CN114478709A

CN114478709A - Long-acting hepatitis virus entry inhibitor

Info

Publication number: CN114478709A
Application number: CN202011270451.9A
Authority: CN
Inventors: 周述靓; 王鹏; 邓岚; 潘文强
Original assignee: Chengdu Aoda Biotechnology Co ltd
Current assignee: Chengdu Aoda Biotechnology Co ltd
Priority date: 2020-11-13
Filing date: 2020-11-13
Publication date: 2022-05-13

Abstract

The invention relates to the field of medicine synthesis, and discloses a long-acting hepatitis virus entry inhibitor. The long-acting hepatitis virus entry inhibitor is used for preparing a pharmaceutical composition for treating diseases, and the pharmaceutical composition is used for treating chronic hepatitis B and hepatitis D. Meanwhile, the invention prolongs the half-life period of the medicament, can provide the long-acting hepatitis virus entry inhibitor and the medicament for patients, reduces the administration frequency and lowers the cost of the patients.

Description

Long-acting hepatitis virus entry inhibitor

Technical Field

The invention relates to the field of medicine synthesis, and particularly relates to a long-acting hepatitis virus entry inhibitor and application thereof.

Background

Viral hepatitis is an infectious disease mainly caused by liver diseases caused by various hepatitis viruses. Clinically, the symptoms of anorexia, nausea, epigastric discomfort, liver pain and hypodynamia are mainly manifested. Some patients may have jaundice fever and hepatomegaly with liver function impairment. Some patients can become chronic, even develop cirrhosis of the liver, and a few can develop liver cancer.

The etiological typing of viral hepatitis is currently recognized by five hepatitis viruses, namely hepatitis A virus, hepatitis B virus, hepatitis C virus, hepatitis D virus and hepatitis E virus, which are respectively written as HAV, HBV, HCV, HDV and HEV, and the rest are RNA viruses except the hepatitis B virus which is a DNA virus.

Hepatitis A Virus (HAV) is a hepadnavirus genus of the picornaviridae family. After human infection with HAV, most of them show subclinical or recessive infection, and only a few of them show acute hepatitis A. Generally, the chronic hepatitis can be completely recovered without conversion to chronic hepatitis or chronic carriers.

Hepatitis B Virus (HBV) is the causative agent of hepatitis B and belongs to the family hepadnaviridae, which contains two genera, orthophagia and avian hepadnaviridae, and it is the genus orthophagia that causes infection in humans. HBV infection is a global public health problem, along with the production and investment of genetic engineering vaccines, the popularization rate of hepatitis B vaccines is increased year by year, and the infection rate is in a descending trend. There are approximately 20 million HBV infected patients worldwide, of which 3.8 million infected patients are chronic infections and the virus is hidden in tissues and organs and cannot be completely eliminated by the immune system and drugs.

Hepatitis C Virus (HCV), is transmitted vertically mainly through blood, sexual life, mother and infant. Unfortunately, the medical community has not yet developed a vaccine effective in preventing hepatitis c. Because hepatitis C virus is RNA virus, it is very easy to mutate, and the difficulty of developing vaccine is very large, because except human and chimpanzee, other animals will not suffer from hepatitis C, so it is difficult to find animal model for vaccine development, but new generation oral direct antiviral medicine can cure hepatitis C.

HDV is a defective single-stranded negative-strand RNA virus that must rely on hepadnaviruses such as HBV to provide an outer shell for replication. HDV is present in the nuclei and in the serum of HBsAg-positive HDV-infected individuals. Replication occurs primarily in hepatocytes. HDV is prone to mutation. The human body can obviously inhibit the synthesis of HBV-DNA after being infected with HDV, the occurrence of HDAg is consistent with the reduction of HBV-DNA in serum, and the HBV-DNA is restored to the original level along with the negative change of HDAg and the occurrence of anti-HD. Mainly transmitted by blood transfusion and blood products, and is similar to the transmission mode of hepatitis B, HDV infection is mostly seen in HBV infected persons, and sporadic HDV infected persons can also be seen. HDV can promote exacerbation of liver damage following superinfection with HBV and is prone to develop chronic active hepatitis, cirrhosis and severe hepatitis.

Researchers have found that a protein called liver bile acid transporter (NTCP, sodium taurocholate cotransporter polypeptide) on the surface of hepatocytes can specifically interact with the key receptor binding domain of HBV envelope proteins, which are receptors required for HBV infection of host cells. Therefore, blocking NTCP would be expected to cure hepatitis B.

Bulevirtide is a drug which contains 47 amino acids and targets NTCP, and has short half-life in vivo, so that patients need to take the Bulevirtide daily and for a long time, the patient compliance is poor, and the clinical cost is high. The invention aims to provide a long-acting hepatitis virus entry inhibitor for patients, reduce the administration frequency and reduce the cost of the patients.

Disclosure of Invention

In view of the above, the present invention provides a long-acting hepatitis virus entry inhibitor and its use.

This is done.

In order to achieve the above object, the present invention provides the following technical solutions:

the invention provides a polypeptide, the structure of which is shown as formula I:

AA1-Gly-Thr-Asn-Leu-Ser-Val-Pro-Asn-Pro-Leu-Gly-Phe-Phe-Pro-

Asp-His-Gln-Leu-Asp-Pro-Ala-Phe-Gly-Ala-Asn-Ser-Asn-Asn-Pro-

Asp-Trp-Asp-Phe-Asn-Pro-Asn-Lys-Asp-His-Trp-Pro-Glu-Ala-Asn-

AA2-Val-Gly-AA3-AA4(R1)-AA5(R2)-AA6

formula I

AA1 in formula I is selected from Ac or H; and/or

AA2 in formula I is selected from Lys, Gln, Glu, Arg or Cit; and/or

AA3 in formula I is selected from PEG_m1(CH₂)_m2CO-, or is absent;

wherein: m1 is an integer from 1 to 10;

m2 is an integer from 1 to 5; and/or

AA5 is selected from Cys when AA4 is absent in formula I; and/or

AA4 is selected from Lys, Dah, Orn, Dab or Dap when AA5 in formula I is absent; and/or

AA6 in formula I is selected from NH₂Or OH; and/or

R1 in formula I is selected from cholesterol succinate monoester, 2-cholesterol acetic acid, 2-cholesterol propionic acid, 3-cholesterol propionic acid, 2-cholesterol butyric acid, 2-cholesterol isobutyric acid, 3-cholesterol butyric acid, 3-cholesterol isobutyric acid, 4-cholesterol butyric acid, 2-cholesterol valeric acid, 2-cholesterol isovaleric acid, 3-cholesterol valeric acid, 5-cholesterol valeric acid, 2-cholesterol caproic acid, 6-cholesterol caproic acid, 2-cholesterol enanthic acid, 7-cholesterol enanthic acid, 2-cholesterol caprylic acid, 8-cholesterol caprylic acid, HO2C (CH2) n1CO- (gamma Glu) n2- (PEGn3(CH 2)₂)n4CO)n5-；

Wherein: n1 is an integer from 10 to 20;

n2 is an integer from 1 to 5;

n3 is an integer from 1 to 30;

n4 is an integer from 1 to 5;

n5 is an integer from 1 to 5; and/or

R2 in formula I is selected from 2-cholesteryl acetate, 2-cholesteryl propionate, 3-cholesteryl propionate, 2-cholesteryl butyrate, 2-cholesteryl isobutyrate, 3-cholesteryl butyrate, 3-cholesteryl isobutyrate, 4-cholesteryl butyrate, 2-cholesteryl valerate, 2-cholesteryl isovalerate, 3-cholesteryl valerate, 5-cholesteryl valerate, 2-cholesteryl hexanoate, 6-cholesteryl hexanoate, 2-cholesteryl heptanoate, 7-cholesteryl heptanoate, 2-cholesteryl octanoate or 8-cholesteryl octanoate.

Based on the research, the invention also provides application of the polypeptide or the derivative thereof in preparing a long-acting hepatitis virus entry inhibitor.

In some embodiments of the invention, the derivative comprises a pharmaceutically acceptable salt, solvate, chelate or non-covalent complex of the polypeptide, based on a mixture of one or more of the prodrugs on which the compound is based.

The invention also provides the application of the polypeptide or the derivative thereof in preparing a medicament for treating liver diseases.

In some embodiments of the invention, the liver disease comprises hepatitis.

In some embodiments of the invention, the hepatitis comprises chronic hepatitis B and/or hepatitis D.

The invention also provides a long-acting hepatitis virus entry inhibitor which comprises the polypeptide or the derivative thereof and pharmaceutically acceptable auxiliary materials.

In addition, the invention also provides a medicament for treating hepatitis, which comprises the polypeptide or the derivative thereof and pharmaceutically acceptable auxiliary materials. In some embodiments of the invention, the hepatitis comprises chronic hepatitis b and/or hepatitis d.

Further details of the invention are set forth below, or some may be appreciated in embodiments of the invention.

Unless otherwise indicated, the amounts of the various ingredients, reaction conditions, and the like used herein are to be construed in any case to mean "about". Accordingly, unless expressly stated otherwise, all numerical parameters set forth in the following specification and attached claims are approximations that may vary depending upon the standard deviation found in the respective experimental conditions.

Herein, when a chemical structural formula and a chemical name of a compound are ambiguous or ambiguous, the compound is exactly defined by the chemical structural formula. The compounds described herein may contain one or more chiral centers, and/or double bonds and the like, and stereoisomers, including isomers of double bonds (e.g., geometric isomers), optical enantiomers, or diastereomers, may also be present. Accordingly, any chemical structure within the scope of the description, whether partial or complete, including similar structures as described above, includes all possible enantiomers and diastereomers of the compound, including any stereoisomer alone (e.g., pure geometric isomers, pure enantiomers, or pure diastereomers), as well as any mixture of such stereoisomers. Mixtures of these racemates and stereoisomers may also be further resolved into the enantiomers or stereoisomers of their constituent members by those skilled in the art using non-stop separation techniques or methods of chiral molecular synthesis.

The compounds of formula I include, but are not limited to, optical isomers, racemates and/or other mixtures of these compounds. In the above case, a single enantiomer or diastereomer, such as an optical isomer, can be obtained by asymmetric synthesis or racemate resolution. Resolution of the racemates can be accomplished by various methods, such as conventional recrystallization from resolution-assisting reagents, or by chromatographic methods. In addition, the compounds of the formula I also comprise the cis and/or trans isomers with double bonds.

The compounds of the present invention include, but are not limited to, the compounds of formula I and all of their pharmaceutically acceptable different forms. The pharmaceutically acceptable different forms of these compounds include various pharmaceutically acceptable salts, solvates, complexes, chelates, non-covalent complexes, prodrugs based on the above and any mixtures of these forms.

In summary, the present invention provides a long-acting hepatitis virus entry inhibitor. The long-acting hepatitis virus entry inhibitor is used for preparing a pharmaceutical composition for treating liver diseases, and the pharmaceutical composition can be used for treating chronic hepatitis B and hepatitis D. Meanwhile, the invention prolongs the half-life period of the medicament, can provide the long-acting hepatitis virus entry inhibitor and the medicament for patients, reduces the administration frequency and lowers the cost of the patients.

Detailed Description

The invention discloses a long-acting hepatitis virus entry inhibitor and application thereof, and a person skilled in the art can realize the long-acting hepatitis virus entry inhibitor by properly improving process parameters by referring to the content. It is expressly intended that all such similar substitutes and modifications which would be obvious to one skilled in the art are deemed to be included in the invention. While the methods and applications of this invention have been described in terms of preferred embodiments, it will be apparent to those of ordinary skill in the art that variations and modifications in the methods and applications described herein, as well as other suitable variations and combinations, may be made to implement and use the techniques of this invention without departing from the spirit and scope of the invention.

The invention firstly provides a compound shown in the structure I, a pharmaceutically acceptable salt, a solvate, a chelate or a non-covalent compound formed by the compound, a prodrug based on the compound, or any mixture of the forms.

AA1-Gly-Thr-Asn-Leu-Ser-Val-Pro-Asn-Pro-Leu-Gly-Phe-Phe-Pro-

Asp-His-Gln-Leu-Asp-Pro-Ala-Phe-Gly-Ala-Asn-Ser-Asn-Asn-Pro-

Asp-Trp-Asp-Phe-Asn-Pro-Asn-Lys-Asp-His-Trp-Pro-Glu-Ala-Asn-

AA2-Val-Gly-AA3-AA4(R1)-AA5(R2)-AA6

Structure I

AA1 in structure I is Ac, or is H;

AA2 in structure I is Lys, or Gln, or Glu, or Arg, or Cit;

AA3 in Structure I is PEG_m1(CH₂)_m2CO-, or is absent;

wherein: m1 is an integer from 1 to 10;

m2 is an integer from 1 to 5;

AA5 is Cys when AA4 is absent in structure I;

AA4 is Lys, or Dah, or Orn, or Dab, or Dap, when AA5 in structure I is absent;

AA6 in Structure I is NH₂Or is OH;

r1 in structure I is cholesterol mono-ester succinate, or is 2-cholesteryl acetate, or is 2-cholesteryl propionate, or is 3-cholesteryl propionate, or 2-cholesteryl butyrate, or is 2-cholesteryl isobutyrate, or is 3-cholesteryl butyrate, or is 3-cholesteryl isobutyrate, 4-cholesteryl butyrate, or is 2-cholesteryl valerate, or is 2-cholesteryl isovalerate, or is 3-cholesteryl valerate, or is 5-cholesteryl valerate, or is 2-cholesteryl hexanoate, or is 6-cholesteryl hexanoate, or is 2-cholesteryl heptanoate, or is 7-cholesteryl heptanoate, or is 2-cholesteryl octanoate, or is 8-cholesteryl octanoate, or is 2-cholesteryl octanoate

HO2C(CH2)n1CO-(γGlu)n2-(PEGn3(CH₂)n4CO)n5-；

Wherein: n1 is an integer from 10 to 20;

n2 is an integer from 1 to 5;

n3 is an integer from 1 to 30;

n4 is an integer from 1 to 5;

n5 is an integer from 1 to 5.

R2 in structure I is 2-cholesteryl acetate, or is 2-cholesteryl propionate, or is 3-cholesteryl propionate, 2-cholesteryl butyrate, or is 2-cholesteryl isobutyrate, or is 3-cholesteryl butyrate, or is 3-cholesteryl isobutyrate, 4-cholesteryl butyrate, or is 2-cholesteryl valerate, or is 2-cholesteryl isovalerate, or is 3-cholesteryl valerate, or is 5-cholesteryl valerate, or is 2-cholesteryl hexanoate, or is 6-cholesteryl hexanoate, or is 2-cholesteryl heptanoate, or is 7-cholesteryl heptanoate, or is 2-cholesteryl octanoate, or is 8-cholesteryl octanoate.

The long-acting hepatitis virus entry inhibitor comprises the formed pharmaceutically acceptable salt, solvate, chelate or non-covalent complex, a prodrug based on the compound, or any mixture of the forms.

The invention also provides pharmaceutical compositions comprising a compound according to the invention and the use of a pharmaceutical composition comprising a compound of the invention for the preparation of a medicament for the treatment of a disease.

Further, the pharmaceutical composition is used for treating chronic hepatitis B and hepatitis D.

The Chinese names corresponding to the English abbreviations involved in the present invention are shown in the following table:

TABLE 1

The long-acting hepatitis virus entry inhibitor and the application thereof provided by the invention have the advantages that the used raw materials and reagents can be purchased from the market.

The invention is further illustrated by the following examples:

EXAMPLE 1 preparation of Compound 1

Ac-Gly-Thr-Asn-Leu-Ser-Val-Pro-Asn-Pro-Leu-Gly-Phe-Phe-Pro-Asp-His-Gln-Leu-Asp-Pro-Ala-Phe-Gly-Ala-Asn-Ser-Asn-Asn-Pro-Asp-Trp-Asp-Phe-Asn-Pro-Asn-Lys-Asp-His-Trp-Pro-Glu-Ala-Asn-Lys-Val-Gly-Lys (AEEA-AEEA-gamma Glu-octadecanedioic acid) -NH-Gly-Lys₂

The preparation method comprises the following steps: preparing peptide resin by adopting a solid-phase polypeptide synthesis method, carrying out acidolysis on the peptide resin to obtain a crude product, and finally purifying the crude product to obtain a pure product; the step of preparing the peptide resin by the solid-phase polypeptide synthesis method is to sequentially insert corresponding protective amino acids or fragments in the following sequences on a carrier resin by the solid-phase coupling synthesis method to prepare the peptide resin:

in the preparation method, the dosage of the Fmoc-protected amino acid or the protected amino acid fragment is 1.2 to 6 times of the total mole number of the charged resin; preferably 2.5 to 3.5 times.

In the preparation method, the substitution value of the carrier resin is 0.2-1.0 mmol/g resin, and the preferable substitution value is 0.3-0.5 mmol/g resin.

In a preferred embodiment of the present invention, the solid-phase coupling synthesis method comprises: and (3) after the Fmoc protecting group of the protected amino acid-resin obtained in the previous step is removed, carrying out coupling reaction with the next protected amino acid. The deprotection time for removing Fmoc protection is 10-60 minutes, and preferably 15-25 minutes. The coupling reaction time is 60-300 minutes, and preferably 100-140 minutes.

The coupling reaction needs to add a condensation reagent, and the condensation reagent is selected from one of DIC (N, N-diisopropyl carbodiimide), N, N-dicyclohexylcarbodiimide, benzotriazole-1-yl-oxy tripyrrolidinophosphonium hexafluorophosphate, 2- (7-aza-1H-benzotriazole-1-yl) -1,1,3, 3-tetramethylurea hexafluorophosphate, benzotriazole-N, N, N ', N' -tetramethylurea hexafluorophosphate or O-benzotriazole-N, N, N ', N' -tetramethylurea tetrafluoroborate; n, N-diisopropylcarbodiimide is preferred. The molar consumption of the condensation reagent is 1.2-6 times of the total molar number of amino groups in the amino resin, and preferably 2.5-3.5 times.

The coupling reaction needs to add an activating reagent, wherein the activating reagent is selected from 1-hydroxybenzotriazole or N-hydroxy-7-azabenzotriazole, and 1-hydroxybenzotriazole is preferred. The amount of the activating agent is 1.2 to 6 times, preferably 2.5 to 3.5 times of the total mole number of the amino groups in the amino resin.

As a preferable scheme of the invention, the reagent for removing Fmoc protection is PIP/DMF (piperidine/N, N-dimethylformamide) mixed solution, and the piperidine content in the mixed solution is 10-30% (V). The dosage of the Fmoc protection removing reagent is 5-15 mL per gram of amino resin, and preferably 8-12 mL per gram of amino resin.

Preferably, the peptide resin is subjected to acidolysis while removing the resin and side chain protecting groups to obtain a crude product:

more preferably, the acidolysis agent used in the acidolysis of the peptide resin is a mixed solvent of trifluoroacetic acid (TFA), 1, 2-Ethanedithiol (EDT) and water, and the volume ratio of the mixed solvent is as follows: 80-95% of TFA, 1-10% of EDT and the balance of water.

More preferably, the volume ratio of the mixed solvent is: 89-91% of TFA, 4-6% of EDT and the balance of water. Optimally, the volume ratio of the mixed solvent is as follows: TFA 90%, EDT 5%, balance water.

The dosage of the acidolysis agent is 4-15 mL per gram of the peptide resin; preferably, 7-10 mL of acidolysis agent is required per gram of peptide resin.

The time for cracking by using the acidolysis agent is 1-6 hours, preferably 3-4 hours at room temperature.

Further, the crude product is purified by high performance liquid chromatography and freeze-dried to obtain a pure product.

1. Synthesis of peptide resins

The peptide resin was prepared by coupling sequentially with protected amino acids as shown in the following table through Fmoc-removal protection and coupling reaction using Rinkamide BHHA resin as a carrier resin. The protected amino acids used in this example correspond to the protected amino acids shown below:

TABLE 2

(1) 1 st protected amino acid inserted into main chain

Dissolving 0.03mol of the 1 st protected amino acid and 0.03mol of HOBt in a proper amount of DMF; and adding 0.03mol DIC slowly into the protected amino acid DMF solution under stirring, and reacting for 30 minutes under stirring at room temperature to obtain an activated protected amino acid solution for later use.

0.01mol of Rink amide MBHA resin (substitution value about 0.4mmol/g) is taken, deprotected by 20% PIP/DMF solution for 25 minutes, washed and filtered to obtain Fmoc-removed resin.

And adding the activated 1 st protected amino acid solution into the Fmoc-removed resin, performing coupling reaction for 60-300 minutes, and filtering and washing to obtain the resin containing 1 protected amino acid.

(2) Protected amino acids of 2-49 th of main chain

And sequentially inoculating the corresponding 2 nd to 49 th protected amino acids by the same method for inoculating the 1 st protected amino acid of the main chain to obtain the resin containing 49 amino acids of the main chain.

(3) Side chain insertion of the 1 st protected amino acid

Dissolving 0.03mol of the 1 st protected amino acid of the side chain and 0.03mol of HOBt in a proper amount of DMF; and adding 0.03mol DIC slowly into the protected amino acid DMF solution under stirring, and reacting for 30 minutes under stirring at room temperature to obtain the activated protected amino acid solution.

Taking 2.5mmol of tetratriphenylphosphine palladium and 25mmol of phenylsilane, dissolving with a proper amount of dichloromethane, deprotecting for 4 hours, filtering and washing to obtain a resin without Alloc for later use.

Adding the activated side chain 1 st protected amino acid solution into the Alloc-removed resin, performing coupling reaction for 60-300 minutes, filtering and washing to obtain the side chain 1 st protected amino acid-containing resin.

(4) 2-4 protective amino acids of grafted side chain

And sequentially inoculating 2 nd to 4 th protected amino acids and single protected fatty acids corresponding to side chains by adopting the same method for inoculating the 1 st protected amino acid to the main chain to obtain the peptide resin.

2. Preparation of crude product

Adding a cleavage reagent (10 mL of cleavage reagent/g of resin) with the volume ratio of TFA, water and EDT (95: 5) into the peptide resin, uniformly stirring, stirring at room temperature for reaction for 3 hours, filtering a reaction mixture by using a sand core funnel, collecting filtrate, washing the resin with a small amount of TFA for 3 times, combining the filtrates, concentrating under reduced pressure, adding anhydrous ether for precipitation, washing the precipitate with anhydrous ether for 3 times, and drying to obtain white-like powder, namely a crude product.

3. Preparation of the pure product

Mixing the crude product with water, stirring, adjusting pH to 8.0 with ammonia water to dissolve completely, filtering the solution with 0.45 μm mixed microporous membrane, and purifying;

purifying by high performance liquid chromatography, wherein the chromatographic packing for purification is 10 μm reversed phase C18, the mobile phase system is 0.1% TFA/water solution-0.1% TFA/acetonitrile solution, the flow rate of a 30mm by 250mm chromatographic column is 20mL/min, eluting by a gradient system, circularly sampling for purification, sampling the crude product solution in the chromatographic column, starting the mobile phase for elution, collecting the main peak, and evaporating acetonitrile to obtain a purified intermediate concentrated solution;

filtering the purified intermediate concentrated solution with 0.45 μm filter membrane for use, and performing salt exchange by high performance liquid chromatography with 1% acetic acid/water solution-acetonitrile as mobile phase system, 10 μm reversed phase C18 as purification chromatographic filler, and 20mL/min of 30 mm/250 mm chromatographic column flow rate (corresponding flow rate can be adjusted according to chromatographic columns of different specifications); the method comprises the steps of adopting a gradient elution and circulation loading method, loading a sample into a chromatographic column, starting mobile phase elution, collecting a map, observing the change of the absorbance, collecting a main salt exchange peak, detecting the purity by using an analysis liquid phase, combining main salt exchange peak solutions, concentrating under reduced pressure to obtain a pure acetic acid aqueous solution, and freeze-drying to obtain 6.2g of a pure product, wherein the purity is 98.1%, the total yield is 10.2%, and the molecular weight is 6074.8 (100% M + H).

EXAMPLE 2 preparation of Compound 2

Ac-Gly-Thr-Asn-Leu-Ser-Val-Pro-Asn-Pro-Leu-Gly-Phe-Phe-Pro-

Asp-His-Gln-Leu-Asp-Pro-Ala-Phe-Gly-Ala-Asn-Ser-Asn-Asn-

Pro-Asp-Trp-Asp-Phe-Asn-Pro-Asn-Lys-Asp-His-Trp-Pro-Glu-

Ala-Asn-Lys-Val-Gly-Lys(PEG₅CH₂CO-gamma Glu-octadecanedioic acid) -NH₂

The procedure is as in example 1, using the protected amino acids as in the following table:

TABLE 3

6.5g of pure product was obtained, with a purity of 97.5%, a total yield of 10.7% and a molecular weight of 6062.2 (100% M + H).

EXAMPLE 3 preparation of Compound 3

Gly-Thr-Asn-Leu-Ser-Val-Pro-Asn-Pro-Leu-Gly-Phe-Phe-Pro-Asp-His-Gln-Leu-Asp-Pro-Ala-Phe-Gly-Ala-Asn-Ser-Asn-Asn-Asn-Pro-Asp-Trp-Asp-Phe-Asn-Pro-Asn-Lys-Asp-His-Trp-Pro-Glu-Ala-Asn-Lys-Val-Gly-Cys (2-cholesteryl acetate) -NH₂

The preparation method is the same as that of example 1, the peptide resin is synthesized, the crude linear peptide after acidolysis is grafted with a side chain under the catalysis of alkali to obtain the crude product, and the used protected amino acids are as follows:

TABLE 4

3.5g of a pure product is obtained, the purity is 98.4 percent, and the total yield is 6.1 percent. The molecular weight was 5719.8 (100% M + H).

EXAMPLE 4 preparation of Compound 4

Gly-Thr-Asn-Leu-Ser-Val-Pro-Asn-Pro-Leu-Gly-Phe-Phe-Pro-Asp-His-Gln-Leu-Asp-Pro-Ala-Phe-Gly-Ala-Asn-Ser-Asn-Asn-Pro-Asp-Trp-Asp-Phe-Asn-Pro-Asn-Lys-Asp-His-Trp-Pro-Glu-Ala-Asn-Lys-Val-Gly-Lys (cholesterol succinate monoester) -NH₂

TABLE 5

6.4g of pure product is obtained, the purity is 97.9 percent, and the total yield is 11.1 percent. The molecular weight was 5785.6 (100% M + H).

EXAMPLE 5 preparation of Compound 5

Gly-Thr-Asn-Leu-Ser-Val-Pro-Asn-Pro-Leu-Gly-Phe-Phe-Pro-Asp-His-Gln-Leu-Asp-Pro-Ala-Phe-Gly-Ala-Asn-Ser-Asn-Asn-Pro-Asp-Trp-Asp-Phe-Asn-Pro-Asn-Lys-Asp-His-Trp-Pro-Glu-Ala-Asn-Lys-Val-Gly-AEEA-Lys (cholesterol succinate monoester) -NH₂

TABLE 6

4.7g of pure product is obtained, the purity is 97.2 percent, and the total yield is 7.9 percent. The molecular weight was 5930.9 (100% M + H).

EXAMPLE 6 preparation of Compound 6

Gly-Thr-Asn-Leu-Ser-Val-Pro-Asn-Pro-Leu-Gly-Phe-Phe-Pro-Asp-His-Gln-Leu-Asp-Pro-Ala-Phe-Gly-Ala-Asn-Ser-Asn-Asn-Pro-Asp-Trp-Asp-Phe-Asn-Pro-Asn-Lys-Asp-His-Trp-Pro-Glu-Ala-Asn-Lys-Val-Gly-PEG₅CH₂CO-Lys (cholesterol succinate monoester) -NH₂

TABLE 7

5.6g of pure product is obtained, the purity is 98.1 percent, and the total yield is 9.2 percent. The molecular weight was 6063.2 (100% M + H).

EXAMPLE 6 determination of Primary pharmacokinetic Properties

Each compound was divided into two dosing groups: SD rats, 4 males and females per group.

Tail vein intravenous injection group: the dose is 1mg/kg, rat orbital veins are respectively bled before (0h) and 30min, 1h, 2h, 4h, 8h, 24h, 48h, 96h and 144h after administration, and plasma samples are centrifugally separated.

Subcutaneous administration group: the dose is 1mg/kg, rat orbital veins are bled before administration (0h) and 1h, 2h, 3h, 4h, 8h, 24h, 48h, 96h, 144h after administration, and plasma samples are centrifuged.

Plasma concentrations of the corresponding compounds in plasma samples of SD rats were measured by the liquid chromatography-mass spectrometry method, and the half-lives of the compounds after intravenous and subcutaneous administration in SD rats under Subcutaneous (SC) administration are shown in the following table:

TABLE 8

Compound (I)	t_1/2(h)
		Compound 1	8.9
Compound 2	8.6
		Compound 3	11.0
Compound 4	12.1
		Compound 5	13.5
Compound 6	14.7

The foregoing is only a preferred embodiment of the present invention, and it should be noted that, for those skilled in the art, various modifications and decorations can be made without departing from the principle of the present invention, and these modifications and decorations should also be regarded as the protection scope of the present invention.

Claims

1. The polypeptide is characterized in that the structure is shown as formula I:

AA1-Gly-Thr-Asn-Leu-Ser-Val-Pro-Asn-Pro-Leu-Gly-Phe-Phe-Pro-Asp-His-Gln-Leu-Asp-Pro-Ala-Phe-Gly-Ala-Asn-Ser-Asn-Asn-Pro-Asp-Trp-Asp-Phe-Asn-Pro-Asn-Lys-Asp-His-Trp-Pro-Glu-Ala-Asn-AA2-Val-Gly-AA3-AA4(R1)-AA5(R2)-AA6

formula I

AA1 in formula I is selected from Ac or H; and/or

AA2 in formula I is selected from Lys, Gln, Glu, Arg or Cit; and/or

AA3 in formula I is selected from PEG_m1(CH₂)_m2CO-, or is absent;

wherein: m1 is an integer from 1 to 10;

m2 is an integer from 1 to 5; and/or

AA5 is selected from Cys when AA4 is absent in formula I; and/or

AA4 is selected from Lys, Dah, Orn, Dab or Dap when AA5 is absent in formula I; and/or

AA6 in formula I is selected from NH₂Or OH; and/or

R1 in formula I is selected from succinic acid cholesterol monoester, 2-cholesterol acetic acid, 2-cholesterol propionic acid, 3-cholesterol propionic acid, 2-cholesterol butyric acid, 2-cholesterol isobutyric acid,3-Cholesterol butanoic acid, 3-Cholesterol isobutyric acid, 4-Cholesterol butanoic acid, 2-Cholesterol pentanoic acid, 2-Cholesterol isovaleric acid, 3-Cholesterol pentanoic acid, 5-Cholesterol pentanoic acid, 2-Cholesterol hexanoic acid, 6-Cholesterol hexanoic acid, 2-Cholesterol heptanoic acid, 7-Cholesterol heptanoic acid, 2-Cholesterol octanoic acid, 8-Cholesterol octanoic acid, HO2C (CH2) n1CO- (gamma Glu) n2- (PEGn3(CH 2)₂)n4CO)n5-；

Wherein: n1 is an integer from 10 to 20;

n2 is an integer from 1 to 5;

n3 is an integer from 1 to 30;

n4 is an integer from 1 to 5;

n5 is an integer from 1 to 5; and/or

2. Use of the polypeptide or derivative thereof according to claim 1 for the preparation of a long-acting hepatitis virus entry inhibitor.

3. The use of claim 2, wherein the derivative comprises a pharmaceutically acceptable salt, solvate, chelate or non-covalent complex of the polypeptide based on a mixture of one or more of the prodrugs on which the compound is based.

4. Use of a polypeptide or derivative thereof according to claim 1 in the manufacture of a medicament for the treatment of liver disease.

5. The use of claim 4, wherein the liver disease comprises hepatitis.

6. The use of claim 5, wherein the hepatitis comprises chronic hepatitis B and/or hepatitis D.

7. The use as claimed in any one of claims 4 to 6 wherein the derivative comprises a pharmaceutically acceptable salt, solvate, chelate or non-covalent complex of the polypeptide based on a mixture of one or more than two prodrugs thereof.

8. A long-acting hepatitis virus entry inhibitor comprising the polypeptide of claim 1 or a derivative thereof and a pharmaceutically acceptable excipient.

9. A medicament for the treatment of hepatitis comprising the polypeptide of claim 1 or a derivative thereof and a pharmaceutically acceptable excipient.

10. The medicament of claim 9, wherein the hepatitis comprises chronic hepatitis B and/or hepatitis D.