = 7) Dosage Blood glucose at different time after administration (mmol/L) AUC.sub.0-24 h Glu Groups (μg/kg) 0 h 2 h 4 h 6 h 10 h 24 h 34 h 48 h (mmol/L hr) Model — 20.23 ± 1.01 19.72 ± 1.86 21.43 ± 1.92 21.12 ± 1.07 22.06 ± 1.83 20.89 ± 1.08 21.19 ± 1.14 19.71 ± 510.66 ± 24.79 control 2.31 GIP 100 20.36 ± 0.74 12.63 ± 1.70 12.46 ± 1.66 13.71 ± 0.86 14.16 ± 1.24 17.84 ± 0.87 — — 363.99 ± 14.21 GLP-1 100 20.30 ± 1.05 12.06 ± 1.36 11.79 ± 0.84 13.02 ± 0.86 14.03 ± 2.41 18.04 ± 0.71 — — 359.61 ± 18.59 Lirag- 100 20.14 ± 0.98 11.56 ± 1.12 10.34 ± 1.30 10.63 ± 0.84 12.03 ± 0.81 17.71 ± 1.08 — — 328.07 ± 19.12 lutide Semag- 100 20.71 ± 1.14  9.12 ± 0.84  8.96 ± 1.40  8.32 ± 0.88  8.29 ± 0.86  8.49 ± 1.12  9.12 ± 0.83 17.83 ± 215.87 ± 18.67 lutide 1.42 ID NO: 1 100 20.31 ± 0.99 10.12 ± 0.87  9.94 ± 0.86  9.83 ± 0.87 10.76 ± 1.12 11.89 ± 2.12 13.93 ± 1.01 17.32 ± 269.99 ± 19.45 0.89 ID NO: 2 100 20.62 ± 1.03  9.14 ± 0.83  8.95 ± 0.84  8.33 ± 1.42  8.31 ± 1.02  8.51 ± 0.86  8.95 ± 0.87 18.01 ± 216.15 ± 18.86 1.12 ID NO: 3 100 20.45 ± 0.91 10.11 ± 1.01  9.52 ± 1.02  9.63 ± 2.42  9.76 ± 1.13 10.01 ± 1.42 11.56 ± 0.97 17.06 ± 246.51 ± 24.91 1.21 ID NO: 4 100 20.43 ± 0.68 10.21 ± 1.06  9.89 ± 0.83  9.63 ± 0.95  9.86 ± 0.96 11.01 ± 0.83 14.56 ± 1.42 17.32 ± 255.33 ± 20.13 0.83 ID NO: 5 100 20.57 ± 0.84 10.12 ± 1.20  9.54 ± 0.84  9.43 ± 0.86  9.76 ± 0.96 10.89 ± 1.43 13.33 ± 2.10 17.32 ± 252.25 ± 19.68 1.12 ID NO: 6 100 20.64 ± 0.77  9.76 ± 0.83  8.96 ± 0.77  8.55 ± 0.96  9.93 ± 0.76 10.53 ± 1.21 13.13 ± 2.21 17.79 ± 246.81 ± 18.95 0.86 ID NO: 7 100 20.64 ± 0.99 10.11 ± 0.88  9.58 ± 0.97  9.66 ± 0.84  9.76 ± 0.79 10.93 ± 0.86 13.54 ± 1.24 17.34 ± 253.35 ± 18.65 1.20 ID NO: 8 100 20.36 ± 0.87 10.56 ± 0.99  9.99 ± 0.86  9.83 ± 0.85  9.54 ± 1.12 11.95 ± 1.26 14.33 ± 1.43 17.16 ± 260.46 ± 21.31 1.52 ID NO: 9 100 20.24 ± 0.95  9.85 ± 0.89  9.51 ± 0.97  8.92 ± 0.86  8.93 ± 0.86 10.57 ± 0.83 12.26 ± 0.94 17.28 ± 240.08 ± 18.97 1.28

TABLE-US-00027 TABLE 4 Decrease rate of random blood glucose and AUC.sub.0-24 h Glu in ob/ob mice after single administration of each test substance (X ± s, n = 7) AUC.sub.0-24 h Glu Dosage Decrease rate of blood glucose at different time after administration (%) Decrease Groups (μg/kg) 2 h 4 h 6 h 10 h 24 h 34 h 48 h rate (%) GIP 100 38.0 38.8 32.7 30.5 12.4 — — 28.7 GLP-1 100 40.6 41.9 35.9 30.9 11.1 — — 29.6 Liraglutide 100 42.6 48.7 47.2 40.3 12.1 — — 35.8 Semaglutide 100 56.0 56.7 59.8 60.0 59.0 56.0 13.9 57.7 ID NO: 1 100 50.2 51.1 51.6 47.0 41.5 31.4 14.7 47.1 ID NO: 2 100 55.7 56.6 59.6 59.7 58.7 56.6 12.7 57.7 ID NO: 3 100 50.6 53.4 52.9 52.3 51.1 43.5 16.6 51.7 ID NO: 4 100 50.0 51.6 52.9 51.7 46.1 28.7 15.2 50.0 ID NO: 5 100 50.8 53.6 54.2 52.6 47.1 35.2 15.8 50.6 ID NO: 6 100 52.7 56.6 58.6 51.9 49.0 36.4 13.8 51.7 ID NO: 7 100 51.0 53.6 53.2 52.7 47.0 34.4 16.0 50.4 ID NO: 8 100 48.1 50.9 51.7 53.1 41.3 29.6 15.7 49.0 ID NO: 9 100 51.3 53.0 55.9 55.9 47.8 39.4 14.6 53.0

TABLE-US-00028 TABLE 5 Effect of single administration of each test substance on random body weight and variable quantity in ob/ob mice (X ± s, n = 7) Random body weight at different Changes of random body weight at different time time after administration (g) after administration (g) Groups Dosage(μg/kg) 0 h 24 h 48 h 0 h 24 h 48 h Model control — 42.9 ± 0.8 43.5 ± 0.8 43.6 ± 0.7 0.0 ± 0.0  0.6 ± 0.1  0.7 ± 0.1 GIP 100 43.5 ± 0.8 43.6 ± 0.8 — 0.0 ± 0.0  0.1 ± 0.1 — GLP-1 100 43.2 ± 0.7 43.1 ± 0.7 — 0.0 ± 0.0 −0.1 ± 0.1 — Liraglutide 100 43.0 ± 1.1 42.6 ± 1.1 — 0.0 ± 0.0 −0.5 ± 0.1 — Semaglutide 100 43.7 ± 1.0 40.7 ± 1.0 41.7 ± 1.0 0.0 ± 0.0 −2.9 ± 0.2 −1.9 ± 0.2 ID NO: 1 100 43.2 ± 0.5 42.0 ± 0.5 42.7 ± 0.6 0.0 ± 0.0 −1.1 ± 0.2 −0.6 ± 0.1 ID NO: 2 100 43.3 ± 0.8 40.3 ± 0.8 41.3 ± 0.8 0.0 ± 0.0 −2.9 ± 0.2 −1.9 ± 0.2 ID NO: 3 100 43.2 ± 0.5 41.6 ± 0.6 42.1 ± 0.6 0.0 ± 0.0 −1.5 ± 0.2 −0.9 ± 0.2 ID NO: 4 100 43.6 ± 1.0 42.2 ± 1.0 42.9 ± 1.1 0.0 ± 0.0 −1.4 ± 0.2 −0.6 ± 0.1 ID NO: 5 100 43.5 ± 0.9 42.0 ± 0.9 42.9 ± 0.9 0.0 ± 0.0 −1.4 ± 0.2 −0.6 ± 0.2 ID NO: 6 100 43.5 ± 0.6 42.1 ± 0.7 42.7 ± 0.7 0.0 ± 0.0 −1.5 ± 0.2 −0.8 ± 0.1 ID NO: 7 100 43.7 ± 1.0 42.1 ± 1.0 43.0 ± 0.9 0.0 ± 0.0 −1.5 ± 0.2 −0.7 ± 0.1 ID NO: 8 100 43.8 ± 0.7 42.7 ± 0.7 43.2 ± 0.7 0.0 ± 0.0 −1.1 ± 0.2 −0.6 ± 0.1 ID NO: 9 100 43.5 ± 0.8 41.5 ± 0.8 42.4 ± 0.9 0.0 ± 0.0 −1.9 ± 0.2 −1.1 ± 0.1