ACTIVE POLYPEPTIDE COMPOUND

Abstract

The present disclosure relates to the field of drug technology, specifically to an active polypeptide compound, which is Y-ID-X or X-ID-Y; wherein Y is a PTH/PTHrP receptor agonist or an osteoclast inhibitor; ID is a peptide bond or a linker in the molecule, which links X to Y; and X is an osteogenic growth peptide receptor agonist, a bone marrow mesenchymal stem cell irritant or a hematopoietic stem cell irritant. The present disclosure also relates to a pharmaceutical composition comprising the compound, and use of the compound and the pharmaceutical composition in the preparation of a medicament for preventing, treating or alleviating diseases or disorders related to osteogenic defects or bone mineral density decreasing.

Claims

1. An active polypeptide compound, which has a structure represented by following Formula (Ia) or Formula (Ib), or is a pharmaceutically acceptable salt thereof,
Y-ID-X Formula (Ia), or
X-ID-Y Formula (Ib), wherein, Y is a PTH/PTHrP receptor agonist or an osteoclast inhibitor; ID is a peptide bond or a linker in the molecule, which links X to Y; and X is an osteogenic growth peptide receptor agonist, a bone marrow mesenchymal stem cell irritant or a hematopoietic stem cell irritant.

2. The active polypeptide compound according to claim 1, wherein Y is M-CSF antagonist, RANKL inhibitor, RANKL antibody, MMP inhibitor, calcitonin, parathyroid hormone or parathyroid hormone-related protein.

3. The active polypeptide compound according to claim 1, wherein Y is a peptide chain having an amino acid sequence as shown in Formula (II):
A.sub.1-Val-Ser-Glu-His-Gln-Leu-A.sub.8-His-Asp-Lys-Gly-Lys-Ser-Ile-Gln-A.sub.17-Leu-Arg-Arg-Arg-A.sub.22-A.sub.23-Leu-A.sub.25-A.sub.26-Leu-A.sub.28-A.sub.29-A.sub.30-A.sub.31-His-Thr-Ala Formula (II); wherein, A.sub.1 is Ala, Val, Leu or Ile; A.sub.8 is Leu or Ile; A.sub.17 is Asp or Glu; A.sub.22 is Glu, Asp or Phe; A.sub.23 is Leu, Ile or Phe; A.sub.25 is Glu, Asp or His; A.sub.26 is Lys, His or Arg; A.sub.28 is Leu, Ile or Val; A.sub.29 is Ala, (N-Me)Ala or Aib; A.sub.30 is Lys or Glu; A.sub.31 is Leu or Ile; the amino terminal of the peptide chain Y is free or chemically modified, and the carboxyl terminal of the peptide chain Y is free or chemically modified.

4. The active polypeptide compound according to claim 1, wherein X is a hematopoietic stem cell irritant, a hematopoietic growth factor, a platelet colony-stimulating factor, a granulocyte colony-stimulating factor, erythropoietin, interleukin 3 or recombinant human interleukin 11.

5. The active polypeptide compound according to claim 1, wherein X is a peptide chain having an amino acid sequence as shown in Formula (IIIa) or Formula (IIIb):
Tyr-(Arg).sub.m-(Gly).sub.n-Phe-Gly-Gly Formula (IIIa)
Gly-Gly-Phe-(Gly).sub.n-(Arg).sub.m-Tyr Formula (IIIb); wherein, m and n are independently 0, 1 or 2; and the amino terminal of the peptide chain X is free or chemically modified, and the carboxyl terminal of the peptide chain X is free or chemically modified.

6. The active polypeptide compound according to claim 5, wherein X is a peptide chain consisting of 5-6 amino acids which has an amino acid sequence as shown in one of the following SEQ ID NO:1-SEQ ID NO:8: TABLE-US-00021 (SEQ ID NO: 1) Tyr-Gly-Phe-Gly-Gly (SEQ ID NO: 2) Tyr-Arg-Phe-Gly-Gly (SEQ ID NO: 3) Tyr-Arg-Gly-Phe-Gly-Gly (SEQ ID NO: 4) Tyr-Pro-Phe-Gly-Gly (SEQ ID NO: 5) Gly-Gly-Phe-Gly-Tyr (SEQ ID NO: 6) Gly-Gly-Phe-Arg-Tyr (SEQ ID NO: 7) Gly-Gly-Phe-Gly-Arg-Tyr (SEQ ID NO: 8) Gly-Gly-Phe-Pro-Tyr.

7. The active polypeptide compound according to claim 1, wherein ID is a linker between X and Y; the linker is an amino-substituted C.sub.1-8 alkyl carboxylic acid, a polyethylene glycol polymer chain or a peptide segment consisting of 1-10 amino acids, and the amino acids in the peptide segment is selected from the group consisting of proline, arginine, alanine, threonine, glutamic acid, aspartic acid, lysine, glutamine, asparagine and glycine.

8. The active polypeptide compound according to claim 7, wherein the linker is one of the following linkers: (1) (Gly-Ser).sub.p, wherein p is 1, 2, 3, 4 or 5; (2) (Gly-Gly-Gly-Gly-Ser).sub.t, wherein t is 1, 2 or 3; (3) Ala-Glu-Ala-Ala-Ala-Lys-Ala; (4) 4-aminobutyric acid or 6-aminocaproic acid; and (5) (PEG).sub.q, wherein q is 1, 2, 3, 4 or 5.

9. The active polypeptide compound according to claim 1, which has a structure as shown in Formula (IV), or is a pharmaceutically acceptable salt of the compound shown as in Formula (IV):
A.sub.1-Val-Ser-Glu-His-Gln-Leu-A.sub.8-His-Asp-Lys-Gly-Lys-Ser-Ile-Gln-A.sub.17-Leu-Arg-Arg-Arg-A.sub.22-A.sub.23-Leu-A.sub.25-A.sub.26-Leu-A.sub.28-A.sub.29-A.sub.30-A.sub.31-His-Thr-Ala-A.sub.35 Formula (IV), wherein, A.sub.1 is Ala, Val, Leu or Ile; A.sub.8 is Leu or Ile; A.sub.17 is Asp or Glu; A.sub.22 is Glu, Asp or Phe; A.sub.23 is Leu, Ile or Phe; A.sub.25 is Glu, Asp or His; A.sub.26 is Lys, His or Arg; A.sub.28 is Leu, Ile or Val; A.sub.29 is Ala, (N-Me)Ala or Aib; A.sub.30 is Lys or Glu; A.sub.31 is Leu or Ile; and A.sub.35 has a peptide chain of the amino acid sequence as shown in Formula (IIIa) or (IIIb):
Tyr-(Arg).sub.m-(Gly).sub.n-Phe-Gly-Gly Formula (IIIa),
Gly-Gly-Phe-(Gly).sub.n-(Arg).sub.m-Tyr Formula (IIIb), wherein, m and n are independently 0, 1 or 2; and the amino terminal of the amino acids shown by A.sub.1 is free or chemically modified, and the carboxyl terminal of the peptide chain A.sub.35 is free or chemically modified.

10. The active polypeptide compound according to claim 9, wherein the chemical modifications of the amino terminal include acylation, sulfonylation, alkylation and PEG modification; and the chemical modifications of the carboxyl terminal include amidation, sulfonylation and PEG modification.

11. The active polypeptide compound according to claim 10, wherein the chemical modification of amino terminal is acetylation, benzoylation or sulfonylation of amino; the alkylation of amino terminal is C.sub.1-6 alkylation or aromatic alkylation; the chemical modification of carboxylic terminal is that the OH in the carboxyl is substituted by NH.sub.2 or sulfamide, or the OH in the carboxyl links to the functionalized PEG molecule.

12. The active polypeptide compound according to claim 1, which is one of the compounds in the following SEQ ID NO:9-SEQ ID NO:15, or a pharmaceutically acceptable salt thereof: TABLE-US-00022 (1) SEQ ID NO: 9 Ala-Val-Ser-Glu-His-Gln-Leu-Leu-His-Asp-Lys-Gly- Lys-Ser-Ile-Gln-Asp-Leu-Arg-Arg-Arg-Glu-Leu-Leu- Glu-Lys-Leu-Leu-(N-Me)Ala-Lys-Leu-His-Thr-Ala- Tyr-Gly-Phe-Gly-Gly; (2) SEQ ID NO: 10 Ala-Val-Ser-Glu-His-Gln-Leu-Leu-His-Asp-Lys-Gly- Lys-Ser-Ile-Gln-Asp-Leu-Arg-Arg-Arg-Glu-Leu-Leu- Glu-Lys-Leu-Leu-Aib-Lys-Leu-His-Thr-Ala-Tyr-Gly- Phe-Gly-Gly; (3) SEQ ID NO: 11 Ala-Val-Ser-Glu-His-Gln-Leu-Leu-His-Asp-Lys-Gly- Lys-Ser-Ile-Gln-Asp-Leu-Arg-Arg-Arg-Glu-Leu-Leu- Glu-Lys-Leu-Leu-Ala-Lys-Leu-His-Thr-Ala-Tyr-Arg- Gly-Phe-Gly-Gly; (4) SEQ ID NO: 12 Ala-Val-Ser-Glu-His-Gln-Leu-Leu-His-Asp-Lys-Gly- Lys-Ser-Ile-Gln-Asp-Leu-Arg-Arg-Arg-Phe-Phe-Leu- His-His-Leu-Ile-Ala-Glu-Ile-His-Thr-Ala-Tyr-Gly- Phe-Gly-Gly; (5) SEQ ID NO: 13 Ala-Val-Ser-Glu-His-Gln-Leu-Leu-His-Asp-Lys-Gly- Lys-Ser-Ile-Gln-Asp-Leu-Arg-Arg-Arg-Phe-Phe-Leu- His-His-Leu-Ile-Aib-Glu-Ile-His-Thr-Ala-Tyr-Arg- Phe-Gly-Gly; (6) SEQ ID NO: 14 Ala-Val-Ser-Glu-His-Gln-Leu-Ile-His-Asp-Lys-Gly- Lys-Ser-Ile-Gln-Glu-Leu-Arg-Arg-Arg-Phe-Phe-Leu- His-His-Leu-Ile-Aib-Glu-Ile-His-Thr-Ala-Tyr-Gly- Phe-Gly-Gly; (7) SEQ ID NO: 15 Ala-Val-Ser-Glu-His-Gln-Leu-Ile-His-Asp-Lys-Gly- Lys-Ser-Ile-Gln-Glu-Leu-Arg-Arg-Arg-Phe-Phe-Leu- His-His-Leu-Leu-Ala-Glu-Ile-His-Thr-Ala-Tyr-Gly- Phe-Gly-Gly.

13. The active polypeptide compound according to claim 1, further includes a compound obtained by chemically modifying the side chain groups of amino acids of the polypeptide compound; or a coordination compound, a complex or a chelate formed by the polypeptide compound and a metal ion; or a hydrate or a solvate formed by the polypeptide compound.

14. The active polypeptide compound according to claim 13, wherein the compound obtained by chemically modifying the side chain groups of amino acids of the polypeptide compound is a thioether or thioglycoside formed from a sulfydryl in a cysteine in the polypeptide compound, or a compound having a disulfide bond formed from a cysteine or a peptide comprising cysteine; or an ester, an ether and a glycoside formed from a phenolic hydroxyl group of a tyrosine in the polypeptide compound; or a compound prepared by substituting a benzene ring of a tyrosine or phenylalanine in the polypeptide compounds.

15. A pharmaceutical composition, comprising the active polypeptide compound according to claim 1, and at least one of a pharmaceutically acceptable adjuvant, excipient, carrier and solvent thereof.

16. The pharmaceutical composition according to claim 15, comprising other therapeutic agents, which are selected from a drug that inhibits bone resorption, a drug that promotes ossification, a drug that promotes bone mineralization and a parathyroid hormone-related protein.

17. The pharmaceutical composition according to claim 16, wherein the drug that inhibits bone resorption includes calcitonin, diphosphonate, oestrogen, a selective oestrogen receptor regulator and isoflavone; the drug that promotes ossification includes fluoride, synthesized steroid, parathyroid hormone and parathyroid hormone-related protein; the drug that promotes bone mineralization includes a calcium agent, vitamin D and active vitamin D; and the parathyroid hormone-related protein is teriparatide or abaloptide.

18. A method of preventing, treating or alleviating diseases or disorders related to osteogenic defects or bone mineral density decreasing, comprising administering a subject in need thereof the compound according to claim 1.

19. The method according to claim 18, wherein the disease is osteoporosis.

Description

BRIEF DESCRIPTION OF DRAWINGS

[0115] In order to explain the technical solutions in the embodiments of the present disclosure or the prior art more clearly, the drawings used in the description of the examples or the prior art will be briefly introduced hereinafter.

[0116] FIG. 1 shows a scanned figure by micron X-ray 3D imaging system of trochlea of thigh bone in the Control group, Model group, abaloptide group (Aba group), 20 μg/kg dosage group of Example 1, 20 μg/kg dosage group of Example 2 and 20 μg/kg dosage group of Example 5.

DETAILED DESCRIPTION

[0117] The present disclosure will be described in further detail below with reference to specific examples, but the embodiments of the present disclosure are not limited thereto. The embodiments of the present disclosure are given merely for the purpose of illustrating the present disclosure, rather than limiting the present disclosure. Therefore, any improvement to the present disclosure under the premise of the method of the present disclosure belongs to the protection scope of the present disclosure. Generally, the compounds of the present disclosure can be prepared by the methods described in the present disclosure. One of ordinary skill in the art can also use well-known methods to select sequential or different synthetic steps to produce polypeptide compounds having the structure described in the present disclosure. The following reaction schemes and examples are provided to further illustrate the content of the present disclosure.

[0118] One of ordinary skill in the art will recognize that the polypeptide compounds described in the present disclosure can be prepared by solid-phase synthesis (SPPS), liquid-phase synthesis, and enzymatic synthesis. The polypeptide compounds of the present disclosure prepared by different preparation methods all fall within the scope of the present disclosure. For example, peptide compounds are usually prepared by solid-phase synthesis. The solid-phase synthesis may be selected from conventional polystyrene-divinylbenzene crosslinked resins, polyacrylamides, polyethylene-glycol resins, and the like, for example: Wang Resin, Fmoc-Pro-CTC, Rink Amide Linker MBHA resin, etc. According to different linking sequences, appropriate resins are selected. For example, the carboxyl group of the carboxyl-terminal amino acid can be first covalently bonded to the polymer solid phase carrier. The protective group of the α-amino group can also be Fmoc, Boc, or Z. From C-terminal to N-terminal, the amino acids are subjected to repetitive process of de-protection, condensation, re-de-protection and condensation according to a certain sequences, giving a peptide chain resin having protective groups, which is subjected to steps of resin removal and de-protection, giving the required peptide chain. The amino of the amino acid on the amino-terminal can also be covalently bonded to the polymer solid phase carrier. By reverse synthesis, from the N-terminal to the C-terminal, the amino acids are subjected to repetitive process of de-protection, condensation, re-de-protection and condensation according to a certain sequences, giving a peptide chain resin having protective groups, which is subjected to steps of resin removal and de-protection, giving the required peptide chain. Terminals of peptide chains obtained by using different kinds of resin sometimes may differ. For example, peptide segment prepared by Wang resin have free carboxyl terminals. Similarly, peptide segment with NH.sub.2 modified-carboxyl terminals are obtained when Rink-AM amino resin is used as a solid phase.

[0119] The amino acid raw materials required for peptide compound synthesis were purchased from GL Biochemical (Shanghai) Co., Ltd.; the solid-phase synthetic resin was purchased from Xi'an Sunresin Technology New Material Co., Ltd.; the amino acid condensation catalysts TBTU and DIEA used were purchased from Suzhou Highfine Biotechnology Co., Ltd. The eluents used in the preparation of HPLC was of chromatographic grade. The reagents were purchased from commercial suppliers such as Aldrich Chemical Company, Arco Chemical Company and Alfa Chemical Company, and were used without further purification, unless otherwise indicated. The analysis and detection instruments used are conventional instruments and equipment in the field. In the examples described below, unless otherwise indicated that all temperatures are set to degrees Celsius, and the given temperature may have a fluctuation range of ±5° C.

[0120] When identifying the structure of the peptide compound, QE identification, N-terminal analysis of protein (by mass spectrometry) and N-terminal sequence analysis of polypeptide protein were used to confirm the primary structure, and circular dichroism scanning analysis was used to confirm the secondary structure.

[0121] In the examples of the present disclosure, the circular dichroism scanning analysis of polypeptide compound is performed with a Chirascan Plus V100 circular dichroism spectrometer (British Applied Optics) to collect the circular dichroism (CD) absorption spectra of the protein test product in far ultraviolet (190-260 nm) and near ultraviolet (250-340 nm) regions, and to analyze the secondary structure by software. In the specific measurement, a scanning wavelength of 180-340 nm was set for background test and blank buffer test, and then circular dichroism far and near ultraviolet absorptions of a 1 mg/mL CSA standard solution was collected in the range of 180-340 nm. All scanned spectra were subjected to subtract baseline and smoothing treatments with software Pro-Data Viewer. The ratio of the peak and valley CD values of a standard sample was calculated, and the effective ratio range is 2.08±0.06. CDNN software was used to fit the secondary structure of the test sample, and the proportions of helix, antiparallel+parallel, beta-turn, and random coil in different wavelength intervals were calculated in the Milli-Degress mode.

[0122] In the QE identification of the polypeptide compound in examples of the present disclosure, protein polypeptide was subjected to enzymolysis with an endonuclease (generally Trypsin), and then LC/MS/MS (nanoLC-QE) was used to analyze the sample after enzymolysis. Finally, the mass spectrometry software such as MASCOT was used to analyze the LC/MS/MS data to obtain the qualitative identification information of the target protein and peptide molecules. In the specific measurement, after the test product was reduced and alkylated, Trypsin (in a mass ratio of 1:50) was added, and enzymolysis was carried out at 37° C. for 20 hours. The enzymolysis product was desalted, freeze-dried, re-dissolved in a 0.1% FA solution, and stored at −20° C. until use. Q Exactive (Thermo Fisher) and Easy-nLC 1000 (Thermo Fisher) were used. The mass-to-charge ratios of polypeptides and polypeptide fragments were collected as follows: 20 fragment spectra (MS2 scan) were collected after each full scan. The raw file of mass spectrometry test was used to search the corresponding database with Mascot2.2 software, finally giving the identified protein results.

[0123] In the examples of the present disclosure, the experimental method of protein N-terminal sequence analysis (by mass spectrometry) of polypeptide compound was performed by: subjecting the protein to enzymolysis respectively with trypsin, chymotrypsin and Glu-C enzyme, and then using LC-MS/MS (Xevo G2-XS QTof, Waters) to analysis the peptide segment sample after enzymolysis. Enzymolysis method: 50 μg of test product was dissolved in in an appropriate amount of guanidine hydrochloride to denature, then after DTT and IAM reactions, the disulfide bond was reduced and protected by alkylation modification, and 1 μg of trypsin, 1 μg of chymotrypsin and 1 μg of Glu-C enzyme were added after dilution, reacted at 37° C. for 20 hours. Finally, the UNIFI software was used to analyze the LC-MS/MS data, and the N-terminal amino acid sequence of the test product was determined to whether be in accordance with the theoretical sequence based on the results of the algorithm. For the specific measurement, the instruments were (1) high-resolution mass spectrometer: XevoG2-XS QTof (Waters), and (2) ultra-high performance liquid chromatography: UPLC (Acquity UPLC I-Class) (Waters).

[0124] The N-terminal sequence analysis of the polypeptide protein of the polypeptide compound in the examples of the present disclosure was performed by analyzing the N-terminal sequence of the test product by a fully automatic protein peptide sequencer. The PPSQ fully automatic protein peptide sequencer (SHIMADZU) was used in the examples of the present disclosure. Sample name, sample number, number of test cycles and selection of a method file were set by software PPSQ Analysis, and the test started after the settings were completed. Data and Atlas Processing: the raw data and spectra generated by PPSQ were identified, peaks were marked up by PPSQ Data Processing software, and the corresponding spectra were derived.

[0125] In the examples of the present disclosure, the preliminary structure of the polypeptide compound was determined by mass spectrometry. High-resolution mass spectrometry was performed with ABSciex 5800 MALDI-TOF/TOF to test the relative molecular mass of the protein, and accurate and reliable relative molecular mass information of the polypeptide was obtained.

[0126] The following abbreviations are used throughout the disclosure:

[0127] Boc: tert-butoxycarbonyl

[0128] DIEA: diisopropylethylamine

[0129] DCM: dichloromethane

[0130] CH.sub.3CN: acetonitrile

[0131] DCM: dichloromethane

[0132] DMF: N,N-dimethylformamide

[0133] DEPBT: 3-(diethoxy orthoacyloxy)-1,2,3-benzotriazin-4-one

[0134] DIEA: diisopropylethylamine

[0135] Et.sub.2O: ethyl ether

[0136] EDT: Ethylene Dithiol

[0137] Fmoc: 9H-fluoren-9-ylmethoxycarbonyl

[0138] H.sub.2O: water

[0139] HBTU: 2-(1H-benzotriazol-1-yl-)-1,1,3,3-tetramethylurea hexafluorophosphate

[0140] NMP: 1-methyl-pyrrolidin-2-one

[0141] Ot-Bu: tert-butoxy

[0142] PyBOP: 1H-benzotriazol-1-yloxytripyrrolidinyl hexafluorophosphate

[0143] Pbf: 2,2,4,6,7-pentamethylbenzodihydrofuran-5-sulfonyl

[0144] t-Bu: tert-butyl

[0145] Trt: Trityl

[0146] TIS: Triisopropyl silane

[0147] T.sub.3P: 1-propyl phosphoric anhydride

[0148] TFA: trifluoroacetic acid

[0149] Trt: trityl

[0150] rt: room temperature

[0151] TA: thioanisole

Examples of Preparation

[0152] In the following specific examples, the peptides in the present disclosure can be prepared by standard solid-phase synthesis method. The preparing process will be described in detail hereinafter. Other peptides in the present disclosure can be prepared by a similar method by one of ordinary skill in the art.

TABLE-US-00004 Example 1: (the polypeptide sequence shown as SEQ ID No.9) Preparation of Ala-Val-Ser-Glu-His-Gln-Leu-Leu- His-Asp-Lys-Gly-Lys-Ser-Ile-Gln-Asp-Leu-Arg-Arg- Arg-Glu-Leu-Leu-Glu-Lys-Leu-Leu-(N-Me)Ala-Lys- Leu-His-Thr-Ala-Tyr-Gly-Phe-Gly-Gly

[0153] This example was synthesized from C terminal to N terminal.

[0154] (1-1) Preprocessing of resin: 30 g 2-CTA resin (degree of substitution 0.93 mmol/g, 27 mmol) was weighed, and placed in a 250 mL test tube. The resin was swelled with 180 mL DCM for 30 min. The solvent was pumped off in vacuum. Then DCM (100 mL) was added in, nitrogen was introduced in and heated, until the temperature reached 25° C. SOCl.sub.2 (10 mL, 5.0 eq) was added dropwise, the temperature was kept at 30-35° C., and reacted for 2 h. After the reaction was completed, nitrogen was introduced in to press and solvent was pumped off in vacuum. DCM (100 mL×3) was added to wash the resin, and the solvent was pumped off each time after washing.

[0155] (1-2) Linking S1 amino acid: Fmoc-Gly-OH (32.5 g, 108 mmol) was weighed, and dissolved in 100 mL DCM. After dissolved completely, DIEA (23 mL, 135 mmol) was added. The obtained mixture was placed in a test tube, nitrogen was introduced, the mixture was stirred and reacted at 20° C.-30° C. for 2 h. 12 mL of mixed solvent of methanol and DIEA (methanol:DIEA=9:1) was added droppedwise, to seal the unreacted sites for 10 min. The solvent was pumped off. The resin was washed with DCM (150 mL×2). After washing, the solvent was pumped off. Then DMF (150 mL×2) was used to wash the resin, and the solvent was pumped off after washing. The resin was then swelled with 120 mL DMF for 30 min. The solvent was pumped off, and de-protection was performed with a piperidine/DMF solution (120 mL) having a volume ratio of 20% for twice. The times were respectively 10 min and 15 min. The temperature of the reaction was controlled at 20° C.-30° C. After the de-protection, the solvent was pumped off, and the resin was washed with DMF (120 mL×6). After washing, the solvent was pumped off, and the resin was left in the test tube. Ninhydrin was used to test the color of resin, and the resin was purple-black. The next step was carried out. The absorbance was detected by spectrophotometric method, and the degree of substitution of the resin was calculated to be 0.8641 mmol/g.

[0156] (1-3) Linking S2 Amino Acid:

[0157] Fmoc-Gly-OH (19.0 g, 63.9 mmol, 3.0 eq.) and PyBOP (33.25 g, 63.9 mmol, 3.0 eq.) were weighted, and dissolved with 50 mL DMF. After dissolved completely, DIEA (10.5 mL, 63.9 mmol) was added. The obtained mixture was placed in a test tube, nitrogen was introduced, the mixture was stirred and reacted at 20° C.-30° C. for 2 h. Ninhydrin was used to test the color of resin, the resin was transparent yellow. After the reaction, the solvent was pumped off, and the resin was washed with DMF (120 mL×3). After washing, the solvent was pumped off, and then de-protection was performed with a piperidine/DMF solution (120 mL) having a volume ratio of 20% for twice. The times were respectively 10 min and 15 min. The temperature of the reaction was controlled at 20° C.-30° C. After the de-protection, the solvent was pumped off, and the resin was washed with DMF (120 mL×6). After washing, the solvent was pumped off, and the resin was left in the test tube. Ninhydrin was used to test the color of resin, and the resin was purple-black. The next step was carried out.

[0158] (1-4) Linking S3 Amino Acid:

[0159] Fmoc-Phe-OH (24.75 g, 63.9 mmol, 3.0 eq.) and PyBOP (33.25 g, 63.9 mmol, 3.0 eq.) were weighted, and dissolved in 50 mL DMF. After dissolved completely, DIEA (10.5 mL, 63.9 mmol) was added. The obtained mixture was placed in a test tube, nitrogen was introduced, the mixture was stirred and the temperature was controlled at 20° C.-30° C. and reacted for 2 h. Ninhydrin was used to test the color of resin, the resin was transparent yellow. After the reaction, the solvent was pumped off, and the resin was washed with DMF (120 mL×3). After washing, the solvent was pumped off, and then de-protection was performed with a piperidine/DMF solution (120 mL) having a volume ratio of 20% for twice. The times were respectively 10 min and 15 min, and the temperature of the reaction was controlled at 20° C.-30° C. After the de-protection, the solvent was pumped off, and the resin was washed with DMF (120 mL×6). After washing, the solvent was pumped off, and the resin was left in the test tube. Ninhydrin was used to test the color of resin, and the resin was purple-black. The next step was carried out.

[0160] (1-5) Steps (1-4) were repeated. S4 amino acid Fmoc-Gly-OH, S5 amino acid Fmoc-Tyr(t-Bu)-OH, S6 amino acid Fmoc-Ala-OH, S7 amino acid Fmoc-Thr(t-Bu)-OH, S8 amino acid Fmoc-His(trt)-OH, S9 amino acid Fmoc-Leu-OH, S10 amino acid Fmoc-Lys(Boc)-OH, S11 amino acid Fmoc-(N-Me)Ala-OH, S12 amino acid Fmoc-Leu-OH, S13 amino acid Fmoc-Leu-OH, S14 amino acid Fmoc-Lys(Boc)-OH, S15 amino acid Fmoc-Glu(Ot-Bu)-OH, S16 amino acid Fmoc-Leu-OH, S17 amino acid Fmoc-Leu-OH, S18 amino acid Fmoc-Glu(Ot-Bu)-OH, S19 amino acid Fmoc-Arg(pbf)-OH, S20 amino acid Fmoc-Arg(pbf)-OH, S21 amino acid Fmoc-Arg(pbf)-OH, S22 amino acid Fmoc-Leu-OH, S23 amino acid Fmoc-Asp(OtBu)-OH, S24 amino acid Fmoc-Gln(trt)-OH, S25 amino acid Fmoc-Ile-OH, S26 amino acid Fmoc-Ser(tBu)-OH, S27 amino acid Fmoc-Lys(Boc)-OH, S28 amino acid Fmoc-Gly-OH, S29 amino acid Fmoc-Lys(Boc)-OH, S30 amino acid Fmoc-Asp(Ot-Bu)-OH, S31 amino acid Fmoc-His(trt)-OH, S32 amino acid Fmoc-Leu-OH, S33 amino acid Fmoc-Leu-OH, S34 amino acid Fmoc-Gln(trt)-OH, S35 amino acid Fmoc-His(trt)-OH, S36 amino acid Fmoc-Glu(Ot-Bu)-OH, S37 amino acid Fmoc-Ser(t-Bu)-OH, S38 amino acid Fmoc-Val-OH and S39 amino acid Fmoc-Ala-OH were successively linked, and a peptide resin was obtained to carry out the next operation.

[0161] (1-6) Resin shrinkage: methanol (80 mL) was firstly added in the test tube, the resin was shrunken for 5 min, and the solvent was pumped off. The shrinkage was repeated for 3 times, 10 min once. Each time after the shrinkage, the solvent was pumped out completely before the next shrinkage. Then the shrunken resin was placed in a vacuum drying oven, dried at 35° C., and 18.82 g peptide resin was obtained.

[0162] (1-7) Peptide segment cracking: 155 mL TFA, 8 mL TIS, 4.12 mL EDT, 2 mL TA, 4.12 mL water and 2 mL anisole were mixed evenly to prepare the lysate. 18.82 g of the peptide resin prepared in step (1-6) was weighed, the lysate and the peptide resin were mixed, sealed and shielded from the light. The mixture was stirred and reacted, the temperature was kept at 25° C.-35° C., and reacted for 2 h. After the reaction, a sand core funnel was used to remove the resin. After removing the solvent in vacuum, methyl tertiary butyl ether (450 mL) was added in the rest liquid, and crystallized at 0° C.-10° C. for 2 h. The mixture was centrifuged to remove the crystallizing solution, and the precipitate was washed with methyl tertiary butyl ether for 3 times. The precipitate was collected, and dried in vacuum at 35° C., to give a polypeptide (SEQ ID NO:9) Ala-Val-Ser-Glu-His-Gln-Leu-Leu-His-Asp-Lys-Gly-Lys-Ser-Ile-Gln-Asp-Leu-Arg-Arg-Arg-Glu-Leu-Leu-Glu-Lys-Leu-Leu-(N-Me)Ala-Lys-Leu-His-Thr-Ala-Tyr-Gly-Phe-Gly-Gly.

[0163] (1-8) Purification: after filtering the peptide crude solution obtained in step (1-7) with a 0.45 μm filter membrane, the solution was subjected to preparative HPLC purification in a 20 mm×150 mm column filled with 10 μm C-18 silica gel. The detection wavelength was 220 nm. The mobile phase A was 0.1% TFA, and the mobile phase B was acetonitrile. Gradient elution was carried out according to the following Table A.

TABLE-US-00005 TABLE A Gradient elution program Flow rate Mobile Mobile Time (mL/ phase phase (min) min) A (%) B (%) 0 8 95 5 0.1 8 75 25 45 8 65 50 60 8 50 50

[0164] Fractions containing target polypeptide product was collected, and the purity was 95.8%. The collected fractions were combined, the solvent was removed in vacuum, and the polypeptide compound was freeze-dried. The obtained end product was identified by analytical RP-HPLC (retention time), LC-MS and MALDI/TOF-MS.

[0165] MALDI/TOF-MS(ESI): 4441.2236 [M+H].sup.+.

[0166] According to QE identification and analysis, the sequence of the obtained polypeptide compound was as that shown as SEQ ID NO:9.

[0167] Assay: moisture content was measured with a moisture titrator by the moisture determination method in Chinese Pharmacopoeia, and TFA content was measured by the acetic acid content detection method in Chinese Pharmacopoeia, and the content of the polypeptide was 83.7%.

TABLE-US-00006 Example 2: (the polypeptide sequence shown as SEQ ID No. 10) Preparation of Ala-Val-Ser-Glu-His-Gln-Leu-Leu- His-Asp-Lys-Gly-Lys-Ser-Ile-Gln-Asp-Leu-Arg-Arg- Arg-Glu-Leu-Leu-Glu-Lys-Leu-Leu-Aib-Lys-Leu-His- Thr-Ala-Tyr-Gly-Phe-Gly-Gly

[0168] The compound was synthesized from C terminal to N terminal:

[0169] (2-1) Preprocessing of resin: 30 g 2-CTA resin (degree of substitution 0.93 mmol/g, 27 mmol) was weighed, disposed in a 250 mL test tube, and swelled in DCM (200 mL) for 30 min. The solvent was pumped out in vacuum. Then DCM (100 mL) was added, nitrogen was introduced, and heated until the temperature reached 25° C. SOCl.sub.2 (10 mL, 5.0 eq) was added dropwise, the temperature was kept at 30-35° C., and the reaction was performed for 2 h. After the reaction was complete, nitrogen was introduced in and solvent was pumped off. DCM (100 mL×3) was added to wash the resin, and the solvent was pumped off each time after washing.

[0170] (2-2) Linking S1 amino acid: Fmoc-Gly-OH (32.5 g, 108 mmol) was weighed and dissolved in 100 mL DCM. After dissolved completely, DIEA (23 mL, 135 mmol) was added. The obtained mixture was added in a test tube, nitrogen was introduced, stirred and reacted at 20° C.-30° C. for 2 h. 12 mL of mixed solvent of methanol and DIEA (methanol:DIEA=9:1) was added, and the unreacted sites was sealed for 10 min. The solvent was pumped off. The resin was washed with DCM (150 mL×2). After washing, the solvent was pumped off. Then DMF (150 mL×2) was used to wash the resin, and the solvent was pumped off after washing. The resin was then swelled with 120 mL DMF for 30 min. The solvent was pumped off, and de-protection was performed with a piperidine/DMF solution (120 mL) having a volume ratio of 20% for twice. The times were respectively 10 min and 15 min, and the temperature of the reaction was controlled at 20° C.-30° C. After the de-protection, the solvent was pumped off, and the resin was washed with DMF (120 mL×6). After washing, the solvent was pumped off, and the resin was left in the test tube. Ninhydrin was used to test the color of resin, and the resin was purple-black. The next step was carried out. The absorbance was detected by spectrophotometric method, and degree of substitution of the resin was calculated to be 0.8641 mmol/g.

[0171] (2-3) Linking S2 Amino Acid:

[0172] Fmoc-Gly-OH (19.0 g, 63.9 mmol, 3.0 eq.) and DEPBT (19.17 g, 63.9 mmol, 3.0 eq.) were weighed, and dissolved in 50 mL DMF. After dissolved completely, DIEA (10.5 mL, 63.9 mmol) was added. The obtained mixture was added in a test tube, nitrogen was introduced, stirred and reacted at 20° C.-30° C. for 2 h. Ninhydrin was used to test the color of resin, the resin was transparent yellow. After the reaction was completed, the solvent was pumped off, and the resin was washed with DMF (120 mL×3). The solvent was pumped off after washing, and de-protection was performed with a piperidine/DMF solution (120 mL) having a volume ratio of 20% for twice. The times were respectively 10 min and 15 min, and the temperature of the reaction was controlled at 20° C.-30° C. After the de-protection, the solvent was pumped off, and the resin was washed with DMF (120 mL×6). After washing, the solvent was pumped off, and the resin was left in the test tube. Ninhydrin was used to test the color of resin, and the resin was purple-black. The next step was carried out.

[0173] (2-4) Linking S3 Amino Acid:

[0174] Fmoc-Phe-OH (24.75 g, 63.9 mmol, 3.0 eq.) and DEPBT (19.17 g, 63.9 mmol, 3.0 eq.) were weighed, dissolved in 50 mL DMF. After dissolved completely, DIEA (10.5 mL, 63.9 mmol) was added. The obtained mixture was added in a test tube, nitrogen was introduced, stirred and reacted at 20° C.-30° C. for 2 h. Ninhydrin was used to test the color of resin, the resin was transparent yellow. After the reaction, the solvent was pumped off, the resin was washed with DMF (120 mL×3), and the solvent was pumped off, and then de-protection was performed with a piperidine/DMF solution (120 mL) having a volume ratio of 20% for twice. The times were respectively 10 min and 15 min, and the temperature of the reaction was controlled at 20° C.-30° C. After the de-protection, the solvent was pumped off, and the resin was washed with DMF (120 mL×6). After washing, the solvent was pumped off, and the resin was left in the test tube. Ninhydrin was used to test the color of resin, and the resin was purple-black. The next step was carried out.

[0175] (2-5) Steps (2-4) were repeated. S4 amino acid Fmoc-Gly-OH, S5 amino acid Fmoc-Tyr(t-Bu)-OH, S6 amino acid Fmoc-Ala-OH, S7 amino acid Fmoc-Thr(t-Bu)-OH, S8 amino acid Fmoc-His(trt)-OH, S9 amino acid Fmoc-Leu-OH, S10 amino acid Fmoc-Lys(Boc)-OH, S11 amino acid Fmoc-Aib-OH, S12 amino acid Fmoc-Leu-OH, S13 amino acid Fmoc-Leu-OH, S14 amino acid Fmoc-Lys(Boc)-OH, S15 amino acid Fmoc-Glu(Ot-Bu)-OH, S16 amino acid Fmoc-Leu-OH, S17 amino acid Fmoc-Leu-OH, S18 amino acid Fmoc-Glu(Ot-Bu)-OH, S19 amino acid Fmoc-Arg(pbf)-OH, S20 amino acid Fmoc-Arg(pbf)-OH, S21 amino acid Fmoc-Arg(pbf)-OH, S22 amino acid Fmoc-Leu-OH, S23 amino acid Fmoc-Asp(Ot-Bu)-OH, S24 amino acid Fmoc-Gln(trt)-OH, S25 amino acid Fmoc-Ile-OH, S26 amino acid Fmoc-Ser(t-Bu)-OH, S27 amino acid Fmoc-Lys(Boc)-OH, S28 amino acid Fmoc-Gly-OH, S29 amino acid Fmoc-Lys(Boc)-OH, S30 amino acid Fmoc-Asp(Ot-Bu)-OH, S31 amino acid Fmoc-His(trt)-OH, S32 amino acid Fmoc-Leu-OH, S33 amino acid Fmoc-Leu-OH, S34 amino acid Fmoc-Gln(trt)-OH, S35 amino acid Fmoc-His(trt)-OH, S36 amino acid Fmoc-Glu(Ot-Bu)-OH, S37 amino acid Fmoc-Ser(t-Bu)-OH, S38 amino acid Fmoc-Val-OH and S39 amino acid Fmoc-Ala-OH were successively linked, and a peptide resin was obtained to carry out the next operation.

[0176] (2-6) Resin shrinkage: methanol (80 mL) was firstly added in the test tube, the resin was shrunken for 5 min, and the solvent was pumped off. The shrinkage was repeated for 3 times, 10 min once. Each time after the shrinkage, the solvent was pumped out completely before the next shrinkage. Then the shrunken resin was placed in a vacuum drying oven, dried at 35° C., and 18.82 g peptide resin was obtained.

[0177] (2-7) Peptide segment cracking: 155 mL TFA, 8 mL TIS, 4.12 mL EDT, 2 mL TA, 4.12 mL water and 2 mL anisole were mixed evenly to prepare the lysate. 18.82 g of the peptide resin prepared in steps (2-6) was weighed, the lysate and the peptide resin were mixed, sealed and shielded from the light. The mixture was stirred and reacted, the temperature was kept at 25° C.-35° C., and reacted for 2 h. After the reaction, a sand core funnel was used to remove the resin. After removing the solvent in vacuum, methyl tertiary butyl ether (450 mL) was added in the rest liquid, and crystallization was performed at low temperature (0° C.-10° C.) for 2 h. The resultant mixture was centrifuged to remove the crystallizing solution, and the precipitate was washed with methyl tertiary butyl ether for 3 times. The precipitate was collected, and dried in vacuum at 35° C., and a polypeptide (SEQ ID NO:10) Ala-Val-Ser-Glu-His-Gln-Leu-Leu-His-Asp-Lys-Gly-Lys-Ser-Ile-Gln-Asp-Leu-Arg-Arg-Arg-Glu-Leu-Leu-Glu-Lys-Leu-Leu-Aib-Lys-Leu-His-Thr-Ala-Tyr-Gly-Phe-Gly-Gly was obtained.

[0178] (2-8) Purification: after filtering the peptide crude solution obtained in steps (2-7) with a 0.45 μm filter membrane, the solution was subjected to preparative HPLC purification in a 20 mm×150 mm column filled with 10 μm C-18 silica gel. The detection wavelength was 220 nm. The mobile phase A was 0.1% TFA, and the mobile phase B was acetonitrile. Gradient elution was carried out according to the following Table A.

TABLE-US-00007 TABLE A Gradient elution program Flow rate Mobile Mobile Time (mL/ phase phase (min) min) A (%) B (%) 0 8 95 5 0.1 8 75 25 45 8 65 50 60 8 50 50

[0179] Fractions containing target polypeptide product were collected, and the purity was 95.8%. The collected fractions were combined, the solvent was removed in vacuum, and the polypeptide compound was freeze-dried. The obtained end product was identified by analytical RP-HPLC (retention time), LC-MS and MALDI/TOF-MS.

[0180] LC-MS(ESI): m/z 1112.2 [M/4+H].sup.+.

[0181] MALDI/TOF-MS(ESI): m/z 4441.4175 [M+H].sup.+.

[0182] According to QE identification and analysis, the sequence of the obtained polypeptide compound was as that shown as SEQ ID NO:10.

[0183] Assay: moisture content was measured with a moisture titrator by the moisture determination method in Chinese Pharmacopoeia, and TFA content was measured by the acetic acid content detection method in Chinese Pharmacopoeia, and the content of polypeptide was 85.9%.

TABLE-US-00008 Example 3: (the polypeptide sequence shown as SEQ ID No. 11) Preparation of Ala-Val-Ser-Glu-His-Gln-Leu-Leu- His-Asp-Lys-Gly-Lys-Ser-Ile-Gln-Asp-Leu-Arg-Arg- Arg-Glu-Leu-Leu-Glu-Lys-Leu-Leu-Ala-Lys-Leu-His- Thr-Ala-Tyr-Arg-Gly-Phe-Gly-Gly

[0184] The compound was synthesized from C terminal to N terminal:

[0185] (3-1) Preprocessing of resin: 30 g 2-CTA resin was weighed (degree of substitution 0.93 mmol/g, 27 mmol), placed in a 250 mL test tube, and swelled in DCM (200 mL) for 30 min. The solvent was pumped out in vacuum. Then DCM (100 mL) was added, nitrogen was introduced, and heated until the temperature reached 25° C. SOCl.sub.2 (10 mL, 5.0 eq) was added dropwise, the temperature was kept at 30-35° C., and reacted for 2 h. After the reaction was completed, nitrogen was introduced and the solvent was pumped off. DCM (100 mL×3) was added to wash the resin, and the solvent was pumped off each time after washing.

[0186] (3-2) Linking S1 amino acid: Fmoc-Gly-OH (32.5 g, 108 mmol) was weighed and dissolved in 100 mL DCM. After dissolved completely, DIEA (23 mL, 135 mmol) was added. The obtained mixture was added in a test tube, nitrogen was introduced, stirred and reacted at 20° C.-30° C. for 2 h. 12 mL of mixed solvent of methanol and DIEA (methanol:DIEA=9:1) was added dropwise, and the unreacted sites were sealed for 10 min. The solvent was pumped off. The resin was washed with DCM (150 mL×2). After washing, the solvent was pumped off. Then DMF (150 mL×2) was used to wash the resin, and the solvent was pumped off after washing. The resin was then swelled with 120 mL DMF for 30 min. The solvent was pumped off, and de-protection was performed with a piperidine/DMF solution (120 mL) having a volume ratio of 20% for twice. The times were respectively 10 min and 15 min, and the temperature of the reaction was controlled at 20° C.-30° C. After the de-protection, the solvent was pumped off, and the resin was washed with DMF (120 mL×6). After washing, the solvent was pumped off, and the resin was left in the test tube. Ninhydrin was used to test the color of resin, and the resin was purple-black. The next step was carried out. The absorbance was detected by spectrophotometric method, and degree of substitution of the resin was calculated to be 0.8641 mmol/g.

[0187] (3-3) Linking S2 Amino Acid:

[0188] Fmoc-Gly-OH (19.0 g, 63.9 mmol, 3.0 eq.) and T.sub.3P (20.33 g, 63.9 mmol, 3.0 eq.) were weighed, and dissolved in 50 mL DMF. After dissolved completely, DIEA (10.5 mL, 63.9 mmol) was added. The obtained mixture was added in a test tube, nitrogen was introduced, stirred and reacted at 20° C.-30° C. for 2 h. Ninhydrin was used to test the color of resin, the resin was transparent yellow. After the reaction was completed, the solvent was pumped off, and the resin was washed with DMF (120 mL×3), 3 min each time. The solvent was pumped off after washing, and de-protection was performed with a piperidine/DMF solution (120 mL) having a volume ratio of 23% for twice. The times were respectively 10 min and 15 min, and the temperature of the reaction was controlled at 20° C.-30° C. After the de-protection, the solvent was pumped off, and the resin was washed with DMF (120 mL×6). After washing, the solvent was pumped off, and the resin was left in the test tube. Ninhydrin was used to test the color of resin, and the resin was purple-black. The next step was carried out.

[0189] (3-4) Linking S3 Amino Acid

[0190] Fmoc-Phe-OH (24.75 g, 63.9 mmol, 3.0 eq.) and T.sub.3P (19.17 g, 63.9 mmol, 3.0 eq.) were weighed, dissolved in 50 mL DMF. After dissolved completely, DIEA (10.5 mL, 63.9 mmol) was added. The obtained mixture was added in a test tube, nitrogen was introduced, stirred and reacted at 20° C.-30° C. for 2 h. Ninhydrin was used to test the color of resin, the resin was transparent yellow. After the reaction was completed, the solvent was pumped off, the resin was washed with DMF (120 mL×3), and the solvent was pumped off after washing, and then de-protection was performed with a piperidine/DMF solution (120 mL) having a volume ratio of 20% for twice. The times were respectively 10 min and 15 min, and the temperature of the reaction was controlled at 20° C.-30° C. After the de-protection, the solvent was pumped off, and the resin was washed with DMF (120 mL×6). After washing, the solvent was pumped off, and the resin was left in the test tube. Ninhydrin was used to test the color of resin, and the resin was purple-black. The next step was carried out.

[0191] (3-5) Steps (3-4) were repeated. S4 amino acid Fmoc-Gly-OH, S5 amino acid Fmoc-Arg(pbf)-OH, S6 amino acid Fmoc-Tyr(t-Bu)-OH, S7 amino acid Fmoc-Ala-OH, S8 amino acid Fmoc-Thr(t-Bu)-OH, S9 amino acid Fmoc-His(trt)-OH, S10 amino acid Fmoc-Leu-OH, S11 amino acid Fmoc-Lys(Boc)-OH, S12 amino acid Fmoc-Ala-OH, S13 amino acid Fmoc-Leu-OH, S14 amino acid Fmoc-Leu-OH, S15 amino acid Fmoc-Lys(Boc)-OH, S16 amino acid Fmoc-Glu(Ot-Bu)-OH, S17 amino acid Fmoc-Leu-OH, S18 amino acid Fmoc-Leu-OH, S19 amino acid Fmoc-Glu(Ot-Bu)-OH, S20 amino acid Fmoc-Arg(pbf)-OH, S21 amino acid Fmoc-Arg(pbf)-OH, S22 amino acid Fmoc-Arg(pbf)-OH, S23 amino acid Fmoc-Leu-OH, S24 amino acid Fmoc-Asp(Ot-Bu)-OH, S25 amino acid Fmoc-Gln(trt)-OH, S26 amino acid Fmoc-Ile-OH, S27 amino acid Fmoc-Ser(t-Bu)-OH, S28 amino acid Fmoc-Lys(Boc)-OH, S29 amino acid Fmoc-Gly-OH, S30 amino acid Fmoc-Lys(Boc)-OH, S31 amino acid Fmoc-Asp(Ot-Bu)-OH, S32 amino acid Fmoc-His(trt)-OH, S33 amino acid Fmoc-Leu-OH, S34 amino acid Fmoc-Leu-OH, S35 amino acid Fmoc-Gln(trt)-OH, S36 amino acid Fmoc-His(trt)-OH, S37 amino acid Fmoc-Glu(Ot-Bu)-OH, S38 amino acid Fmoc-Ser(t-Bu)-OH, S39 amino acid Fmoc-Val-OH and S40 amino acid Fmoc-Ala-OH were successively connected, and a peptide resin was obtained to carry out the next operation.

[0192] (3-6) Resin shrinkage: methanol (80 mL) was firstly added in the test tube, the resin was shrunken for 5 min, and the solvent was pumped off. The shrinkage was repeated for 3 times, 10 min once. Each time after the shrinkage, the solvent was pumped out completely before the next shrinkage. Then the shrunken resin was placed in a vacuum drying oven, dried at 35° C., and 19.56 g peptide resin was obtained.

[0193] (3-7) Peptide segment cracking: 155 mL TFA, 8 mL TIS, 4.12 mL EDT, 2 mL TA, 4.12 mL water and 2 mL anisole were mixed evenly to prepare the lysate. 18.82 g of the peptide resin prepared in steps (3-6) was weighed, the lysate and the peptide resin were mixed, sealed and shielded from the light. The mixture was stirred and reacted, the temperature was kept at 25° C.-35° C., and reacted for 2 h. After the reaction was completed, a sand core funnel was used to remove the resin. After removing the solvent in vacuum, methyl tertiary butyl ether (450 mL) was added in the rest liquid, and crystallization was performed at low temperature (0° C.-10° C.) for 2 h. The mixture was centrifuged to remove the crystallizing solution, and the obtained precipitate was washed with methyl tertiary butyl ether for 3 times. The precipitate was collected, and dried in vacuum at 35° C., to obtain polypeptide (SEQ ID NO:11) Ala-Val-Ser-Glu-His-Gln-Leu-Leu-His-Asp-Lys-Gly-Lys-Ser-Ile-Gln-Asp-Leu-Arg-Arg-Arg-Glu-Leu-Leu-Glu-Lys-Leu-Leu-Ala-Lys-Leu-His-Thr- Ala-Tyr-Arg-Gly-Phe-Gly-Gly.

[0194] (3-8) Purification: after filtering the peptide crude solution obtained in steps (3-7) with a 0.45 μm filter membrane, the mixture was subjected to preparative HPLC purification in a 20 mm×150 mm column filling with 10 μm C-18 silica gel. The detection wavelength was 220 nm. The mobile phase A was 0.1% TFA, and the mobile phase B was acetonitrile. Gradient elution was carried out according to the following Table A.

TABLE-US-00009 TABLE A Gradient elution program Flow rate Mobile Mobile Time (mL/ phase phase (min) min) A(%) B(%) 0 8 95 5 0.1 8 75 25 45 8 65 50 60 8 50 50

[0195] Fractions containing target polypeptide product were collected, and the purity was 96.1%. The collected fractions were combined, the solvent was removed in vacuum, and the polypeptide compound was freeze-dried. The obtained end product was identified by analytical RP-HPLC (retention time) and MALDI/TOF-MS.

[0196] MALDI/TOF-MS(ESI): m/z 4585.2 [M+H].sup.+.

[0197] According to QE identification and analysis, the sequence of the obtained polypeptide compound was as that shown as SEQ ID NO: 11.

[0198] Assay: moisture content was measured with a moisture titrator by the moisture determination method in Chinese Pharmacopoeia, and TFA content was measured by the acetic acid content detection method in Chinese Pharmacopoeia, and the content of polypeptide was 81.97%.

TABLE-US-00010 Example 4: (the polypeptide sequence shown as SEQ ID No. 12) Preparation of Ala-Val-Ser-Glu-His-Gln-Leu-Leu- His-Asp-Lys-Gly-Lys-Ser-Ile-Gln-Asp-Leu-Arg-Arg- Arg-Phe-Phe-Leu-His-His-Leu-Ile-Ala-Glu-Ile-His- Thr-Ala-Tyr-Gly-Phe-Gly-Gly

[0199] The process as shown in Example 2 was adopted. 2-CTA resin having a degree of substitution of 0.93 mmol/g was used. The resin was firstly swelled to prepare 2-CTA resin into CTC resin. Then the side chain-protected amino acids Fmoc-Gly-OH, Fmoc-Gly-OH, Fmoc-Phe-OH, Fmoc-Gly-OH, Fmoc-Tyr(t-Bu)-OH, Fmoc-Ala-OH, Fmoc-Thr(t-Bu)-OH, Fmoc-His(trt)-OH, Fmoc-Ile-OH, Fmoc-Glu(Ot-Bu)-OH, Fmoc-Ala-OH, Fmoc-Ile-OH, Fmoc-Leu-OH, Fmoc-His(trt)-OH, Fmoc-His(trt)-OH, Fmoc-Leu-OH, Fmoc-Phe-OH, Fmoc-Phe-OH, Fmoc-Arg(pbf)-OH, Fmoc-Arg(pbf)-OH, Fmoc-Arg(pbf)-OH, Fmoc-Leu-OH, Fmoc-Asp(Ot-Bu)-OH, Fmoc-Gln(trt)-OH, Fmoc-Ile-OH, Fmoc-Ser(t-Bu)-OH, Fmoc-Lys(Boc)-OH, Fmoc-Gly-OH, Fmoc-Lys(Boc)-OH, Fmoc-Asp(Ot-Bu)-OH, Fmoc-His(trt)-OH, Fmoc-Leu-OH, Fmoc-Leu-OH, Fmoc-Gln(trt)-OH, Fmoc-His(trt)-OH, Fmoc-Glu(Ot-Bu)-OH, Fmoc-Ser(t-Bu)-OH, Fmoc-Val-OH, Fmoc-Ala-OH were successively linked to give the peptide resin. Finally, a lysate prepared by evenly mixing TFA, TIS, EDT, TA, water and anisole was used to treat the peptide resin. The side chain protective groups were removed, and the resin cracked at the same time, to give a crude peptide product (SEQ ID NO: 12): Ala-Val-Ser-Glu-His-Gln-Leu-Leu-His-Asp-Lys-Gly-Lys-Ser-Ile-Gln-Asp-Leu-Arg-Arg-Arg-Phe-Phe-Leu-His-His-Leu-Ile-Ala-Glu-Ile-His-Thr-Ala-Tyr-Gly-Phe-Gly-Gly.

[0200] The polypeptide crude product was purified by reverse phase preparative high pressure liquid phase chromatography (HPLC), the detection wavelength was 220 nm, the mobile phase A was 0.1% TFA, and the mobile phase B was acetonitrile. The fractions containing pure products were combined, and freeze-dried to give the polypeptide product. The purity detected by HPLC was 94.8%. The obtained end product was identified by MALDI/TOF-MS.

[0201] MALDI/TOF-MS(ESI) (ESI): m/z 4450.12 [M+H].sup.+.

[0202] According to QE identification and analysis, the sequence of the obtained polypeptide compound was as that shown as SEQ ID NO: 12.

TABLE-US-00011 Example 5: (the polypeptide sequence shown as SEQ ID No. 13) Preparation of Ala-Val-Ser-Glu-His-Gln-Leu-Leu- His-Asp-Lys-Gly-Lys-Ser-Ile-Gln-Asp-Leu-Arg-Arg- Arg-Phe-Phe-Leu-His-His-Leu-Ile-Aib-Glu-Ile-His- Thr-Ala-Tyr-Arg-Phe-Gly-Gly

[0203] The process as shown in Example 2 was adopted. 2-CTA resin having a degree of substitution of 0.93 mmol/g was used. The resin was firstly swelled to prepare 2-CTA resin into CTC resin. Then the side chain-protected amino acids Fmoc-Gly-OH, Fmoc-Gly-OH, Fmoc-Phe-OH, Fmoc-Arg(pbf)-OH, Fmoc-Tyr(t-Bu)-OH, Fmoc-Ala-OH, Fmoc-Thr(t-Bu)-OH, Fmoc-His(trt)-OH, Fmoc-Ile-OH, Fmoc-Glu(Ot-Bu)-OH, Fmoc-Aib-OH, Fmoc-Ile-OH, Fmoc-Leu-OH, Fmoc-His(trt)-OH, Fmoc-His(trt)-OH, Fmoc-Leu-OH, Fmoc-Phe-OH, Fmoc-Phe-OH, Fmoc-Arg(pbf)-OH, Fmoc-Arg(pbf)-OH, Fmoc-Arg(pbf)-OH, Fmoc-Leu-OH, Fmoc-Asp(Ot-Bu)-OH, Fmoc-Gln(trt)-OH, Fmoc-Ile-OH, Fmoc-Ser(t-Bu)-OH, Fmoc-Lys(Boc)-OH, Fmoc-Gly-OH, Fmoc-Lys(Boc)-OH, Fmoc-Asp(Ot-Bu)-OH, Fmoc-His(trt)-OH, Fmoc-Leu-OH, Fmoc-Leu-OH, Fmoc-Gln(trt)-OH, Fmoc-His(trt)-OH, Fmoc-Glu(Ot-Bu)-OH, Fmoc-Ser(t-Bu)-OH, Fmoc-Val-OH and Fmoc-Ala-OH were successively linked to give the peptide resin. Finally, a lysate prepared by evenly mixing TFA, TIS, EDT, TA, water and anisole was used to treat the peptide resin. The side chain protective groups were removed, and the resin cracked at the same time, to give a crude peptide product (SEQ ID NO: 13): Ala-Val-Ser-Glu-His-Gln-Leu-Leu-His-Asp-Lys-Gly-Lys-Ser-Ile-Gln-Asp-Leu-Arg-Arg-Arg-Phe-Phe-Leu-His-His-Leu-Ile-Aib-Glu-Ile-His-Thr-Ala-Tyr-Arg-Phe- Gly-Gly.

[0204] The polypeptide crude product was purified by reverse phase preparative high pressure liquid phase chromatography (HPLC), the detection wavelength was 220 nm, the mobile phase A was 0.1% TFA, and the mobile phase B was acetonitrile. The fractions containing pure products were combined, and freeze-dried to give the polypeptide product. The purity detected by HPLC was 94.8%. The obtained end product was identified by MALDI/TOF-MS.

[0205] MALDI/TOF-MS(ESI): m/z 4599.253 [M+H].sup.+.

[0206] According to QE identification and analysis, the sequence of the obtained polypeptide compound was as that shown as SEQ ID NO: 13.

TABLE-US-00012 Example 6: (the polypeptide sequence shown as SEQ ID No. 14) Preparation of Ala-Val-Ser-Glu-His-Gln-Leu-Ile- His-Asp-Lys-Gly-Lys-Ser-Ile-Gln-Glu-Leu-Arg-Arg- Arg-Phe-Phe-Leu-His-His-Leu-Ile-Aib-Glu-Ile-His- Thr-Ala-Tyr-Gly-Phe-Gly-Gly

[0207] The process as shown in Example 2 was adopted. 2-CTA resin having a degree of substitution of 0.93 mmol/g was used. The resin was firstly swelled to prepare 2-CTA resin into CTC resin. Then the side chain-protected amino acids Fmoc-Gly-OH, Fmoc-Gly-OH, Fmoc-Phe-OH, Fmoc-Gly-OH, Fmoc-Tyr(t-Bu)-OH, Fmoc-Ala-OH, Fmoc-Thr(t-Bu)-OH, Fmoc-His(trt)-OH, Fmoc-Ile-OH, Fmoc-Glu(Ot-Bu)-OH, Fmoc-Aib-OH, Fmoc-Ile-OH, Fmoc-Leu-OH, Fmoc-His(trt)-OH, Fmoc-His(trt)-OH, Fmoc-Leu-OH, Fmoc-Phe-OH, Fmoc-Phe-OH, Fmoc-Arg(pbf)-OH, Fmoc-Arg(pbf)-OH, Fmoc-Arg(pbf)-OH, Fmoc-Leu-OH, Fmoc-Glu(Ot-Bu)-OH, Fmoc-Gln(trt)-OH, Fmoc-Ile-OH, Fmoc-Ser(t-Bu)-OH, Fmoc-Lys(Boc)-OH, Fmoc-Gly-OH, Fmoc-Lys(Boc)-OH, Fmoc-Asp(Ot-Bu)-OH, Fmoc-His(trt)-OH, Fmoc-Ile-OH, Fmoc-Leu-OH, Fmoc-Gln(trt)-OH, Fmoc-His(trt)-OH, Fmoc-Glu(Ot-Bu)-OH, Fmoc-Ser(t-Bu)-OH, Fmoc-Val-OH and Fmoc-Ala-OH were successively linked to give the peptide resin. Finally, a lysate prepared by evenly mixing TFA, TIS, EDT, TA, water and anisole was used to treat the peptide resin. The side chain protective groups were removed, and the resin cracked at the same time, to give a crude peptide product (SEQ ID NO: 14): Ala-Val-Ser-Glu-His-Gln-Leu-Ile-His-Asp-Lys-Gly-Lys-Ser-Ile-Gln-Glu-Leu-Arg-Arg-Arg-Phe-Phe-Leu-His-His-Leu-Ile-Aib-Glu-Ile-His-Thr-Ala-Tyr-Gly-Phe-Gly-Gly.

[0208] The polypeptide crude product was purified by reverse phase preparative high pressure liquid phase chromatography (HPLC), the detection wavelength was 220 nm, the mobile phase A was 0.1% TFA, and the mobile phase B was acetonitrile. The fractions containing pure products were combined, and freeze-dried to give the polypeptide product. The purity detected by HPLC was 94.8%. The obtained end product was identified by MALDI/TOF-MS.

[0209] MALDI/TOF-MS(ESI): m/z 4514.15 [M+H].sup.+.

[0210] According to QE identification and analysis, the sequence of the obtained polypeptide compound was as that shown as SEQ ID NO: 14.

TABLE-US-00013 Example 7: (the polypeptide sequence shown as SEQ ID No. 15) Preparation of Ala-Val-Ser-Glu-His-Gln-Leu-Ile- His-Asp-Lys-Gly-Lys-Ser-Ile-Gln-Glu-Leu-Arg-Arg- Arg-Phe-Phe-Leu-His-His-Leu-Leu-Ala-Glu-Ile-His- Thr-Ala-Tyr-Gly-Phe-Gly-Gly

[0211] The process as shown in Example 2 was adopted. 2-CTA resin having a degree of substitution of 0.93 mmol/g was used. The resin was firstly swelled to prepare 2-CTA resin into CTC resin. Then the side chain-protected amino acids Fmoc-Gly-OH, Fmoc-Gly-OH, Fmoc-Phe-OH, Fmoc-Gly-OH, Fmoc-Tyr(t-Bu)-OH, Fmoc-Ala-OH, Fmoc-Thr(t-Bu)-OH, Fmoc-His(trt)-OH, Fmoc-Ile-OH, Fmoc-Glu(Ot-Bu)-OH, Fmoc-Ala-OH, Fmoc-Leu-OH, Fmoc-Leu-OH, Fmoc-His(trt)-OH, Fmoc-His(trt)-OH, Fmoc-Leu-OH, Fmoc-Phe-OH, Fmoc-Phe-OH, Fmoc-Arg(pbf)-OH, Fmoc-Arg(pbf)-OH, Fmoc-Arg(pbf)-OH, Fmoc-Leu-OH, Fmoc-Glu(Ot-Bu)-OH, Fmoc-Gln(trt)-OH, Fmoc-Ile-OH, Fmoc-Ser(t-Bu)-OH, Fmoc-Lys(Boc)-OH, Fmoc-Gly-OH, Fmoc-Lys(Boc)-OH, Fmoc-Asp(Ot-Bu)-OH, Fmoc-His(trt)-OH, Fmoc-Ile-OH, Fmoc-Leu-OH, Fmoc-Gln(trt)-OH, Fmoc-His(trt)-OH, Fmoc-Glu(Ot-Bu)-OH, Fmoc-Ser(t-Bu)-OH, Fmoc-Val-OH and Fmoc-Ala-OH were successively linked to give the peptide resin. Finally, a lysate prepared by evenly mixing TFA, TIS, EDT, TA, water and anisole was used to treat the peptide resin. The side chain protective groups were removed, and the resin cracked at the same time, to give a crude peptide product (SEQ ID NO: 15): Ala-Val-Ser-Glu-His-Gln-Leu-Ile-His-Asp-Lys-Gly-Lys-Ser-Ile-Gln-Glu-Leu-Arg-Arg-Arg-Phe-Phe-Leu-His-His-Leu-Leu-Ala-Glu-Ile-His-Thr-Ala-Tyr-Gly-Phe-Gly-Gly.

[0212] The polypeptide crude product was purified by revers phase preparative high pressure liquid phase chromatography (HPLC), the detection wavelength was 220 nm, the mobile phase A was 0.1% TFA, and the mobile phase B was acetonitrile. The fractions containing pure products were combined, and freeze-dried to give the polypeptide product. The purity detected by HPLC was 94.8%. The obtained end product was identified by MALDI/TOF-MS.

[0213] MALDI/TOF-MS(ESI): m/z 4514.15 [M+H].sup.+.

[0214] According to QE identification and analysis, the sequence of the obtained polypeptide compound was as that shown as SEQ ID NO: 15.

Examples of Effect

(I) Research on Osteoporosis Therapeutic Effect of the Compound in the Present Disclosure on Ovary Removed SD Rats

[0215] Experimental method: female SD rats aged around 22 weeks were used in the experiment. Feeding conditions: the animal room had a temperature of 21±5° C., and a relative humidity of 35±10%; and the animal room was exposed to light for 12 h and darkness for 12 h in each day. The animals had free access to water. The SD rats were subjected to ovariectomy (OVX), then fed for another 3 months to be induced to osteoporosis model. The rats were grouped according to bone mineral density of thigh bone: {circle around (1)} Sham group: normal saline of the same volume was subcutaneously administered; {circle around (2)} OVX group (model group): normal saline of the same volume was subcutaneously administered; {circle around (3)} 5 μg dosage group of positive drug abaloptide (Aba-5): 5 μg/kg abaloptide (Aba) was subcutaneously administered. Administration groups of three dosages were set for each test compound. 2.5 μg/kg dosage group of the test compound: 2.5 μg/kg test compound was subcutaneously administered. 5 μg/kg dosage group of the test compound: 5 μg/kg test compound was subcutaneously administered. 10 μg/kg dosage group of the test compound: 10 μg/kg test compound was subcutaneously administered. The administration was carried out 5 times a week, successively lasted for 25 weeks. The test compounds were polypeptide compounds in examples 1-5 (as shown in SEQ ID NO:9-SEQ ID NO:13). Abaloptide was a drug for treating post-menopausal osteoporosis in the market.

[0216] Detection method: times and contents of detection indicators were shown hereinafter.

[0217] 1) Bone mineral density (BMD) was detected with a dual-energy X-ray bone densitometry (DXA) before model establishment (−13 w), before administration (0 w), and 6 w, 12 w, 25 w, 37 w and 49 w after administration.

[0218] 2) Blood was collected from eye orbit before administration (0 w) and 25 w after administration, and a blood routine examination was carried out with whole blood.

[0219] 3) 25 w after administration, the serum was kept, and bone turnover markers (CTx (β-CTX, serum), OC (serum) and serum procollagen type 1 N-terminal propeptide (PINP)) were detected, and calcium, phosphorus and alkaline phosphatase (ALP) were detected.

[0220] 4) After administration, thigh bones of both sides and lumbar vertebra of rats were collected, and subjected to MicroCT, histomorphology detection and biomechanics detections.

[0221] Statistical analysis of data: the data were treated with SPSS 20 software. The data in line with normality test were subjected to one-way analysis of variance; and the data not in line with normality test were subjected to rank sum test. The data were represented by mean value±standard error.

Experimental Results

[0222] (1) The bone mineral densities of thigh and lumbar vertebra were counted, and percentages of density changes of thigh bone and lumbar vertebra at each detection time point were calculated, shown in Table 1.

Percentage of bone mineral density change=(bone mineral density at the detection time point−bone density before administration)/bone density before administration*100%

TABLE-US-00014 TABLE 1 Bone mineral density and percentages of change of rats in each group 0 w Bone mineral density 25 w Bone mineral density (g/cm.sup.2) (g/cm.sup.2) Dosage lumbar lumbar 0-25 w percentage of change % Group (μg/kg) thigh bone vertebra thigh bone vertebra thigh bone lumbar vertebra Sham — 0.26 ± 0 0.28 ± 0.01 0.28 ± 0 0.29 ± 0.01 7.6 ± 2.03 4.73 ± 3.19 Model/OVX — 0.23 ± 0*** 0.25 ± 0*** 0.22 ± 0*** 0.21 ± 0*** −3.77 ± 1.19*** −14.41 ± 1.58*** OVX + Aba 5 0.23 ± 0*** 0.24 ± 0*** 0.3 ± 0*### 0.3 ± 0### 30.28 ± 2***### 21.66 ± 2.12***### 2.5 0.23 ± 0.01** 0.21 ± 0.00*** 0.28 ± 0.01* 0.23 ± 0.01* 30.75 ± 3.69# 2.82 ± 4.46 OVX + 5 0.23 ± 0.01** 0.22 ± 0.00*** 0.30 ± 0.01## 0.27 ± -0.01 37.67 ± 2.70## 24.82 ± 4.93## Example 1 10 0.25 ± 0.01** 0.22 ± 0.00*** 0.31 ± 0.01# 0.27 ± 0.01# 38.15 ± 4.87# 24.91 ± 7.23# 2.5 0.23 ± 0*** 0.24 ± 0*** 0.28 ± 0### 0.29 ± 0.01### 22.21 ± 1.54***### 18.49 ± 2.13### OVX + 5 0.23 ± 0*** 0.24 ± 0*** 0.3 ± 0### 0.31 ± 0.01### 29.17 ± 1.86***### 29.61 ± 2.26***### Example 2 10 0.23 ± 0*** 0.24 ± 0.01*** 0.31 ± 0***### 0.31 ± 0.01### 33.68 ± 1.84***### 30.94 ± 2.24***### 2.5 0.23 ± 0*** 0.24 ± 0*** 0.26 ± 0### 0.27 ± 0### 14.03 ± 1.81***### 12.4 ± 2.2## OVX + 5 0.23 ± 0*** 0.24 ± 0*** 0.29 ± 0.02### 0.29 ± 0.01### 27.19 ± 1.33***### 20.41 ± 2.56### Example 3 10 0.23 ± 0*** 0.24 ± 0.01*** 0.29 ± 0### 0.29 ± 0.01### 28.7 ± 1.6***### 22.46 ± 2.25### 2.5 0.23 ± 0*** 0.24 ± 0*** 0.27 ± 0### 0.28 ± 0.01### 17.23 ± 1.4***### 14.05 ± 2.79*### OVX + 5 0.23 ± 0*** 0.24 ± 0*** 0.28 ± 0### 0.31 ± 0.01### 31.61 ± 1.54***### 25.79 ± 2.06***### Example 4 10 0.23 ± 0*** 0.24 ± 0.01*** 0.29 ± 0*### 0.32 ± 0.01### 35.37 ± 1.94***### 24.06 ± 2.19***### 2.5 0.23 ± 0.01** 0.21 ± 0.00*** 0.3 ± 0.01# 0.26 ± 0.01* 40.87 ± 4.24# 18.36 ± 4.34 OVX + 5 0.23 ± 0.01** 0.22 ± 0.00*** 0.32 ± 0.01## 0.29 ± 0.02## 46.29 ± 2.42## 34.29 ± 7.15## Example 5 10 0.25 ± 0.01** 0.22 ± 0.00*** 0.3 ± 0.01# 0.27 ± 0.01# 35.57 ± 3.17# 32.67 ± 5.38# Note: compared with Sham group, *represented for P < 0.05, **represented for P < 0.01, and ***represented for P < 0.001; compared with OVX group, #represented for P < 0.05, ##represented for P < 0.01, and ###represented for P < 0.001; and compared with Aba-5 μ/kg group, & represented for P < 0.05, && represented for P < 0.01, and &&& represented for P < 0.001.

[0223] Result and discussion: on the base of Table 1, compared with OVX model group, the test compound groups of examples 1-5 were administered at dosages of 2.5 μg/kg, 5 μg/kg and 10 μg/kg for 25 weeks, bone mineral densities of thigh bone and lumbar vertebra significantly increased in OVX induced rat osteoporosis model. In addition, there was a dose-response relationship between the test compounds in examples 1-5 and the increase of bone mineral density of osteoporosis rat.

[0224] Compared with sham group, the test compound groups of examples 1-5 were administered at dosages of 2.5 μg/kg, 5 μg/kg and 10 μg/kg for 25 weeks, bone mineral densities of thigh bone and lumbar vertebra of osteoporosis rat increased, and there was no significant difference in bone mineral density compared with the bone mineral density of sham group in the end. In some administration groups, the bone mineral densities of rats were even higher than that of the sham group, indicating that 25 weeks after administration, bone mineral densities of thigh bone and lumbar vertebra were already close to normal level. This demonstrated that the polypeptide compound in the present disclosure can facilitate ossification, and increase bone mineral density.

[0225] After administering the marketed drug abaloptide at a dosage of 5 μg/kg for 25 weeks, the bone mineral density of thigh bone of osteoporosis rat increased by 30.28±2%. When the test compounds 1-5 were administered at a dosage of 5 μg/kg, the bone mineral density of thigh bone of osteoporosis rat respectively increased by 37.67±2.70%, 29.17±1.86%, 27.19±1.33%, 31.61±1.54 and 46.29±2.42%. After administering the marketed drug abaloptide at a dosage of 5 μg/kg for 25 weeks, the bone mineral density of lumbar vertebra of osteoporosis rat increased by 21.66±2.12%. When the test compounds 1-5 were administered at a dosage of 5 μg/kg, the bone mineral density of lumbar vertebra of osteoporosis rat respectively increased by 24.82±4.93%, 29.61±2.26%, 20.41±2.56%, 25.79±2.06% and 34.29±7.15%.

[0226] (2) Blood routine examination results of animals in each group were counted, shown in Table 2.

TABLE-US-00015 TABLE 2 immunity-related indicators of peripheral blood of rats in each group immunity-related indicators of peripheral blood Dosage WBC Lymph Gran Gran/ Group (μg/kg) (10.sup.9/L) (10.sup.9/L) (10.sup.9/L) lymph Sham — 6.32 ± 3.48 ± 2.57 ± 73.79 ± 0.38 0.16 0.25 6.5 Model/ — 6.25 ± 3.89 ± 2.12 ± 55.16 ± OVX 0.38 0.23 0.16 2.96 OVX+ 5 4.31 ± 2.72 ± 1.45 ± 53.27 ± Aba 0.22*## 0.13### 0.09* 2.19* OVX+ 2.5 6.47 ± 3.85 ± 2.13 ± 74.12 ± Exam- 1.1&& 0.96&& 0.44& 0.17&& ple 1 5 6.06 ± 3.12 ± 2.72 ± 70.30 ± 1.26&& 0.99 0.44&&& 0.15&& 10 6.98 ± 3.4 ± 2.17 ± 71.22 ± 0.88&&& 0.58 0.488z 0.11&& OVX + 2.5 6.13 ± 3.63 ± 2.27 ± 64.17 ± Exam- 0.32&& 0.21&& 0.16& 4.328z ple 2 5 6.31 ± 3.4 ± 2.7 ± 75.15 ± 0.4&& 0.18 0.27&&& 5.87#&& 10 6.61 ± 3.7 ± 2.7 ± 67.79 ± 0.47&&& 0.21&& 0.27&&& 4.48 OVX + 2.5 6.65 ± 3.68 ± 2.36 ± 74.77 ± Exam- 0.63&& 0.27&& 0.33& 4.59##&&& ple 3 5 6.55 ± 3.97 ± 2.92 ± 72.27 ± 0.75&& 0.28&&& 0.43#&&& 5.29#&& 10 6.12 ± 3.7 ± 2.03 ± 70.35 ± 0.47&& 0.27&& 0.26& 4.41#&& OVX + 2.5 6.5 ± 3.29 ± 2.08 ± 61.06 ± Exam- 0.35&& 0.24 0.14& 3.2**& ple 4 5 6.87 ± 3.4 ± 2.2 ± 60.15 ± 0.39&&& 0.23 0.168z 2.14**& 10 6.74 ± 3.49 ± 2.87 ± 63.82 ± 0.41&& 0.28& 0.11#&&& 2.55*#&& OVX + 2.5 6.48 ± 3.58 ± 2.58 ± 65.15 ± Exam- 4.47&& 2.41& 1.91&& 3.23&& ple 5 5 6.34 ± 3.8 ± 2.18 ± 73.15 ± 2.03& 1.26&& 1.02& 4.78#&& 10 6.8 ± 3.4 ± 2.57 ± 66.54 ± 1.38&& 1.02 0.31&& 2.38#&& Note: WBC: white blood cell, Lymph: lymphocyte, Gran: neutrophile granulocyte; compared with Sham group, *represented for P <0.05, **represented for P <0.01, and ***represented for P<0.001; compared with OVX group, #represented for P <0.05, ##represented for P <0.01, ###represented for P <0.001; and compared with Aba-5 μ/kg group, &represented for P <0.05, &&represented for P <0.01, and &&&represented for P <0.001.

[0227] Results and discussion: on the basis of Table 2, it could be concluded that compared with Sham group, the number of white blood cells, lymphocytes and neutrophile granulocytes of osteoporosis rat in OVX group did not significantly change, indicating that karyocyte level in peripheral blood of osteoporosis rat kept normal.

[0228] In abaloptide control group, after administering for 25 weeks, the number of white blood cell significantly decreased compared with OVX osteoporosis model control group and Sham control group, and the number of lymphocytes significantly decreased compared with Sham control group, and the number of neutrophile granulocyte significantly decreased compared with OVX group.

[0229] Compared with OVX model group and Sham control group, after administering the test compounds in examples 1-5 at dosages of 2.5 μg/kg, 5 μg/kg and 10 μg/kg for 25 weeks, the number of white blood cells, lymphocytes and neutrophile granulocyte kept normal, indicating that the compound of the present disclosure stabilized the number of karyocyte in peripheral blood while facilitating ossification and improving bone mineral density of osteoporosis rat.

[0230] Compared with 5 μg/kg administration group of abaloptide, after administering the test compounds in examples 1-5 at dosages of 2.5 μg/kg, 5 μg/kg and 10 μg/kg for 25 weeks, the number of white blood cells, lymphocytes and neutrophile granulocyte in peripheral blood was significantly higher than that of abaloptide group. During the administration period of abaloptide, mononuclear cell, lymphocyte and white blood cells in peripheral blood significantly decreased, while the compounds in the present disclosure significantly stablized the karyocyte level in peripheral blood cells, and overcame the adverse effects of abaloptide.

(II) Research on Retinoic Acid-Induced Osteoporosis Therapeutic Effect of the Compound in the Present Disclosure

[0231] Experimental method: SD rats were administered by gavage to induce a rat osteoporosis model. After model establishment, the rats were randomly grouped according to bone mineral densities of thigh bone: {circle around (1)} vehicle group (Control group): normal saline was subcutaneously administered in an equal volume to the test compound, and soybean oil was administered by gavage in an equal volume to retinoic acid; {circle around (2)} Model group: normal saline of the same volume was subcutaneously administered; and administered with retinoic acid (RA) by gavage every other day to maintain osteoporosis state; {circle around (3)} 20 μg dosage group of positive drug abaloptide (Aba-20): 10 μg/kg abaloptide was subcutaneously administered, and retinoic acid was administered by gavage every other day. Administration groups of three dosages were set for each test compound. −10 μg/kg dosage group of the test compound: 10 μg/kg test compound was subcutaneously administered, and retinoic acid was administered by gavage every other day. −20 μg/kg dosage group of the test compound: 20 μg/kg test compound was subcutaneously administered, and retinoic acid was administered by gavage every other day. −40 μg/kg dosage group of the test compound: 40 μg/kg test compound was subcutaneously administered, and retinoic acid was administered by gavage every other day. The administration was carried out 5 times a week, with normal saline as the diluent of test drug, and the administration successively lasted for 12 weeks. The induction dosage of retinoic acid was about 80 mg/kg and the maintenance dosage after model establishment was about 30 mg/Kg, and solvent of inducing agent was soybean oil.

[0232] Times and contents of detection indicators were shown hereinafter.

[0233] 1) Bone mineral density (BMD) was detected with a dual-energy X-ray bone densitometry (DXA) before administration (0 w), and 12 w after administration.

[0234] 2) Blood was collected from eye orbit before administration (0 w) and 12 w after administration, the serum was removed to carry out a detection of calcium, phosphorus and alkaline phosphatase (ALP); and the other part of whole blood was subjected to a blood routine examination.

[0235] 3) 12 w after administration, the serum of blood collected from eye orbit was subjected to detection of bone turnover markers (CTx (β-CTX, serum), OC (serum) and serum procollagen type 1 N-terminal propeptide (PINP)).

[0236] After administration completed, tibia on the left was used to prepare a bone marrow smear. The thigh bone on the left was subjected to three-point bending test, and the thigh bone on the right was subjected to CT scan and was used to prepare a pathological section.

[0237] Statistical analysis of data: the data were treated with SPSS 20 software. The data in line with normality test were subjected to one-way analysis of variance; and the data not in line with normality test were subjected to rank sum test. The data were represented by mean value±standard error.

Experimental Results

[0238] (1) Influence of the Compound on Bone Marrow Karyocyte of Retinoic Acid-Induced Osteoporosis Rats

[0239] The number of karyocyte in bone marrow cavity of retinoic acid-induced osteoporosis rats was counted, shown in Table 3.

TABLE-US-00016 TABLE 3 number of bone marrow karyocyte 12 weeks after administration Dosage Number of cells Group (μg/kg) (*10.sup.5) Control — 235.35 ± 26.08 Model/RA — 190.44 ± 29.83 RA + Aba 20 112.21 ± 13.85***# RA + 10 187.66 ± 15.78&& Example 1 20 195.18 ± 15.22& 40 189.5 ± 27.29& RA + 10 210.43 ± 23.99&& Example 2 20 205.81 ± 14.39& 40 187.5 ± 27.29& RA + 10 186.57 ± 16.89& Example 3 20 201.82 ± 12.93& 40 193.15 ± 20.31& RA + 10 198.21 ± 13.76& Example 4 20 201.82 ± 12.93& 40 203.45 ± 27.29& RA + 10 206.11 ± 20.32&& Example 5 20 200.33 ± 9.93& 40 210.00 ± 26.14&& Note: Aba: positive drug abaloptide; RA: retinoic acid; compared with Control group, ***represented for p <0.001; compared with RA group, #represented for p <0.05; compared with Aba-20 μg/kg group, && represented for p <0.01; and compared with Aba-20 μg/kg group, & represented for p <0.05.

[0240] Results and discussion: after administering for 12 w, compared with Control group, the number of karyocyte in bone marrow cavity significantly decreased in 20 μg/kg group of abaloptide (P<0.001); compared with model group, the number of karyocyte in bone marrow cavity significantly decreased in 20 μg/kg group of abaloptide (P<0.05). Retinoic acid-induced osteoporosis rat did not have bone marrow suppression, and abaloptide had obvious inhibiting effects on bone marrow of normal rats and osteoporosis rats.

[0241] Compared with 20 μg/kg dosage group of abaloptide, the numbers of karyocyte in 10 and 20 μg/kg dosage groups of test compounds 1-5 significantly increased (P<0.05). The numbers of bone marrow karyocyte in administration groups of test compounds 1-5 were close to normal level (there was no significant difference compared with control group). The compounds in the present disclosure stablized the number of bone marrow karyocyte.

[0242] (2) Influence of the Compound on Microstructure of Bone of Retinoic Acid-Induced Osteoporosis Rats

[0243] After the experiment was completed, thigh bone on the right side of rats were collected, and subjected to CT detection, and bone surface area/bone volume ratio (BV/TV), the number of trabeculae (TbN) and trabeculae spacing (TbSp) were calculated, and the results were shown in Table 4. An example was selected from each of Control group, Model group, abaloptide group, 20 μg/kg dosage group of Example 1, 20 μg/kg dosage group of Example 2 and 20 μg/kg dosage group of Example 5, and the scanned figures by micron X-ray 3D imaging system of trochlea of their thigh bone were shown in FIG. 1.

TABLE-US-00017 TABLE 4 Effect of the compound on the bone microstructure of rats with retinoic acid-induced osteoporosis Indicators of CT Dosage TbN TbSp Group μg/kg BV/TV (mm.sup.−1) (mm) Control — 0.32 ± 5.22 ± 0.13 ± 0.02 0.19 0.01 Model/RA — 0.15 ± 2.9 ± 0.32 ± 0.02.sup.** 0.43.sup.*** 0.05.sup.*** RA + Aba 20 0.28 ± 3.4 ± 0.22 ± 0.02 0.2.sup.*** 0.02 RA + 10 0.34 ± 4.24 ± 0.17 ± Example 1 0.03.sup.### 0.16.sup.# 0.02.sup.## 20 0.32 ± 4.01 ± 0.19 ± 0.01.sup.## 0.34 0.03.sup.## 40 0.41 ± 4.25 ± 0.14 ± 0.02.sup.### 0.15.sup.# 0.01.sup.### RA + 10 0.35 ± 4.10 ± 0.17 ± Example 2 0.04.sup.### 0.28 0.03.sup.## 20 0.32 ± 4.06 ± 0.18 ± 0.03.sup.## 0.36 0.04.sup.## 40 0.40 ± 4.38 ± 0.15 ± 0.04.sup.### 0.17.sup.# 0.04.sup.## RA + 10 0.33 ± 4.17 ± 0.20 ± Example 3 0.01.sup.### 0.28.sup.# 0.04.sup.# 20 0.30 ± 4.21 ± 0.17 ± 0.02.sup.## 0.16.sup.# 0.04.sup.## 40 0.39 ± 4.22 ± 0.15 ± 0.05.sup.## 0.37.sup.# 0.02.sup.## RA + 10 0.35 ± 4.23 ± 0.17 ± Example 4 0.01.sup.### 0.21 0.03.sup.## 20 0.37 ± 4.56 ± 0.18 ± 0.02.sup.## 0.33.sup.# 0.04.sup.## 40 0.40 ± 4.58 ± 0.15 ± 0.03.sup.### 0.15.sup.# 0.04.sup.## RA + 10 0.33 ± 4.19 ± 0.16 ± Example 5 0.03.sup.### 0.28.sup.# 0.03.sup.## 20 0.36 ± 4.42 ± 0.15 ± 0.02.sup.## 0.16.sup.# 0.06.sup.## 40 0.39 ± 4.48 ± 0. 14 ± 0.02.sup.### 0.09.sup.# 0.03.sup.## Note: compared with Control group, *represented for p <0.05, **represented for p <0.01, ***represented for p <0.001; compared with RA group, #represented for p <0.05, ##represented for p <0.01, and ###represented for p <0.001; compared with Aba-20 μg/kg group, & represented for p <0.05; and n = 6-8.

[0244] Results and discussion: it can be concluded from Table 4 that, after administering for 12 w, compared with Control group, the bone volume/total volume BV/TV (P<0.01) and the number of trabeculae TbN (P<0.001) of RA group significantly decreased, and the trabeculae spacing TbSp (P<0.001) significantly increased, indicating that microstructure of bone tissue of rats in Model (RA) group was seriously damaged.

[0245] Compared with model group/RA, the bone volume/total volume BV/TV and the number of trabeculae TbN of each dosage group of the test compounds 1-5 significantly increased, and the trabeculae spacing TbSp (P<0.001) significantly decreased, indicating that bone mass of thigh bone of rats increased and microstructure damage of bone tissue was repaired after administering the test compounds of the present disclosure.

[0246] Under the same dosage (20 μg/kg), the test compounds 1-5 were obviously superior to abaloptide in aspects of improving bone surface area/total volume ratio (BV/TV), number of trabeculae (TbN) and trabeculae spacing (TbSp), showing that the compounds of the present disclosure were superior to abaloptide in treating high-turnover osteoporosis.

[0247] (3) Influence of the Compound on Biomechanics of Retinoic Acid-Induced Osteoporosis Rats

[0248] After the experiment, thigh bones on the left of rats were collected, subjected to three-point mechanical test, and the results of experiments were shown in Table 5.

TABLE-US-00018 TABLE 5 Influence of the compound on three-point mechanical test of osteoporosis rats Peak load of Dosage three-point Group (μg/kg) mechanics (N) Control — 144.08 ± 7.87 RA(Model) — 88.05 ± 7.1*** RA + Aba 20 119.57 ± 6.32# RA + 10 98.11 ± Example 1 5.43** 20 123.54 ± 3.95# 40 130.38 ± 8.54## RA + 10 97.87 ± Example 2 8.71** 20 122.02 ± 6.65# 40 133.38 ± 9.70## RA + 10 97.65 ± Example 3 8.53** 20 126.54 ± 6.76# 40 131.74 ± 5.89## RA + 10 99.01 ± Example 4 9.12** 20 125.43 ± 7.44# 40 136.96 ± 8.52## RA + 10 98.47 ± Example 5 8.84** 20 127.02 ± 8.63# 40 132.38 ± 7.32## Note: Aba: positive drug abaloptide; RA: inducing agent retinoic acid; compared with Control group, **represented for p <0.01, and ***represented for p <0.001; compared with RA group, #represented for p <0.05, and ##represented for p <0.01; and n = 5-14.

[0249] Results and discussion: after administering for 12 w, thigh bone three-point mechanical test was carried out. Compared with Control group, the peak load of Model group significantly decreased (P<0.001), indicating that the peak load of thigh bone of retinoic acide-induced osteoporosis rats significantly decreased.

[0250] Compared with Model group, peak loads of thigh bone of rats in the 20 μg/kg, 40 μg/kg dosage groups of test compound significantly increased (P<0.05). Peak load of 20 μg/kg dosage group of abaloptide did not significantly increase compared with Model group. In the aspect of improving peak load of thigh bone of osteoporosis rat, the compounds of the present disclosure were superior to abaloptide.

[0251] In summary, compared with abaloptide, the compounds in the present disclosure can significantly stablize karyocyte level in both bone marrow and peripheral blood. The compounds in the present disclosure can increase the peak load of retinoic acid-induced osteoporosis rat, and the effect is better than that of abaloptide. The compounds of the present disclosure can also improve microstructure of bone, specifically including improving bone surface area/total volume ratio (BV/TV), trabeculae number (TbN) and trabecular spacing, and the effect is better than the marketed drug abaloptide.

[0252] The present disclosure also relates to the following embodiments:

[0253] 1. An active polypeptide compound, which has a structure represented by following Formula (Ia) or Formula (Ib), or is a pharmaceutically acceptable salt thereof,

Y-ID-X Formula (Ia), or

X-ID-Y Formula (Ib),

[0254] wherein,

[0255] Y is a PTH/PTHrP receptor agonist or an osteoclast inhibitor;

[0256] ID is a peptide bond or a linker in the molecule, which links X to Y; and

[0257] X is an osteogenic growth peptide receptor agonist, a bone marrow mesenchymal stem cell irritant or a hematopoietic stem cell irritant.

[0258] 2. The active polypeptide compound according to embodiment 1, wherein Y is M-CSF antagonist, RANKL inhibitor, RANKL antibody, MMP inhibitor, calcitonin, parathyroid hormone or parathyroid hormone-related protein.

[0259] 3. The active polypeptide compound according to embodiment 1 or 2, wherein Y is a peptide chain having an amino acid sequence as shown in Formula (II):

A.sub.1-Val-Ser-Glu-His-Gln-Leu-A.sub.8-His-Asp-Lys-Gly-Lys-Ser-Ile-Gln-A.sub.17-Leu-Arg-Arg-Arg-A.sub.22-A.sub.23-Leu-A.sub.25-A.sub.26-Leu-A.sub.28-A.sub.29-A.sub.30-A.sub.31-His-Thr-Ala Formula (II);

[0260] wherein, A.sub.1 is Ala, Val, Leu or Ile;

[0261] A.sub.8 is Leu or Ile;

[0262] A.sub.17 is Asp or Glu;

[0263] A.sub.22 is Glu, Asp or Phe;

[0264] A.sub.23 is Leu, Ile or Phe;

[0265] A.sub.25 is Glu, Asp or His;

[0266] A.sub.26 is Lys, His or Arg;

[0267] A.sub.28 is Leu, Ile or Val;

[0268] A.sub.29 is Ala, (N-Me)Ala or Aib;

[0269] A.sub.30 is Lys or Glu;

[0270] A.sub.31 is Leu or Ile;

[0271] the amino terminal of the peptide chain Y is free or chemically modified, and the carboxyl terminal of the peptide chain Y is free or chemically modified.

[0272] 4. The active polypeptide compound according to any one of embodiments 1-3, wherein X is a hematopoietic stem cell irritant, a hematopoietic growth factor, a platelet colony-stimulating factor, a granulocyte colony-stimulating factor, erythropoietin, interleukin 3 or recombinant human interleukin 11.

[0273] 5. The active polypeptide compound according to any one of embodiments 1-4, wherein X is a peptide chain having an amino acid sequence as shown in Formula (IIIa) or Formula (IIIb):

Tyr-(Arg)m-(Gly).sub.n-Phe-Gly-Gly Formula (IIIa)

Gly-Gly-Phe-(Gly).sub.n-(Arg).sub.m-Tyr Formula (IIIb);

[0274] wherein, m and n are independently 0, 1 or 2; and

[0275] the amino terminal of the peptide chain X is free or chemically modified, and the carboxyl terminal of the peptide chain X is free or chemically modified.

[0276] 6. The active polypeptide compound according to embodiment 5, wherein X is a peptide chain consisting of 5-6 amino acids which has an amino acid sequence as shown in one of the following SEQ ID NO:1-SEQ ID NO:8:

TABLE-US-00019 (SEQ ID NO: 1) Tyr-Gly-Phe-Gly-Gly (SEQ ID NO: 2) Tyr-Arg-Phe-Gly-Gly (SEQ ID NO: 3) Tyr-Arg-Gly-Phe-Gly-Gly (SEQ ID NO: 4) Tyr-Pro-Phe-Gly-Gly (SEQ ID NO: 5) Gly-Gly-Phe-Gly-Tyr (SEQ ID NO: 6) Gly-Gly-Phe-Arg-Tyr (SEQ ID NO: 7) Gly-Gly-Phe-Gly-Arg-Tyr (SEQ ID NO: 8) Gly-Gly-Phe-Pro-Tyr.

[0277] 7. The active polypeptide compound according to any one of embodiments 1-6, wherein ID is a linker between X and Y; the linker is an amino-substituted C.sub.1-8 alkyl carboxylic acid, a polyethylene glycol polymer chain or a peptide segment consisting of 1-10 amino acids, and the amino acids in the peptide segment is selected from the group consisting of proline, arginine, alanine, threonine, glutamic acid, aspartic acid, lysine, glutamine, asparagine and glycine.

[0278] 8. The active polypeptide compound according to embodiment 7, wherein the linker is one of the following linkers:

[0279] (1) (Gly-Ser).sub.p, wherein p is 1, 2, 3, 4 or 5;

[0280] (2) (Gly-Gly-Gly-Gly-Ser).sub.t, wherein t is 1, 2 or 3;

[0281] (3) Ala-Glu-Ala-Ala-Ala-Lys-Ala;

[0282] (4) 4-aminobutyric acid or 6-aminocaproic acid; and

[0283] (5) (PEG).sub.q, wherein q is 1, 2, 3, 4 or 5.

[0284] 9. The active polypeptide compound according to any one of embodiments 1-8, which has a structure as shown in Formula (IV), or is a pharmaceutically acceptable salt of the compound shown as in Formula (IV):

[0285] wherein, A.sub.1 is Ala, Val, Leu or Ile;

[0286] A.sub.8 is Leu or Ile;

[0287] A.sub.17 is Asp or Glu;

[0288] A.sub.22 is Glu, Asp or Phe;

[0289] A.sub.23 is Leu, Ile or Phe;

[0290] A.sub.25 is Glu, Asp or His;

[0291] A.sub.26 is Lys, His or Arg;

[0292] A.sub.28 is Leu, Ile or Val;

[0293] A.sub.29 is Ala, (N-Me)Ala or Aib;

[0294] A.sub.30 is Lys or Glu;

[0295] A.sub.31 is Leu or Ile; and

[0296] A.sub.35 has a peptide chain of the amino acid sequence as shown in Formula (IIIa) or (IIIb):

Tyr-(Arg).sub.m-(Gly).sub.n-Phe-Gly-Gly Formula (IIIa),

Gly-Gly-Phe-(Gly).sub.n-(Arg).sub.m-Tyr Formula (IIIb),

[0297] wherein, m and n are independently 0, 1 or 2; and

[0298] the amino terminal of the amino acids shown by A.sub.1 is free or chemically modified, and the carboxyl terminal of the peptide chain A.sub.35 is free or chemically modified.

[0299] 10. The active polypeptide compound according to embodiment 9, wherein the chemical modifications of the amino terminal include acylation, sulfonylation, alkylation and PEG modification; and the chemical modifications of the carboxyl terminal include amidation, sulfonylation and PEG modification.

[0300] 11. The active polypeptide compound according to embodiment 10, wherein the chemical modification of amino terminal is acetylation, benzoylation or sulfonylation of amino; the alkylation of amino terminal is C.sub.1-6 alkylation or aromatic alkylation; the chemical modification of carboxylic terminal is that the OH in the carboxyl is substituted by NH.sub.2 or sulfamide, or the OH in the carboxyl links to the functionalized PEG molecule.

[0301] 12. The active polypeptide compound according to any one of embodiments 1-11, which is one of the compounds in the following SEQ ID NO:9-SEQ ID NO:15, or a pharmaceutically acceptable salt thereof:

TABLE-US-00020 (1) SEQ ID NO: 9 Ala-Val-Ser-Glu-His-Gln-Leu-Leu-His-Asp-Lys-Gly- Lys-Ser-Ile-Gln-Asp-Leu-Arg-Arg-Arg-Glu-Leu-Leu- Glu-Lys-Leu-Leu-(N-Me)Ala-Lys-Leu-His-Thr-Ala- Tyr-Gly-Phe-Gly-Gly; (2) SEQ ID NO: 10 Ala-Val-Ser-Glu-His-Gln-Leu-Leu-His-Asp-Lys-Gly- Lys-Ser-Ile-Gln-Asp-Leu-Arg-Arg-Arg-Glu-Leu-Leu- Glu-Lys-Leu-Leu-Aib-Lys-Leu-His-Thr-Ala-Tyr-Gly- Phe-Gly-Gly; (3) SEQ ID NO: 11 Ala-Val-Ser-Glu-His-Gln-Leu-Leu-His-Asp-Lys-Gly- Lys-Ser-Ile-Gln-Asp-Leu-Arg-Arg-Arg-Glu-Leu-Leu- Glu-Lys-Leu-Leu-Ala-Lys-Leu-His-Thr-Ala-Tyr-Arg- Gly-Phe-Gly-Gly; (4) SEQ ID NO: 12 Ala-Val-Ser-Glu-His-Gln-Leu-Leu-His-Asp-Lys-Gly- Lys-Ser-Ile-Gln-Asp-Leu-Arg-Arg-Arg-Phe-Phe-Leu- His-His-Leu-Ile-Ala-Glu-Ile-His-Thr-Ala-Tyr-Gly- Phe-Gly-Gly; (5) SEQ ID NO: 13 Ala-Val-Ser-Glu-His-Gln-Leu-Leu-His-Asp-Lys-Gly- Lys-Ser-Ile-Gln-Asp-Leu-Arg-Arg-Arg-Phe-Phe-Leu- His-His-Leu-Ile-Aib-Glu-Ile-His-Thr-Ala-Tyr-Arg- Phe-Gly-Gly; (6) SEQ ID NO: 14 Ala-Val-Ser-Glu-His-Gln-Leu-Ile-His-Asp-Lys-Gly- Lys-Ser-Ile-Gln-Glu-Leu-Arg-Arg-Arg-Phe-Phe-Leu- His-His-Leu-Ile-Aib-Glu-Ile-His-Thr-Ala-Tyr-Gly- Phe-Gly-Gly; (7) SEQ ID NO: 15 Ala-Val-Ser-Glu-His-Gln-Leu-Ile-His-Asp-Lys-Gly- Lys-Ser-Ile-Gln-Glu-Leu-Arg-Arg-Arg-Phe-Phe-Leu- His-His-Leu-Leu-Ala-Glu-Ile-His-Thr-Ala-Tyr-Gly- Phe-Gly-Gly.

[0302] 13. The active polypeptide compound according to claim 1, further includes a compound obtained by chemically modifying the side chain groups of amino acids of the polypeptide compound; or

[0303] a coordination compound, a complex or a chelate formed by the polypeptide compound and a metal ion; or

[0304] a hydrate or a solvate formed by the polypeptide compound.

[0305] 14. The active polypeptide compound according to embodiment 13, wherein the compound obtained by chemically modifying the side chain groups of amino acids of the polypeptide compound is a thioether or thioglycoside formed from a sulfydryl in a cysteine in the polypeptide compound, or a compound having a disulfide bond formed from a cysteine or a peptide comprising cysteine; or

[0306] an ester, an ether and a glycoside formed from a phenolic hydroxyl group of a tyrosine in the polypeptide compound; or

[0307] a compound prepared by substituting a benzene ring of a tyrosine or phenylalanine in the polypeptide compounds.

[0308] 15. A pharmaceutical composition, comprising the active polypeptide compound according to any one of embodiments 1-14, and at least one of a pharmaceutically acceptable adjuvant, excipient, carrier and solvent thereof.

[0309] 16. The pharmaceutical composition according to embodiment 15, comprising other therapeutic agents, which are selected from a drug that inhibits bone resorption, a drug that promotes ossification, a drug that promotes bone mineralization and a parathyroid hormone-related protein.

[0310] 17. The pharmaceutical composition according to embodiment 16, wherein the drug that inhibits bone resorption includes calcitonin, diphosphonate, oestrogen, a selective oestrogen receptor regulator and isoflavone; the drug that promotes ossification includes fluoride, synthesized steroid, parathyroid hormone and parathyroid hormone-related protein; the drug that promotes bone mineralization includes a calcium agent, vitamin D and active vitamin D; and the parathyroid hormone-related protein is teriparatide or abaloptide.

[0311] 18. Use of the compound according to any one of embodiments 1-4 and the pharmaceutical composition according to any one of embodiments 15-18 in the preparation of a medicament for preventing, treating or alleviating diseases or disorders related to osteogenic defects or bone mineral density decreasing, and the diseases include osteoporosis.

[0312] The above contents are further detailed descriptions of the present disclosure in combination with specific preferred embodiments, but it cannot be considered that the specific implementations of the present disclosure are limited to these descriptions. For one of ordinary skill in the art to which the present disclosure pertains, without deviating from the concept of the present disclosure, several simple deductions or replacements can also be made, which should all be regarded as belonging to the protection scope of the present disclosure.

ACTIVE POLYPEPTIDE COMPOUND

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Abstract

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