WO2020259539A1 - 4-吡啶取代酞嗪酮类化合物、其制备方法、药物组合物及用途 - Google Patents

4-吡啶取代酞嗪酮类化合物、其制备方法、药物组合物及用途 Download PDF

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WO2020259539A1
WO2020259539A1 PCT/CN2020/097944 CN2020097944W WO2020259539A1 WO 2020259539 A1 WO2020259539 A1 WO 2020259539A1 CN 2020097944 W CN2020097944 W CN 2020097944W WO 2020259539 A1 WO2020259539 A1 WO 2020259539A1
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胡有洪
左建平
陈五红
杨莉
赵琪良
刘菲菲
曾艳萍
童贤崑
曾丽敏
魏爱环
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中国科学院上海药物研究所
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Definitions

  • the invention belongs to the technical field of medicine, and relates to 4-pyridine substituted phthalazinone compounds, a preparation method thereof, a pharmaceutical composition and uses.
  • hepatitis B virus infection is still a worldwide disease that seriously endangers public health.
  • interferon and nucleoside anti-hepatitis B virus drugs are the two main methods for the treatment of hepatitis B virus infection.
  • interferon has shortcomings such as poor tolerance, multiple adverse reactions, and high cost; and the six nucleoside drugs currently on the market (lamivudine, adefovir dipivoxil, entecarn, telbivudine, Tenofovir and Kravudine) both act on the reverse transcriptase of hepatitis B virus.
  • the present invention provides a non-nucleoside small molecule HBV virus inhibitor with novel structure and oral administration.
  • the compound in the Chinese patent application with application number 201810905518.8 is a new type of high-efficiency and low-toxicity non-nucleoside anti-HBV small molecule candidate compound (the entire content of the patent application is incorporated into this application). However, the bioavailability and druggability of this compound need to be further improved.
  • the present invention provides a 4-pyridine-substituted phthalazinone compound as shown in formula I.
  • This compound has improved pharmacophysical properties and in vivo pharmacokinetic properties, high oral bioavailability, and good druggability , Is a novel structure, oral non-nucleoside small molecule HBV inhibitor.
  • the present invention provides a compound of formula I or a pharmaceutically acceptable salt thereof:
  • R A is selected from halo or cyano
  • R B is selected from halogen
  • R C is selected from H, alkyl, cycloalkyl
  • L is a linking group, a group obtained by replacing any carbon atom selected from an alkylene group and an alkylene group with at least one group selected from -O- and -CO-, a cycloalkylene group;
  • R D is selected from
  • R 1 is H, alkyl, aryl, substituted with a group selected from hydroxyl, mercapto, alkylthio, aminoalkanoyl, carboxyl, amino, guanidino, ureido, aryl, hydroxyaryl, and heteroaryl ⁇ alkyl;
  • R 2 is hydrogen or alkyl
  • R 3 is hydrogen, tert-butoxycarbonyl
  • W is selected from alkyl, haloalkyl, cycloalkyl, heterocyclyl, heterochain alkyl, aryl, heteroaryl, alkoxy, amino; wherein, the alkyl, cycloalkyl, heterocyclyl, hetero Alkyl, aryl, alkoxy, and amino groups are optionally independently substituted with 1, 2, 3, or 4 groups selected from alkyl, carboxy, alkoxy, heterocyclyl, aryl, and heteroaryl. ;
  • X is a single bond or an alkylene group
  • R 5 and R 6 are each independently selected from -OH, aryloxy group, alkyl group, any carbon atom is replaced by at least one group selected from -O-, -CO-, -S- group, amino acid
  • the amino acid residue mentioned here refers to the group obtained by removing one hydrogen from the ⁇ -amino group of the amino acid.
  • the hydroxyl group in the carboxyl group of the amino acid residue is optionally substituted by an alkoxy group, an aryloxy group, or a heteroaryloxy group.
  • amino acid mentioned here is selected from the following amino acids of L configuration, D configuration, or a mixed configuration of L and D: glycine, proline, alanine, valine, Norvaline, Leucine, Isoleucine, Norleucine, 2-tert-Butylglycine, 2-Phenylglycine, Phenylalanine, Tyrosine, Tryptophan, Histidine, Serine, homoserine, threonine, cysteine, S-methylcysteine, homocysteine, methionine, asparagine, glutamine, aspartic acid, glutamic acid, lysine Acid, hydroxylysine, ornithine, 2,4-diaminobutyric acid, 2,3-diaminopropionic acid, arginine, citrulline; or R 5 , R 6 and P together form a heterocyclic group
  • the heterocyclic group is optionally substituted by a halogenated
  • halogen is selected from F, Cl, Br or I;
  • alkyl groups in any groups such as “alkyl”, “haloalkyl”, “alkoxy”, “alkylthio” and the like are independently C 1 -C 20 linear or branched.
  • alkylene group in any group such as “alkylene”, “heterocyclylenealkyl”, “aminoalkanoyl” and the like are each independently selected from C 1 -C 20 linear or Branch alkylene, optionally C 1 -C 15 linear or branched alkylene, optionally C 1 -C 10 linear or branched alkylene, optionally C 1 -C 8 Linear or branched alkylene, optionally C 1 -C 6 linear or branched alkylene, optionally C 1 -C 4 linear or branched alkylene, optionally C 2 -C 6 linear or branched chain alkylene, optionally C 3 -C 6 linear or branched alkylene;
  • the "hetero-chain hydrocarbon group” is a linear or branched saturated or unsaturated C 1 -C chain containing 1, 2, 3 or 4 heteroatoms selected from N, O and S. 20 Hetero-chain hydrocarbon structure, optionally, a linear or branched saturated or unsaturated C 1 -C 20 hetero-chain hydrocarbon structure containing 1, 2, 3 or 4 N in the chain, optionally, It is a linear or branched saturated or unsaturated C 1 -C 10 heterochain hydrocarbon structure containing 1, 2, 3 or 4 N in the chain;
  • the "cycloalkyl group” is a C 3 -C 10 monocyclic or bicyclic cycloalkyl group, optionally, a C 3 -C 7 monocyclic cycloalkyl group, optionally, a cyclopropyl group, Cyclobutyl, cyclopentyl, cyclohexyl or cycloheptyl;
  • the "cycloalkylene” is a C 3 -C 10 monocyclic or bicyclic cycloalkylene, alternatively, a C 3 -C 8 monocyclic cycloalkylene, optionally, a C 3 -C 8 monocyclic cycloalkylene Cyclopropyl, cyclobutylene, cyclopentylene, cyclohexylene, cycloheptylene and cyclooctylene.
  • the heterocyclic ring contained in any group such as “heterocyclylene”, “heterocyclylene”, “heterocyclylenealkyl” and the like contains 1, 2, or 3 selected groups on the ring.
  • a 3-10 membered non-aromatic heterocyclic ring from heteroatoms of N, O, S optionally, the heterocyclic ring is a 3-10 membered non-aromatic heterocyclic ring containing 1 or 2 heteroatoms selected from N, O Aromatic ring;
  • the heterocyclic ring is a 3-6 membered non-aromatic ring containing 1 or 2 heteroatoms selected from N and O; alternatively, the heterocyclic ring is a ring containing 1 or 2
  • the heterocyclic ring is a 3-6 membered non-aromatic ring with one or two heteroatoms selected from N and S on the ring;
  • the aryl group in any of the "aryl”, “hydroxyaryl”, “haloaryl”, “aryloxy” and other groups is a 6-10 membered aryl group; Phenyl or naphthyl, optionally phenyl, 1-naphthyl, 2-naphthyl;
  • the ring of the heteroaryl group contained in any group in the "heteroaryloxy group", “heteroaryl group” etc., “aryloxy group” contains 1-3 selected from N, O and 5-10 membered heteroaromatic ring of heteroatom in S; alternatively, 5-10 membered heteroaromatic ring containing 1-2 heteroatoms selected from N, O and S; alternatively, the The heteroaromatic ring is selected from pyridine ring, pyrrole ring, pyrimidine ring, pyrazine ring, pyridazine ring, thiophene ring, furan ring; optionally, the heteroaryl group is selected from pyridine-2-yl, pyridine-3- Yl, pyridin-4-yl, pyridazin-3-yl, pyridazin-4-yl, pyrimidin-2-yl, pyrimidin-4-yl, pyrimidin-5-yl, pyrazin-2
  • alkyl is a group obtained by replacing any carbon atom of an alkylene group with at least one group selected from -O- and -CO-
  • "Aryl”, “heteroaryl” or “benzyl” are each independently substituted with a substituent selected from the following groups: alkyl (such as methyl, ethyl, n-propyl, isopropyl, n-butyl, Isobutyl, tert-butyl), alkenyl (such as vinyl, propynyl), alkynyl (such as ethynyl, propynyl), hydroxyl, alkoxy (such as methoxy, ethoxy, n-propoxy) Group, isopropoxy, tert-butoxy), acyloxy (such as formyloxy, acetoxy, propionyloxy, butyryloxy, isopropylcarbonyloxy,
  • R A is selected from chlorine and cyano
  • R B is selected from chlorine
  • R C is selected from H, methyl, cyclopropyl
  • L is selected from -CH 2 CH 2 -, -CH 2 CH 2 CH 2 -, -CH 2 CH 2 CH 2 CH 2 -, CH 2 CH 2 OCH 2 CH 2 -,
  • R D is selected from amino acid residues
  • the amino acid residue mentioned here refers to the group obtained by removing the hydroxyl group from the carboxyl group in the amino acid
  • the amino acid residue is selected from the following amino acids in the L configuration, D configuration, or mixed configuration of L and D Residues: glycine, proline, alanine, valine, norvaline, leucine, isoleucine, norleucine, 2-tert-butylglycine, 2-phenylglycine, benzene Alanine, tyrosine, phenylethyline, tryptophan, histidine, serine, homoserine, threonine, cysteine, S-methylcysteine, homocysteine, Methionine, asparagine, glutamine, aspartic acid, glutamic acid, lysine, hydroxylysine, ornithine, 2,4-diaminobutyric acid, 2,3-diaminopropionic acid, Arginine, cit
  • W is selected from C 1-6 alkyl, carboxy C 1-6 alkyl, C 5-10 heterocyclyl C 1-6 alkyl, C 1-6 alkoxy C 6-10 aryl, C 1-6 Alkoxy, one or two C 1-6 alkyl substituted amino groups, C 5-10 heterocyclic group;
  • X is a single bond, -CH 2 -, or -CH 2 (CH 3 )-;
  • R 5 , R 6 and P together form a C 5-10 heterocyclic group, the C 5-10 heterocyclic group is optionally substituted by a halogenated phenyl group, and the C 5-10 heterocyclic group is optionally combined with a phenyl group.
  • the compound of formula I is selected from the following compounds:
  • the pharmaceutically acceptable salts include anionic and cationic salts of the compound of formula I.
  • the pharmaceutically acceptable salt includes the alkali metal salt, alkaline earth metal salt, and ammonium salt of the compound of formula I; optionally, the alkali metal includes sodium, potassium, lithium, cesium, and the alkaline earth Metals include magnesium, calcium, and strontium.
  • the pharmaceutically acceptable salt includes a salt formed by a compound of formula I and an organic base; optionally, the organic base includes trialkylamine, pyridine, quinoline, piperidine, imidazole, and picoline , Dimethylaminopyridine, dimethylaniline, N-alkylmorpholine, 1,5-diazabicyclo[4.3.0]nonene-5, 1,8-diazabicyclo[5.4.0] ⁇ Monocarbene-7, 1,4-diazabicyclo[2.2.2]octane; optionally, the trialkylamine includes trimethylamine, triethylamine, N-ethyldiisopropylamine; optional Specifically, the N-alkylmorpholine includes N-methylmorpholine.
  • the organic base includes trialkylamine, pyridine, quinoline, piperidine, imidazole, and picoline , Dimethylaminopyridine, dimethylaniline, N-alkylmorpholine, 1,5-diaza
  • the pharmaceutically acceptable salt includes a salt formed by a compound of formula I and an acid; optionally, the acid includes an inorganic acid and an organic acid; optionally, the inorganic acid includes hydrochloric acid and hydrobromic acid , Hydroiodic acid, sulfuric acid, nitric acid, phosphoric acid, carbonic acid; alternatively, the organic acid includes formic acid, acetic acid, propionic acid, oxalic acid, malonic acid, succinic acid, fumaric acid, maleic acid, lactic acid, malic acid , Citric acid, citric acid, tartaric acid, carbonic acid, picric acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, glutamic acid, pamoic acid.
  • the acid includes an inorganic acid and an organic acid
  • the inorganic acid includes hydrochloric acid and hydrobromic acid , Hydroiodic acid, sulfuric acid, nitric acid, phospho
  • the reaction route includes: condensing Hu1 compound with R D OH to form an ester to obtain a compound of formula I, wherein, in formula Hu1 and formula I, R A , R B , and L are as defined in any one of claims 1-5 Said, R C is H; in formula I, R D is defined as described in any one of claims 1-5;
  • the reaction route includes: condensing Hu1-1 compound with R D OH to form an ester reaction to obtain a compound of formula Hu1-2, wherein R D in the Hu1-2 compound is defined by any one of claims 1-5 containing protection Then, the compound of formula Hu1-2 is reacted with a halide into R C , and the protecting group in R D is optionally removed to obtain the compound of formula I, wherein, formula Hu1-1, formula Hu1-2 and In formula I, R A , R B and L are defined as described in any one of claims 1-5; in formula I, R D is defined as described in any one of claims 1-5; in formula I, R C is a group other than hydrogen as defined in any one of claims 1-5;
  • a protecting group is used to protect the compound of formula Hu1, Hu1-1, and the compound of formula R D OH, and after the condensation reaction to an ester, deprotection is optionally performed;
  • the removal of these protective groups is carried out according to a conventional method, preferably by reaction with a strong acid such as hydrochloric acid, hydrobromic acid or trifluoroacetic acid in an inert solvent such as 1,4-dioxane or dichloromethane.
  • a strong acid such as hydrochloric acid, hydrobromic acid or trifluoroacetic acid in an inert solvent such as 1,4-dioxane or dichloromethane.
  • a suitable condensing agent is selected from carbodiimides such as N,N'-dicyclohexylcarbodiimide (DCC) or N-(3-diimide). Methylaminoisopropyl)-N'-ethylcarbodiimide hydrochloride (EDC).
  • DCC N,N'-dicyclohexylcarbodiimide
  • EDC Methylaminoisopropyl)-N'-ethylcarbodiimide hydrochloride
  • the condensation to ester reaction is preferably carried out in the presence of a base.
  • a base it is preferable to use alkali metal carbonates such as sodium carbonate or potassium carbonate, or organic bases such as triethylamine, N-methylmorpholine, N-methylpiperidine, N,N-diisopropylethylamine Or 4-N,N-dimethylaminopyridine.
  • the reaction temperature of the condensation to ester is 0°C to +60°C, preferably +20°C to +40°C.
  • the reaction is preferably carried out at atmospheric pressure.
  • the compound of formula I or a pharmaceutically acceptable salt thereof can also be prepared by the following method:
  • the amino acid residues here refer to the groups obtained by removing one hydrogen from the ⁇ -amino group of the amino acid.
  • the hydroxyl group in the carboxyl group of the amino acid residue is optionally alkoxy, Group substitution of aryloxy, heteroaryloxy or benzyloxy;
  • the amino acid mentioned here is selected from the following amino acids in the L configuration, D configuration, or mixed configuration of L and D: glycine, proline, propyl Acid, valine, norvaline, leucine, isoleucine, norleucine, 2-tert-butylglycine, 2-phenylglycine, phenylalanine, tyrosine, color Acid, histidine, serine, homoserine, threonine, cysteine, S-methylcysteine, homocysteine, methionine, asparagine, glutamine, aspartic acid , Glutamic acid, lysine, hydroxylysine, ornithine, 2,4-diaminobutyric acid, 2,3-diaminopropionic acid, arginine, citrulline;
  • X is as described above, and Q 1 and Q 2 are each independently H and an alkyl group;
  • R A , R B , R C , L and X are as described above;
  • the methods for introducing R 5 and R 6 groups include phosphoramidating the compound of formula Hu3 with the corresponding amino acid to obtain the corresponding phosphoramide formula I, or reacting with the corresponding halide or alcohol to obtain the phosphoric acid ester formula I compound.
  • the Hu3 compound and the corresponding amino acid undergo amino acid condensation to form a phosphoramide reaction.
  • the suitable condensing agent is selected from carbodiimides such as N,N'-dicyclohexylcarbodiimide (DCC) or N-(3-dimethyl Aminoisopropyl)-N'-ethylcarbodiimide hydrochloride (EDC).
  • DCC N,N'-dicyclohexylcarbodiimide
  • EDC N-(3-dimethyl Aminoisopropyl)-N'-ethylcarbodiimide hydrochloride
  • the condensation reaction to amide is preferably carried out in the presence of a base.
  • alkali metal carbonates such as sodium carbonate or potassium carbonate
  • organic bases such as triethylamine, N-methylmorpholine, N-methylpiperidine, N,N-diisopropylethylamine Or 4-N,N-dimethylaminopyridine.
  • the Hu3 compound and the corresponding alcohol undergo an alkyd condensation reaction to form a phosphate ester.
  • the suitable condensing agent is selected from carbodiimides such as N,N'-dicyclohexylcarbodiimide (DCC) or N-(3-dimethyl Aminoisopropyl)-N'-ethylcarbodiimide hydrochloride (EDC).
  • DCC N,N'-dicyclohexylcarbodiimide
  • EDC N-(3-dimethyl Aminoisopropyl)-N'-ethylcarbodiimide hydrochloride
  • the condensation reaction to phosphate ester is preferably carried out in the presence of a base.
  • alkali metal carbonates such as sodium carbonate or potassium carbonate
  • organic bases such as triethylamine, N-methylmorpholine, N-methylpiperidine, N,N-diisopropylethylamine Or 4-N,N-dimethylaminopyridine.
  • the Hu3 compound undergoes nucleophilic substitution to phosphoric acid ester under alkaline conditions with the corresponding chlorinated compound.
  • sodium hydride and potassium hydride are preferred.
  • the protective group When a multifunctional organic compound reacts, in order to make the reaction only occur at the desired functional group and avoid other functional groups from being affected, the other functional groups should be protected before the reaction and restored after the reaction is complete.
  • Protecting group is a general term for such groups that can provide protection to the other functional groups.
  • the protecting group of the amino group may be a tert-butoxy carbyl (Boc).
  • the protective group of the hydroxyl or carboxyl functional group preferably uses tert-butyl, ethyl, or methyl.
  • the removal of these protective groups is carried out according to a conventional method, preferably by reaction with a strong acid such as hydrochloric acid, hydrobromic acid or trifluoroacetic acid in an inert solvent such as 1,4-dioxane or dichloromethane.
  • a strong acid such as hydrochloric acid, hydrobromic acid or trifluoroacetic acid in an inert solvent such as 1,4-dioxane or dichloromethane.
  • the compound of formula Hu1 can be prepared according to conventional methods in the literature (for example, it can be prepared by referring to the preparation method in the patent application with application number 201810905518.8).
  • the above-mentioned pharmaceutically acceptable salt of the compound of formula I can be prepared by dissolving the compound of formula I in a corresponding acid-saturated alcohol solution, for example: dissolving the compound of formula I in hydrogen chloride saturated The methanol solution is stirred at room temperature for 30 minutes, and the solvent is evaporated to dryness to obtain the corresponding hydrochloride of the compound of formula I.
  • a pharmaceutical composition which comprises a therapeutically effective amount of one or more of the above-mentioned compound of formula I or a pharmaceutically acceptable salt thereof and optionally a pharmaceutically acceptable carrier.
  • the above-mentioned pharmaceutically acceptable carriers refer to conventional pharmaceutical carriers in the pharmaceutical field, for example: diluents, such as water, etc.; fillers, such as starch, sucrose, etc.; binders, such as cellulose derivatives, alginate, gelatin, poly Vinylpyrrolidone; wetting agents such as glycerin; disintegrating agents such as agar, calcium carbonate and sodium bicarbonate; absorption promoters such as quaternary ammonium compounds; surfactants such as cetyl alcohol; adsorption carriers such as kaolin and bentonite ; Lubricants, such as talc, calcium and magnesium stearate, and polyethylene glycol.
  • other adjuvants such as flavoring and sweetening agents, can also be added to the above-mentioned pharmaceutical composition.
  • the above-mentioned compound of formula I, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition can be administered to patients in need of such treatment by oral, rectal or parenteral administration.
  • oral administration it can be made into conventional solid preparations, such as tablets, powders, granules, capsules, etc., or into liquid preparations, such as water or oil suspensions, or other liquid preparations, such as syrups;
  • parenteral administration it can be made into a solution for injection, water or oily suspension, etc.
  • the present invention also provides the use of the above-mentioned compound of formula I or a pharmaceutically acceptable salt thereof, or the above-mentioned pharmaceutical composition in the preparation of a medicine for preventing and/or treating hepatitis B disease.
  • the present invention also provides the use of the aforementioned compound of formula I or a pharmaceutically acceptable salt thereof, or the aforementioned pharmaceutical composition in the preparation of hepatitis B virus inhibitors.
  • the present invention also provides the use of the above-mentioned compound of formula I or a pharmaceutically acceptable salt thereof, or the above-mentioned pharmaceutical composition in the preparation of a drug for inhibiting virus replication, the virus including hepatitis A virus, hepatitis B virus, hepatitis C virus, and influenza virus , Adenovirus, HIV, herpes virus, human papilloma virus.
  • the present invention also provides a method for preventing and/or treating hepatitis B disease, which comprises administering an effective amount of the above-mentioned compound of formula I or a pharmaceutically acceptable salt thereof, or the above-mentioned pharmaceutical composition to a patient in need.
  • the present invention also provides a method for inhibiting hepatitis B virus, which comprises administering an effective amount of the above-mentioned compound of formula I or a pharmaceutically acceptable salt thereof, or the above-mentioned pharmaceutical composition to a patient in need.
  • the present invention also provides a method for inhibiting virus replication, which comprises administering to a host an effective amount of the above-mentioned compound of formula I or a pharmaceutically acceptable salt thereof, or the above-mentioned pharmaceutical composition.
  • the viruses include hepatitis A virus, hepatitis B virus, hepatitis C virus, influenza virus, adenovirus, HIV, herpes virus, and human papilloma virus.
  • the compound has a highly effective activity of inhibiting HBV DNA replication at the cellular level, and is less toxic to the growth of HepG2.2.15 cells.
  • the compound has good liver targeting, can stably exist and be enriched in the liver, and is an effective hepatitis B virus inhibitor.
  • Figure 1 shows a schematic diagram of the change of the virus HBV DNA content in the peripheral blood of AAV mice in Test Example 3 over time.
  • Figure 2 shows a schematic diagram of the change of the viral HBsAg content in the peripheral blood of the AAV mice in Test Example 3 over time.
  • Figure 3 shows a schematic diagram of the change of the viral HBeAg content in the peripheral blood of the AAV mice in Test Example 3 over time.
  • Figure 4 shows a schematic diagram of the viral HBcAg content in the liver of AAV mice 4 weeks after administration in Test Example 3.
  • the core protein-positive cells appear dark black, and the negative cells appear light black.
  • the proton nuclear magnetic resonance spectrum was recorded with a Bruker AMX-400, Gemini-300 or AMX-600 nuclear magnetic resonance instrument, and the unit of chemical shift ⁇ is ppm.
  • the specific rotation is measured by Perkin-Elmer 241 automatic polarimeter, and the microwave used is a CEM-discovery microwave reactor.
  • all reaction solvents were purified according to conventional methods.
  • Silica gel (200-300 mesh) for column chromatography is produced by Qingdao Ocean Chemical Branch.
  • Thin layer chromatography uses GF254 high-efficiency plate, which is produced by Yantai Chemical Research Institute.
  • the preparative thin-layer chromatography plate was prepared by the Shanghai Institute of Materia Medica, Chinese Academy of Sciences, and the stationary phase was prepared with GF254 (HG/T2354-92) silica gel and sodium carboxymethyl cellulose (800-1200), respectively, Qingdao Ocean Chemical Co., Ltd. and Produced by China National Pharmaceutical (Group) Shanghai Chemical Reagent Company.
  • all solvents are analytical reagents, and all reagents used are purchased from Sinopharm Chemical Reagent Co., Ltd. Use iodine, ultraviolet fluorescence and other methods to develop color.
  • the organic solvent was evaporated under reduced pressure in a rotary evaporator.
  • Example 1 The synthetic route of compound 1 is as follows:
  • 3,6-Dichlorophthalazine i-1 (10g, 50.3mmol) was added to acetic acid (150mL) and refluxed at 120°C for 6 hours. After the completion of the reaction was monitored by TLC, the reaction was stopped and the reaction solution was cooled to room temperature. The acetic acid was removed by rotary evaporation to obtain a white solid compound i-2.
  • Boc-L-valine (1.85g, 8.5mmol), bicyclohexylcarbodiimide (1.4g, 6.8mmol) and 4-dimethylaminopyridine (0.34g, 2.8mmol) were added to compound 1 (2.63g 5.67mmol) in dichloromethane (50ml) solution, after the addition, continue to react at room temperature for 1 hour. After the reaction, filter with suction and evaporate the dichloromethane from the filtrate under reduced pressure. The residue is dissolved in ethyl acetate, washed with water and saturated brine successively, the ethyl acetate layer is collected, dried over anhydrous sodium sulfate, and the filtrate is evaporated to dryness. The crude product was obtained, and the crude product was purified by silica gel column chromatography (PE/EA 75/25) to obtain white solid I-1 (3.4 g, 92%).
  • Step b
  • the organic phase is anhydrous After drying over sodium sulfate, filtering, and evaporating the solvent under reduced pressure, the residue was subjected to silica gel column chromatography to obtain white powdery solid compound I-33' (47 mg, 55%).
  • Step b
  • Step b
  • Step b
  • the residue (280mg, 0.44mmol) was dissolved in 20mL of acetonitrile, 0.5mL of thionyl chloride was added, and the reaction was carried out at 75°C for 2 hours. The reaction solution was concentrated to dryness. The residue was dissolved in dry 20mL of dichloromethane and cooled to -78.
  • reaction solution was concentrated to dryness, the residue was dissolved in dry dichloromethane (20ml), L-alanine ethyl ester hydrochloride (0.344g, 2.24mmol), diisopropylethylamine (0.46g) were added , 3.58mmol), react for 1.5 hours, add water, extract 3 times with dichloromethane, wash the organic phase with water, collect the dichloromethane layer, dry with anhydrous sodium sulfate, filter with suction, and evaporate the dichloromethane in the filtrate under reduced pressure to obtain The crude product was purified by silica gel column chromatography (dichloromethane/methanol 90/10) to obtain compound I-41 (0.31 g, 45%) as a white solid.
  • phosphine oxychloride (2.3 g, 15 mmol) was added to a solution of 1 (0.464 g, 1 mmol) in triethyl phosphate (10 mL) and stirred for 1 h.
  • Triethylamine/ammonium bicarbonate buffer solution (1M, pH 7)
  • dichloromethane and water to the residue
  • extract the aqueous phase with dichloromethane 3 times combine the organic phases, dry, and filter.
  • the solvent was removed to obtain the crude product.
  • the crude product was dissolved in a small amount of dichloromethane, and a large amount of cold ether was added to precipitate a solid, which was filtered and washed with ether to obtain the product compound 4 (0.45 g 83%).
  • Step b
  • Step b
  • Test Example 1 The effect of the compound prepared in the example on the ability of hepatitis B virus DNA replication
  • Human hepatoma cell line HepG2.2.15 stably transfected with full-length HBV (provided by Shanghai Institute of Materia Medica, Chinese Academy of Sciences)
  • Incubator (ThermoForma3111); Microplate reader (Molecular Devices Spectra Max 190); Electronic balance; Microscope; Biological safety cabinet (Heal Force safe15); Centrifuge (Eppendorf Centrifuge 5810R); Real-Time PCR (FASTA GEN-DNA fast2000)
  • Lamivudine (3TC) a combination of medicinal chemistry from the Shanghai Institute of Materia Medica, Chinese Academy of Sciences, and DMEM/High Glucose medium (Dulbecco’s modified Eagle’s medium, Hyclone) was used to prepare a 40mM stock solution for use.
  • DMEM/High Glucose culture medium Dulbecco's modified Eagle's medium 1 ⁇ (Hyclone) phosphate buffered saline (PBS, pH7.3, 1L): NaCl, 8.0g; Na 2 HPO 4 , 1.16g; KH 2 PO 4 , 0.2 g; KCl: 0.2g;
  • MTT solution MTT (Sigma), formulated with PBS to 5mg/ml;
  • HepG2.2.15 cells were cultured and passaged according to conventional methods.
  • the medium used is DMEM, which contains 10% (v/v) bovine serum and the selective antibiotic G418, and cultured in an incubator at 37°C and 5% CO 2 for 8 days (change the medium on the 4th day).
  • the compound to be tested was prepared into a 40mM stock solution with DMSO, and the DMEM medium containing 10% Hyclone TM Fetal Bovine Serum was mixed with the specified highest concentration solution and diluted; the positive drug was lamivudine, which also contained 10% Hyclone TM Fetal Bovine Serum's DMEM medium is prepared to the specified concentration.
  • HepG2.2.15 cells were seeded in 96-well plates at 5 ⁇ 10 3 cells/well, cultured for eight days under the action of drugs as described above, 200 ⁇ l of supernatant was taken out and MTT solution was added, lysate was added after 4 hours of culture, and enzymes were used after 12 hours of culture
  • the OD 570 was measured by the calibration instrument and compared with the absorbance of the control well to calculate the percentage of viable cells and calculate the concentration CC 50 required to cause half of the cytotoxicity.
  • step 4) Add all the liquid from step 3) to the DNeasy Mini spin column (Qiagen, Blood & Tissue Kit), centrifuge at 6000rcf*1min, discard the supernatant;
  • Standard curve 1e 7 -1e 4 IU/ml, 1 ⁇ l loading, set a blank hole to detect whether the reaction system is contaminated;
  • the first stage 93°C, 2min
  • the first step 93°C, 45s
  • the second step 55°C, 1min
  • the third stage 45 cycles
  • the first step 93°C, 30s
  • the second step 55°C, 45s
  • Test Collect data after completing the second step of the third stage (55°C, 45s).
  • Table 1 Toxicity of the compounds of formula I in the examples to HepG2.2.15 cells and the activity of inhibiting HBV DNA
  • CC 50 is the effect of the compounds of the examples on the growth of HepG2.2.15 cells, the half (50%) lethal concentration.
  • the IC 50 is the concentration at which the compound of the example inhibits hepatitis B virus DNA replication by half (50%).
  • the IC 50 of the compounds of the examples are all less than 10 ⁇ M, have high activity of inhibiting HBV DNA replication at the cellular level, and are less toxic to the growth of HepG2.2.15 cells.
  • ICR human-1 male mice (body weight: ⁇ 20g), the number of animals in each group is 3 at each time point, and they are administered by intragastric administration at a dose of 20 mg/kg to give compound I-2, compound I-36 and comparative compound 1.
  • the blood sampling time points are arranged as 0min, 15min, 30min, 1h, 2h, 4h, 8h and 24h for the oral group.
  • plasma was prepared by centrifugation, and aliquoted in 20 ⁇ L, and stored at -70°C until analysis. The pharmacokinetic parameters were analyzed and processed using Kinetica 5.0 software (US) non-compartmental model.
  • the PK data of Compound 1 in Table 2 showed that very limited drug-related compounds (including Compound 1 and active metabolite 1-M) were detected in the plasma after oral administration of Compound 1.
  • Compounds of formula I-2 and compounds of formula I-36 can effectively deliver drugs into plasma after oral administration, and the drugs include compound 1 and active metabolite compound 1-M.
  • the PK data in Table 2 shows that the compound represented by formula I of the present invention is a non-nucleoside small molecule HBV virus inhibitory drug suitable for oral administration.
  • mice Take 6-8 weeks old Balb/c male mice (weight: ⁇ 20g), and inject the recombinant virus AAV8-HBV virus solution into the tail vein at a dose of 1 ⁇ 10 11 vg/mouse (Viral genome, vg) to construct the wild type HBV chronic infection mouse model.
  • the infected mice with continuous and stable virus content in the serum were randomly divided into three groups, each with six mice, respectively, the model group (0.5% CMC-Na solvent control group), compound I-2 100mg/ The kg dose group and the entecavir (ETV) 1 mg/kg dose group (positive drug group).
  • the compound I-2 group was orally administered twice a day (Bid), and the model group and the positive drug group were orally administered once a day (Qd).
  • blood was collected from the orbit every week, and the levels of virus and virus-related secreted proteins (HBeAg and HBsAg) in the peripheral blood were detected by qPCR and immunofluorescence respectively; after 4 weeks of drug treatment, the mice were euthanized and the livers were taken , Immunohistochemical detection of HBcAg level in the liver.
  • HBsAg and HBeAg are used as clinical indicators to judge the status of hepatitis B virus infection and functional cure, which have great clinical significance.
  • the compounds of the present invention are particularly suitable for preventing or treating liver diseases, including, for example, liver cancer, liver infections, including hepatitis infections.

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Abstract

4-吡啶取代酞嗪酮类化合物、其制备方法、药物组合物及用途，所述4-吡啶取代酞嗪酮类化合物结构如式I所示，所述式I化合物具有改善的药物理化性质和体内药代动力学性质，口服生物利用度高，成药性好，是一类结构新颖的、可口服的非核苷类小分子HBV病毒抑制剂。

Description

4-吡啶取代酞嗪酮类化合物、其制备方法、药物组合物及用途 技术领域
本发明属于医药技术领域，涉及4-吡啶取代酞嗪酮类化合物、其制备方法、药物组合物及用途。
背景技术
全球每年约有75万人死于乙肝及其并发症，包括肝细胞癌、肝硬化、肝功能衰竭等。因此，乙型肝炎病毒感染仍是一种严重危害公众健康的世界性疾病。
目前，干扰素和核苷类抗乙肝病毒药是治疗乙型肝炎病毒感染的两种主要手段。但是干扰素存在耐受性差、不良反应多、费用昂贵等缺点；而目前已上市的6种核苷类药物(拉米夫定、阿德福韦酯、恩替卡恩、替比夫定、替诺福韦和克拉夫定)均作用于乙型肝炎病毒的逆转录酶。由于现有的治疗手段远远不能满足乙肝治疗的临床需求，研究和开发更多作用于新靶点、新机制及全新结构母核的非核苷类小分子抗乙型肝炎病毒药物日益迫切，是目前药物化学领域研究的热点，且具有十分重要的理论、经济和社会意义。
为此，本发明提供一种结构新颖的、可口服的非核苷类小分子HBV病毒抑制剂。
发明内容
申请号为201810905518.8的中国专利申请中的化合物是一类高效、低毒的新型非核苷类抗HBV小分子候选化合物(该专利申请中全部内容被引入本申请中)。不过，该化合物生物利用度和成药性有待于进一步改进。
为此，本发明提供一种如式I所示的4-吡啶取代酞嗪酮类化合物，该类化合物具有改善的药物理化性质和体内药代动力学性质，口服生物利用度高，成药性好，是一类结构新颖的、可口服的非核苷类小分子HBV病毒抑制剂。
鉴于此，本发明提供一种式I化合物或其药学上可接受的盐：
Figure PCTCN2020097944-appb-000001
其中，
R A选自卤素或氰基；
R B选自卤素；
R C选自H、烷基、环烷基；
L为连接基团，选自亚烷基、亚烷基的任意碳原子被选自-O-、-CO-中至少一种基团置换所得的基团、亚环烷基；
R D选自
Figure PCTCN2020097944-appb-000002
其中，
R 1为H，烷基，芳基，被选自羟基、巯基、烷硫基、氨基烷酰基、羧基、氨基、胍基、脲基、芳基、羟基芳基、杂芳基的基团取代的烷基；
R 2为氢或烷基；
R 3为氢、叔丁氧羰基；
W选自烷基、卤代烷基、环烷基、杂环基、杂链烃基、芳基、杂芳基、烷氧基、氨基；其中，所述烷基、环烷基、杂环基、杂链烃基、芳基、烷氧基、氨基任选各自独立地被1、2、3或4个选自烷基、羧基、烷氧基、杂环基、芳基、杂芳基的基团取代；
X为单键或亚烷基；
R 5，R 6各自独立地选自-OH、芳氧基、烷基的任意碳原子被选自-O-、-CO-、-S-中至少一种基团置换所得的基团、氨基酸残基，此处所述氨基酸残基是指氨基酸的α-氨基除去一个氢所得到的基团，所述氨基酸残基中羧基中的羟基任选被烷氧基、芳氧基、杂芳氧基或苄氧基的基团置换；此处所述氨基酸选自L构型、D构型、或L和D混合构型的下列氨基酸：甘氨酸、脯氨酸、丙氨酸、缬氨酸、正缬氨酸、亮氨酸、异亮氨酸、正亮氨酸、2-叔丁基甘氨酸、2-苯基甘氨酸、苯基丙氨酸、酪氨酸、色氨酸、组氨酸、丝氨酸、高丝氨酸、苏氨酸、半胱氨酸、S-甲基半胱氨酸、高半胱氨酸、蛋氨酸、天冬酰胺、谷氨酰胺、天冬氨酸、谷氨酸、赖氨酸、羟基赖氨酸、鸟氨酸、2,4-二氨基丁酸、2,3-二氨基丙酸、精氨酸、瓜氨酸；或者R 5、R 6与P一起形成杂环基，所述杂环基任选被卤代芳基取代，所述杂环基任选与芳基稠合。
可选地，所述“卤素”选自F、Cl、Br或I；
可选地，所述“烷基”、“卤代烷基”、“烷氧基”、“烷硫基”等任意基团中所具有的烷基各自独立地为C 1-C 20直链或支链烷基，可选地为C 1-C 15直链或支链烷基，可选地为C 1-C 10直链或支链烷基，可选地为C 1-C 7直链或支链烷基，可选地为C 1-C 6直链或支链烷基，可选地为C 1-C 5直链或支链烷基，可选地为C 1-C 4直链或支链烷基，可选地，选自甲基、乙基、正丙基、异丙基、正丁基、异丁基、叔丁基、仲丁基、正戊基，1-甲基丁基、2-甲基丁基、3-甲基丁基、异戊基、1-乙基丙基、新戊基、正己基、1-甲基戊基、2-甲基戊基、3-甲基戊基、异己基、1,1-二甲基丁基、2,2-二甲基丁基、3,3-二甲基丁基、1,2-二甲基丁基、1,3-二甲基丁基、2,3-二甲基丁基、2-乙基丁基、正庚基、2-甲基己基、3-甲基己基、2,2-二甲基戊基、3,3-二甲基戊基、2,3-二甲基戊基、2,4-二甲基戊基、3-乙基戊基或2,2,3-三甲基丁基；可选地，选自甲基、乙基、丙基、异丙基、丁基或异丁基。
可选地，所述“亚烷基”、“亚杂环基烷基”、“氨基烷酰基”等任意基团中所具有的亚烷基各自独立地选自C 1-C 20直链或支链亚烷基，可选地为C 1-C 15直链或支链亚烷基，可选地为C 1-C 10直链或支链亚烷基，可选地为C 1-C 8直链或支链亚烷基，可选地为C 1-C 6直链或支链亚烷基，可选地为C 1-C 4直链或支链亚烷基，可选地为C 2-C 6直链或支链亚烷基，可选地为C 3-C 6直链或支链亚烷基；
可选地，所述“杂链烃基”为链上含1个、2个、3个或4个选自N、O、S的杂原子的直链或支链饱和或不饱和C 1-C 20杂链烃结构，可选地，为链上含1个、2个、3个或4个N的直链或支链饱和或不饱和C 1-C 20杂链烃结构，可选地，为链上含1个、2个、3个或4个N的直链或支链饱和或不饱和C 1-C 10杂链烃结构；
可选地，所述“环烷基”为C 3-C 10单环或双环环烷基，可选地，为C 3-C 7单环环烷基，可选地，为环丙基、环丁基、环戊基、环已基或环庚基；
可选地，所述“亚环烷基”为C 3-C 10单环或双环亚环烷基，可选地，为C 3-C 8单环亚环烷基，可选地，为亚环丙基、亚环丁基、亚环戊基、亚环己基、亚环庚基及亚环辛基。
可选地，所述“亚杂环基”、“杂环基”、“亚杂环基烷基”等任意基团中所具有的杂环为环上含有1个、2个或3个选自N、O、S的杂原子的3-10元非芳香杂环，可选地，所述杂环为环上含有1个或2个选自N、O的杂原子的3-10元非芳香环；所述杂环为环上含有1个或2个选自N、O的杂原子的3-6元非芳香环；可选地，所述杂环为环上含有1个或2个选自N、S的杂原子的3-10元非芳香环；所述杂环为环上含有1个或2个选自N、S的杂原子的3-6元非芳香环；
可选地，所述“芳基”、“羟基芳基”、“卤代芳基”、“芳氧基”等任意基团中所具有 的芳基为6-10元芳基；可选为苯基或萘基，可选为苯基、1-萘基、2-萘基；
可选地，所述“杂芳氧基”、“杂芳基”等、“芳氧基”中任意基团中所含的杂芳基的环为含有1-3个选自N、O和S中的杂原子的5-10元杂芳环；可选地，为含有1-2个选自N、O和S中的杂原子的5-10元杂芳环；可选地，所述杂芳环选自吡啶环、吡咯环、嘧啶环、吡嗪环、哒嗪环、噻吩环、呋喃环；可选地，所述杂芳基为选自吡啶-2-基、吡啶-3-基、吡啶-4-基、哒嗪-3-基、哒嗪-4-基、嘧啶-2-基、嘧啶-4-基、嘧啶-5-基、吡嗪-2-基、吡嗪-3-基、吲哚基、异吲哚基、吲唑基、吲嗪基、嘌呤基、喹嗪基、喹啉基、异喹啉基、噌啉基、酞嗪基、萘啶基、喹唑啉基、喹喔啉基、噻吩并[2,3-b]呋喃基、呋喃并[3,2-b]-吡喃基、吡啶并[2,3-d]噁嗪基、吡唑并[4,3-d]噁唑基、咪唑并[4,5-d]噻唑基、吡嗪并[2,3-d]哒嗪基、咪唑并[2,1-b]噻唑基、咪唑并[1,2-b][l,2,4]三嗪基、苯并噻吩基、苯并噁唑基、苯并咪唑基、苯并噻唑基、苯并噁庚因基、苯并噁嗪基、苯并呋喃基、苯并三唑基、吡咯并[2,3-b]吡啶基、吡咯并[3,2-c]吡啶基、吡咯并[3,2-b]吡啶基、咪唑并[4,5-b]吡啶基、咪唑并[4,5-c]吡啶基、吡唑并[4,3-d]吡啶基、吡唑并[4,3-c]吡啶基、吡唑并[3,4-c]吡啶基、吡唑并[3,4-d]吡啶基、吡唑并[3,4-b]吡啶基、咪唑并[1,2-a]吡啶基、吡唑并[1,5-a]吡啶基、吡咯并[1,2-b]哒嗪基、咪唑并[1,2-c]嘧啶基、吡啶并[3,2-d]嘧啶基、吡啶并[4,3-d]嘧啶基、吡啶并[3,4-d]嘧啶基、吡啶并[2,3-d]嘧啶基、吡啶并[2,3-b]吡嗪基、吡啶并[3,4-b]吡嗪基、嘧啶并[5,4-d]嘧啶基、吡唑并[2,3-b]吡嗪基、嘧啶并[4,5-d]嘧啶基，可选为吡啶-2-基、吡啶-3-基、吡啶-4-基、嘧啶-2-基、嘧啶-4-基或嘧啶-5-基；
除非另有说明，所述“烷基”、“亚烷基”、“亚烷基的任意碳原子被选自-O-、-CO-中至少一种基团置换所得的基团”、“芳基”、“杂芳基”或“苄基”各自独立地被选自下列基团的取代基取代：烷基(如甲基、乙基、正丙基、异丙基、正丁基、异丁基、叔丁基)、烯基(如乙烯基、丙烯基)、炔基(如乙炔基、丙炔基)、羟基、烷氧基(如甲氧基、乙氧基、正丙氧基、异丙氧基、叔丁氧基)、酰氧基(如甲酰氧基、乙酰氧基、丙酰氧基、丁酰氧基、异丙基羰基氧基、叔丁基羰基氧基)、硫基、烷硫基(如甲硫基、乙硫基、正丙硫基、异丙硫基、叔丁硫基)、氨基、烷氨基(如甲氨基、乙氨基、二甲氨基、二乙氨基、正丙氨基、异丙氨基、叔丁氨基)、酰氨基(如甲酰氨基、乙酰氨基、丙酰氨基、丁酰氨基、异丙基羰基氨基、叔丁基羰基氨基)、卤素(如氟、氯、溴、碘)、氰基、硝基、酰基(如甲酰基、乙酰基、羧基、甲氧碳基、乙氧羰基、异丙氧羰基、叔丁氧羰基、氨基羰基、N,N-二甲基氨基羰基)；可选为甲基、乙基、羟基、甲氧基、乙氧基、乙酰氧基、氨基、二甲氨基、乙酰氨基、氟、氯、氰基、乙酰基、甲氧羰基、乙氧羰基、氨基羰基或N,N-二甲基氨基羰基；
可选地，
R A选自氯、氰基；
R B选自氯；
R C选自H、甲基、环丙基；
L选自-CH 2CH 2-、-CH 2CH 2CH 2-、-CH 2CH 2CH 2CH 2-、CH 2CH 2OCH 2CH 2-、
Figure PCTCN2020097944-appb-000003
Figure PCTCN2020097944-appb-000004
R D选自氨基酸残基、
Figure PCTCN2020097944-appb-000005
其中，此处所述氨基酸残基是指氨基酸中的羧基除去羟基所得的基团，所述氨基酸残基为选自L构型、D构型、或L和D混合构型的下述氨基酸的残基：甘氨酸、脯氨酸、丙氨酸、缬氨酸、正缬氨酸、亮氨酸、异亮氨酸、正亮氨酸、2-叔丁基甘氨酸、2-苯基甘氨酸、苯丙氨酸、酪氨酸、苯乙氨酸、色氨酸、组氨酸、丝氨酸、高丝氨酸、苏氨酸、半胱氨酸、S-甲基半胱氨酸、高半胱氨酸、蛋氨酸、天冬酰胺、谷氨酰胺、天冬氨酸、谷氨酸、赖氨酸、羟基赖氨酸、鸟氨酸、2,4-二氨基丁酸、2,3-二氨基丙酸、精氨酸、瓜氨酸；所述氨基酸中α氨基上的氢原子任选被选自叔丁氧羰基的基团置换，所述氨基酸中α碳上的氢原子任选被烷基置换；
W选自C 1-6烷基、羧基C 1-6烷基、C 5-10杂环基C 1-6烷基、C 1-6烷氧基C 6-10芳基、C 1-6烷氧基、一个或两个C 1-6烷基取代的氨基、C 5-10杂环基；
X为单键、-CH 2-、或-CH 2(CH 3)-；
R 5，R 6各自独立地选自-OH、C 6-10芳氧基、C 1-6烷基-O(C＝O)O-C 1-6亚烷氧基、C 1-6烷基(C＝O)O-C 1-6亚烷氧基、C 1-16烷基-O-C 1-6亚烷氧基、氨基酸残基，此处所述氨基酸残基是指氨基酸的α-氨基除去一个氢所得到的基团，所述氨基酸残基中羧基中的羟基任选被C 1-6烷氧基置换；此处所述氨基酸选自L构型、D构型、或L和D混合构型的下列氨基酸：丙氨酸、缬氨酸、正缬氨酸、亮氨酸、异亮氨酸、正亮氨酸、2-叔丁基甘氨酸、2-苯基甘氨酸、苯基丙氨酸、酪氨酸、色氨酸、组氨酸、丝氨酸、高丝氨酸、苏氨酸、半胱氨酸、S-甲基半胱氨酸、高半胱氨酸、蛋氨酸、天冬酰胺、谷氨酰胺、天冬氨酸、谷氨酸、赖氨酸、羟基赖氨酸、鸟氨酸、2,4-二氨基丁酸、2,3-二氨基丙酸、精氨酸、瓜氨酸，优选丙氨酸；
或者R 5、R 6与P一起形成C 5-10杂环基，所述C 5-10杂环基任选被卤代苯基取代，所述 C 5-10杂环基任选与苯基稠合；
可选地，上述式I化合物或其药学上可接受的盐中，其中所述式I化合物选自下列化合物：
Figure PCTCN2020097944-appb-000006
Figure PCTCN2020097944-appb-000007
Figure PCTCN2020097944-appb-000008
Figure PCTCN2020097944-appb-000009
Figure PCTCN2020097944-appb-000010
Figure PCTCN2020097944-appb-000011
Figure PCTCN2020097944-appb-000012
可选地，所述药学上可接受的盐包括式I化合物的阴离子盐和阳离子盐。
可选地，所述药学上可接受的盐包括式I化合物的碱金属的盐、碱土金属的盐、铵盐；可选地，所述碱金属包括钠、钾、锂、铯，所述碱土金属包括镁、钙、锶。
可选地，所述药学上可接受的盐包括式I化合物与有机碱形成的盐；可选地，所述有机碱包括三烷基胺、吡啶、喹啉、哌啶、咪唑、甲基吡啶、二甲氨基吡啶、二甲基苯胺、N-烷基吗啉、1,5-二氮杂双环[4.3.0]壬烯-5、1,8-二氮杂双环[5.4.0]十一碳烯-7、1,4-二氮杂双环[2.2.2]辛烷；可选地，所述三烷基胺包括三甲胺、三乙胺、N-乙基二异丙胺；可选地，所述N-烷基吗啉包括N-甲基吗啉。
可选地，所述药学上可接受的盐包括式I化合物与酸形成的盐；可选地，所述酸包括无机酸、有机酸；可选地，所述无机酸包括盐酸、氢溴酸、氢碘酸、硫酸、硝酸、磷酸、碳酸；可选地，所述有机酸包括甲酸、乙酸、丙酸、草酸、丙二酸、琥珀酸、富马酸、马来酸、乳 酸、苹果酸、柠檬酸、枸橼酸、酒石酸、碳酸、苦味酸、甲磺酸、乙磺酸、对甲苯磺酸、谷氨酸、双羟萘酸。
根据本发明的另一方面，还提供了上述式I化合物或其药学上可接受的盐的制备方法，该方法可通过以下反应路线1实现：
Figure PCTCN2020097944-appb-000013
该反应路线包括：将Hu1化合物与R DOH进行缩合成酯反应，得到式I化合物，其中，式Hu1和式I中，R A、R B、L的定义如权利要求1-5任一项所述，R C为H；式I中，R D定义如权利要求1-5任一项所述；
或者，该方法通过以下反应路线2实现：
Figure PCTCN2020097944-appb-000014
该反应路线包括：将Hu1-1化合物与R DOH进行缩合成酯反应，得到式Hu1-2化合物，其Hu1-2化合物中的R D为权利要求1-5任一项所定义的含有保护基的基团，然后将式Hu1-2化合物与卤化物反应引入R C，任选地脱去R D中的保护基团，得到式I化合物，其中，式Hu1-1、式Hu1-2和式I中，R A、R B、L的定义如权利要求1-5任一项所述；式I中，R D的定义如权利要求1-5任一项所述；式I中，R C为权利要求1-5任一项所定义的不为氢的基团；
任选地，上述反应路线1和反应路线2中，采用保护基团对式Hu1化合物、式Hu1-1、式R DOH化合物进行保护，进行缩合成酯反应后，任选地进行脱保护；
这些保护基的脱去根据常规方法，优选通过在惰性溶剂例如1,4-二氧六环、二氯甲烷中与强酸例如盐酸、氢溴酸或三氟乙酸进行反应实施。
R DOH为式HOOCC(R 1)(R 2)NHR 3、式WCOOH、式PG-(P＝O)R 5R 6化合物，其中R 1、R 2、R 3、W定义如上所述；PG为保护基团，PG优选为五氟苯氧基；R 5，R 6各自独立地选自-OH、芳氧基、氨基酸残基，此处所述氨基酸残基是指氨基酸的α-氨基除去一个氢所得到的基团，所述氨基酸残基中羧基中的羟基任选被烷氧基、芳氧基、杂芳氧基或苄氧基的基团置换；此处所述氨基酸选自L构型、D构型、或L和D混合构型的下列氨基酸：甘氨酸、脯氨酸、丙氨酸、缬氨酸、正缬氨酸、亮氨酸、异亮氨酸、正亮氨酸、2-叔丁基甘氨酸、2-苯基甘氨酸、苯基丙氨酸、酪氨酸、色氨酸、组氨酸、丝氨酸、高丝氨酸、苏氨酸、半胱氨酸、S-甲基半胱氨酸、高半胱氨酸、蛋氨酸、天冬酰胺、谷氨酰胺、天冬氨酸、谷氨酸、赖氨酸、羟基赖氨酸、鸟氨酸、2,4-二氨基丁酸、2,3-二氨基丙酸、精氨酸、瓜氨酸；或者R 5、R 6与P一起形成杂环基，所述杂环基任选被卤代芳基取代，所述杂环基任选与芳基稠合。
可选地，上述制备方法中的所述缩合成酯反应中，合适的缩合剂选自碳二亚胺例如N,N’-二环己基碳二亚胺(DCC)或N-(3-二甲基氨基异丙基)-N’-乙基碳二亚胺盐酸盐(EDC)。
缩合成酯反应优选在碱的存在下进行。作为碱，优选使用碱金属碳酸盐，例如碳酸钠或碳酸钾，或有机碱例如三乙胺，N-甲基吗啉，N-甲基哌啶，N,N-二异丙基乙胺或4-N,N-二甲基氨基吡啶。
可选地，所述缩合成酯反应的温度为0℃至+60℃，优选+20℃至+40℃。所述反应优选在常压进行。
或者，上述式I化合物或其药学上可接受的盐还可以采用以下方法制备：
将式Hu1化合物与式Hu2化合物反应，得到式Hu3化合物，再引入R 5和R 6基团，得到式I化合物。
Figure PCTCN2020097944-appb-000015
Figure PCTCN2020097944-appb-000016
式I中，R A、R B、R C、L如上所述；R D为
Figure PCTCN2020097944-appb-000017
其中，X如上所述，R 5，R 6各自独立地选自-OH、芳氧基、烷基的任意碳原子被选自-O-、-CO-、-S-中至少一种基团置换所得的基团、氨基酸残基，此处所述氨基酸残基是指氨基酸的α-氨基除去一个氢所得到的基团，所述氨基酸残基中羧基中的羟基任选被烷氧基、芳氧基、杂芳氧基或苄氧基的基团置换；此处所述氨基酸选自L构型、D构型、或L和D混合构型的下列氨基酸：甘氨酸、脯氨酸、丙氨酸、缬氨酸、正缬氨酸、亮氨酸、异亮氨酸、正亮氨酸、2-叔丁基甘氨酸、2-苯基甘氨酸、苯基丙氨酸、酪氨酸、色氨酸、组氨酸、丝氨酸、高丝氨酸、苏氨酸、半胱氨酸、S-甲基半胱氨酸、高半胱氨酸、蛋氨酸、天冬酰胺、谷氨酰胺、天冬氨酸、谷氨酸、赖氨酸、羟基赖氨酸、鸟氨酸、2,4-二氨基丁酸、2,3-二氨基丙酸、精氨酸、瓜氨酸；
式Hu2中，X如上所述，Q 1、Q 2各自独立地为H、烷基；
式Hu3中，R A、R B、R C、L、X如上所述；
R 5和R 6基团的引入方法包括，将式Hu3化合物与相应的氨基酸进行磷酰胺化反应制得相应的磷酰胺式I化合物，或者与相应的卤代物或醇进行反应制得磷酸酯式I化合物。
Hu3化合物与相应的氨基酸进行胺酸缩合成磷酰胺反应，合适的缩合剂选自碳二亚胺例如N,N’-二环己基碳二亚胺(DCC)或N-(3-二甲基氨基异丙基)-N’-乙基碳二亚胺盐酸盐(EDC)。缩合成酰胺反应优选在碱的存在下进行。作为碱，优选使用碱金属碳酸盐，例如碳酸钠或碳酸钾，或有机碱例如三乙胺，N-甲基吗啉，N-甲基哌啶，N,N-二异丙基乙胺或4-N,N-二甲基氨基吡啶。
Hu3化合物与相应的醇进行醇酸缩合成磷酸酯反应，合适的缩合剂选自碳二亚胺例如N,N’-二环己基碳二亚胺(DCC)或N-(3-二甲基氨基异丙基)-N’-乙基碳二亚胺盐酸盐(EDC)。缩合成磷酸酯反应优选在碱的存在下进行。作为碱，优选使用碱金属碳酸盐，例如碳酸钠或碳酸钾，或有机碱例如三乙胺，N-甲基吗啉，N-甲基哌啶，N,N-二异丙基乙胺或4-N,N-二甲 基氨基吡啶。
Hu3化合物与相应的氯代物在碱性条件下进行亲核取代成磷酸酯反应，作为碱，优选氢化钠，氢化钾。
关于保护基团：当多官能团有机化合物进行反应时，为使反应只发生在所希望的官能团处，而避免其他官能团遭受影响，在反应前将其他官能团先加以保护，当反应完成后再恢复。保护基团是这种能够对所述其他官能团提供保护的一类基团的总称。所述氨基的保护基团可以为叔丁氧基碳基(Boc)。羟基或羧基官能团的保护基优选使用叔丁基、乙基、甲基。这些保护基的脱去根据常规方法，优选通过在惰性溶剂例如1,4-二氧六环、二氯甲烷中与强酸例如盐酸、氢溴酸或三氟乙酸进行反应实施。
式Hu1化合物可以按照文献常规方法制备(例如可参照申请号为201810905518.8的专利申请中的制备方法制备)。
可选地，上述式I化合物的药学上可接受的盐，可以通过将式I化合物溶于相应的酸饱和的醇溶液中进行反应而制备，例如：将式I所示的化合物溶于氯化氢饱和的甲醇溶液，室温搅拌30分钟，将溶剂蒸干，即制得相应的式I化合物的盐酸盐。
另一发面，还提供一种药物组合物，其包含治疗有效量的上述式I化合物或其药学上可接受的盐中的一种或多种以及任选存在的药学上可接受的载体。
上述药学上可接受的载体是指药学领域常规的药物载体，例如：稀释剂，如水等；填充剂，如淀粉、蔗糖等；粘合剂，如纤维素衍生物、藻酸盐、明胶、聚乙烯吡咯烷酮；湿润剂，如甘油；崩解剂，如琼脂、碳酸钙和碳酸氢钠；吸收促进剂，如季铵化合物；表面活性剂，如十六烷醇；吸附载体，如高岭土和皂粘土；润滑剂，如滑石粉、硬脂酸钙和硬脂酸镁、和聚乙二醇等。另外，还可以在上述药物组合物中加入其它辅剂，如香味剂和甜味剂等。
上述的式I化合物或其药学上可接受的盐，或药物组合物可以通过口服、直肠或肠外给药的方式施用于需要这种治疗的患者。用于口服时，可以将其制成常规的固体制剂，如片剂、粉剂、粒剂、胶囊等，或制成液体制剂，如水或油悬浮剂，或其它液体制剂，如糖浆等；用于肠外给药时，可将其制成注射用的溶液、水或油性悬浮剂等。
本发明还提供上述的式I化合物或其药学上可接受的盐，或者上述的药物组合物在制备预防和/或治疗乙型肝炎疾病的药物中的用途。
本发明还提供上述的式I化合物或其药学上可接受的盐，或者上述的药物组合物在制备乙型肝炎病毒抑制剂中的用途。
本发明还提供上述的式I化合物或其药学上可接受的盐，或者上述的药物组合物在制备 抑制病毒复制的药物中的用途，所述病毒包括甲肝病毒、乙肝病毒、丙肝病毒、流感病毒、腺病毒、艾滋病毒、疱疹病毒、人乳头瘤病毒。
本发明还提供一种预防和/或治疗乙型肝炎疾病的方法，包括给需要的患者施用有效量的上述式I化合物或其药学上可接受的盐，或者上述的药物组合物。
本发明还提供一种抑制乙型肝炎病毒的方法，包括给需要的患者施用有效量的上述式I化合物或其药学上可接受的盐，或者上述的药物组合物。
本发明还提供一种抑制病毒复制的方法，包括给宿主施用有效量的上述式I化合物或其药学上可接受的盐，或者上述的药物组合物。所述病毒包括甲肝病毒、乙肝病毒、丙肝病毒、流感病毒、腺病毒、艾滋病毒、疱疹病毒、人乳头瘤病毒。
根据一方面的具体实施方式，化合物在细胞水平上具有高效的抑制HBV DNA复制的活性，对HepG2.2.15细胞的生长毒性较小。
根据一方面的具体实施方式，化合物具有较好的肝脏靶向性，能在肝脏稳定存在并富集，是有效的乙型肝炎病毒抑制剂。
附图说明
图1显示的是测试实施例3中AAV小鼠外周血中病毒HBV DNA含量随时间变化示意图。
图2显示的是测试实施例3中AAV小鼠外周血中病毒HBsAg含量随时间变化示意图。
图3显示的是测试实施例3中AAV小鼠外周血中病毒HBeAg含量随时间变化示意图。
图4显示的是测试实施例3中给药4周后AAV小鼠肝脏中病毒HBcAg含量示意图；图中，核心蛋白阳性的细胞呈现深黑色，阴性细胞呈现浅黑色。
具体实施方式
以下对本发明的具体实施方式进行详细的说明。应当理解的是，此处所描述的具体实施方式仅用于示例性地对本发明进行说明，并不用于限制本发明。
在以下实施例中，核磁共振氢谱用BrukerAMX-400型、Gemini-300型或AMX–600型核磁共振仪记录，化学位移δ的单位为ppm。比旋光度由Perkin-Elmer241型自动旋光仪测定，所用微波为CEM-discovery微波反应器。如无特别说明，所有反应溶剂均按照常规方法进行纯化。柱层析用硅胶(200-300目)为青岛海洋化工分厂生产。薄层层析使用GF254高效板，为烟台化工研究所生产。制备型薄层层析板由中国科学院上海药物研究所制备，固定相 采用GF254(HG/T2354-92)硅胶和羧甲基纤维素钠(800-1200)制备，分别为青岛海洋化工有限公司和中国医药(集团)上海化学试剂公司生产。如无特别标注，所有溶剂均为分析纯试剂，所用试剂均购自国药集团化学试剂有限公司。采用碘、紫外荧光等方法显色。减压蒸除有机溶剂在旋转蒸发仪中进行。
实施例1：化合物1的合成路线如下：
Figure PCTCN2020097944-appb-000018
步骤(i)：
将3,6-二氯酞嗪i-1(10g,50.3mmol)加入到醋酸(150mL)中，120℃回流6小时，TLC监测反应完全后，停止反应并将反应液冷却至室温，减压旋蒸除去醋酸得到白色固体化合物i-2。
步骤(ii)：
将化合物i-2(1.0g,5.5mmol)溶于DMF(70mL)中，依次加入4-氰基苄氯(1.26g,6.6mmol)和Cs 2CO 3(2.1g,6.6mmol)。反应体系在50℃下搅拌5小时后，TLC监测反应完全。向反应溶液中加入乙酸乙酯(100mL)和水(100mL)，分出有机层，有机层依次用水(100mLx4)和饱和食盐水(100mL)洗涤。有机相用无水硫酸钠干燥半个小时后过滤浓缩得粗产物，该粗产物用柱层析(PE:EA＝100:0～10:1)分离纯化得白色固体化合物i-3。
步骤(iii)：
将化合物i-3(97mg,0.33mmol)溶于1,4-二氧六环(20mL)和水(5mL)的混合溶剂中，依次加入2,6-二氟-吡啶-3-硼酸(68.3mg,0.43mmol)、Pd(PPh 3) 4(76.0mg,0.033mmol)和K 3PO 4(139mg,0.66mmol)。N 2置换3次后100℃反应过夜。次日TLC监测反应完全，将反应体系冷却至室温。减压旋蒸除去溶剂后，将残余物用乙酸乙酯萃取(30mL)溶解，有机层依次用水(30mLx3)和饱和食盐水(30mL)洗涤，无水硫酸钠干燥、过滤、浓缩得粗产物，该粗产物用柱层析(PE:EA＝67:33)分离纯化得化合物i-4。
Figure PCTCN2020097944-appb-000019
步骤(iv)：
将化合物i-4(150mg,0.4mmol)溶于无水乙醇(10mL)中，加入3-氨基-1-丙醇(4mmol)，回流反应过夜。次日旋转蒸干溶剂，残余物溶于乙酸乙酯(50mL)和水(50mL)，有机层用饱和食盐水(30mL)洗涤，无水硫酸钠干燥、过滤、浓缩得粗产物，该粗产物用柱层析(PE:EA＝50:50)分离纯化得白色固体化合物i-5。
步骤(v)：
将化合物i-5(0.19mmol)溶于无水乙腈(10mL)中，加入N-氯代丁二酰亚胺(25mg,0.19mmol)，回流5小时。旋干溶剂得粗产物，粗产物用柱层析(PE:EA＝60:40)分离纯化得白色固体化合物1(收率81％)。 1H NMR(400MHz,Chloroform-d)δ8.53–8.46(m,1H),7.85–7.78(m,2H),7.68–7.51(m,6H),5.87(t,J＝5.9Hz,1H),5.48(s,2H),3.80(t,J＝5.6Hz,2H),3.69(q,J＝6.1Hz,2H),1.96–1.85(m,2H).
化合物I-1的合成路线如下所示：
Figure PCTCN2020097944-appb-000020
步骤a：
Boc-L-缬氨酸(1.85g,8.5mmol)、二环已基碳二亚胺(1.4g,6.8mmol)以及4-二甲氨基吡啶(0.34g,2.8mmol)，分别加到化合物1(2.63g 5.67mmol)的二氯甲烷(50ml)溶液中，加完后继续室温反应1小时。反应结束后，抽滤，减压蒸去滤液中的二氯甲烷，残余物用乙酸乙酯溶解，依次用水、饱和食盐水洗涤，收集乙酸乙酯层，无水硫酸钠干燥，滤液蒸干，得粗产物， 粗品用硅胶柱层析(PE/EA 75/25)纯化，得白色固体I-1(3.4g,92％)。
步骤b：
将I-1(3.4g 5.6mmol)溶于25mL二氯甲烷中，加入5mL三氟乙酸，油浴温度40℃反应3h,加入饱和碳酸氢钠溶液调节pH值至中性，二氯甲烷萃取3次，无水硫酸钠干燥，过滤，旋干溶剂，粗品经硅胶柱层析(二氯甲烷/甲醇94/6)得白色固体I-2(2.7g,94％)。
Figure PCTCN2020097944-appb-000021
Figure PCTCN2020097944-appb-000022
Figure PCTCN2020097944-appb-000023
Figure PCTCN2020097944-appb-000024
Figure PCTCN2020097944-appb-000025
Figure PCTCN2020097944-appb-000026
Figure PCTCN2020097944-appb-000027
Figure PCTCN2020097944-appb-000028
Figure PCTCN2020097944-appb-000029
Figure PCTCN2020097944-appb-000030
Figure PCTCN2020097944-appb-000031
Figure PCTCN2020097944-appb-000032
Figure PCTCN2020097944-appb-000033
实施例2：
Figure PCTCN2020097944-appb-000034
步骤a:
化合物I-1(85mg,0.13mmol)溶于干燥的四氢呋喃(5mL)中,0℃下，滴加到60％NaH(25mg,64mmol)中，搅拌30分钟。加入碘甲烷(9.58uL，0.15mmol)的四氢呋喃(2mL)溶液，加毕，反应液逐渐升至室温搅拌过夜，反应液倒入冰水中，乙酸乙酯(20mL×3)萃取，有机相无水硫酸钠干燥，过滤，减压蒸除溶剂，残余物经硅胶柱层析，得白色粉末状固体化合物I-33’(47mg,55％)。
步骤b:
将I-33’(40mg 0.059mmol)溶于5mL二氯甲烷中，加入0.5mL三氟乙酸，油浴温度40℃反应3h,加入饱和碳酸氢钠溶液调节pH值至中性，二氯甲烷萃取3次，无水硫酸钠干燥，过滤，旋干溶剂，粗品经硅胶柱层析(二氯甲烷/甲醇95/5)得白色固体I-34(32mg,94％)。
Figure PCTCN2020097944-appb-000035
Figure PCTCN2020097944-appb-000036
实施例3：
Figure PCTCN2020097944-appb-000037
步骤:将化合物1(300mg,0.65mmol)溶解于四氢呋喃中，氮气保护、-5℃条件下，依次加入叔丁基溴化镁的四氢呋喃(1.35mmol,1.35mL,c 1mol/L)溶液，N-[(S)-(2,3,4,5,6-五氟苯氧基)苯氧基磷酰基]-L-丙氨酸异丙酯B(351mg,0.775mmol)的四氢呋喃溶液，反应液逐步升 温至5℃，反应17h。TLC监测无原料，加水淬灭反应，旋干溶剂后，加入100mL水,乙酸乙酯萃取3次(50mL*3)，饱和食盐水洗涤，无水硫酸钠干燥，过滤，旋干溶剂得粗产物。该粗产物经硅胶柱层析纯化(石油醚/乙酸乙酯90/10-50/50)得白色固体化合物I-36(280mg,52％)。
Figure PCTCN2020097944-appb-000038
实施例4：
Figure PCTCN2020097944-appb-000039
步骤a:
将化合物1(4.64g,10mmol)溶于50mLDMF中,搅拌下分批加入80％NaH(0.64g,26.7mmol)，室温搅拌15分钟。冰盐浴(-10℃)下滴加对甲苯磺酰氧甲基膦酸二乙酯(0.45g,14mmol)，加毕，保持温度在-10～0℃之间搅拌1h，后升至室温搅拌4～5h。滴加冰醋酸调pH至中性(约1mL)，过滤，滤饼用二氯甲烷洗涤,合并滤液，有机相用水(30mL×3)洗涤，水相再用二氯甲烷(20mL×3)萃取，合并二氯甲烷相，无水硫酸钠干燥，过滤，减压蒸除溶剂，残余物中加入甲苯，冷冻过夜。过滤,滤饼于50℃减压干燥,得白色粉末状固体化合物2(2.88g,47％)。
步骤b:
在100mL反应瓶中，将化合物2(2.5g,4.07mmol)溶于20mL氢溴酸的水溶液中，升温水解，充分反应后冷却，加入二氯甲烷萃取,有机层干燥，过滤，浓缩，粗品经硅胶柱层析(二氯甲烷/甲醇85/15)分离得到白色固体化合物3(1.68g,74％)。
步骤c:
100mL反应瓶中，分别投入化合物3(557mg,1mmol)、氯甲基碳酸异丙酯(760mg,5mmol)、N-甲基吡咯烷酮(880mg,8.8mmol)和三乙胺(310mg,3mmol)，室温搅拌12h，然后升温至50～60℃，搅拌4h。冷却至室温，加入乙酸异丙酯250mL，室温搅拌1～2h。过滤，滤饼用20mL乙酸异丙酯洗涤，滤液用饱和氯化钠水溶液(100mL×3)洗涤，有机层用无水硫酸钠干燥，过滤，滤液减压浓缩得油状物，油状物加入丙酮10mL，搅拌10min，充分搅拌均匀后，加入无水乙醚20mL析晶，此时溶液变浑浊，继续搅拌7～8h。过滤，无水乙醚洗涤，晾干得到 白色固体粉末,即为目标化合物I-38(395mg,收率50％)。
Figure PCTCN2020097944-appb-000040
实施例5：
Figure PCTCN2020097944-appb-000041
步骤a：
将化合物3(0.5g,0.9mmol)、苯酚(84mg,0.9mmol)、二环已基碳二亚胺(0.22g,1.07mmol) 以及三乙胺(0.181g,1.8mmol)，分别加到N-甲基吡咯烷酮(20mL)溶液中，100℃下反应24小时。反应结束后，加入大量水，二氯甲烷萃取3次，有机相用水洗，无水硫酸钠干燥，抽滤，减压蒸去二氯甲烷，得残余物280mg，直接下一步反应。
步骤b：
残余物(280mg,0.44mmol)溶于20mL乙腈中，加入0.5mL二氯亚砜，75℃反应2小时，将反应液浓缩至干，残留物溶于干燥20mL二氯甲烷中，冷却至-78℃，加入L-丙氨酸异丙酯盐酸盐(61mg,0.44mmol))，三乙胺(0.181g,1.8mmol)，反应1.5小时，加入水，二氯甲烷萃取3次，有机相用水洗，收集二氯甲烷层，无水硫酸钠干燥，抽滤，减压蒸去二氯甲烷得粗品，粗品用硅胶柱层析(二氯甲烷/甲醇85/15)纯化，得白色固体化合物I-40(0.13g,41％)。
Figure PCTCN2020097944-appb-000042
实施例6：
Figure PCTCN2020097944-appb-000043
步骤a：
将化合物3(0.5g,0.9mmol)、草酰氯(0.2mL)、1滴N,N-二甲基甲酰分别加到二氯甲烷(20mL)溶液中，回流反应3小时。将反应液浓缩至干，残留物溶于干燥二氯甲烷(20ml)中，加入L-丙氨酸乙酯盐酸盐(0.344g,2.24mmol))，二异丙基乙基胺(0.46g,3.58mmol)，反应1.5小时，加入水，二氯甲烷萃取3次，有机相用水洗，收集二氯甲烷层，无水硫酸钠干燥，抽滤，减压蒸去滤液中的二氯甲烷得粗品，粗品用硅胶柱层析(二氯甲烷/甲醇90/10)纯化，得白色固体化合物I-41(0.31g,45％)。
Figure PCTCN2020097944-appb-000044
实施例7：
Figure PCTCN2020097944-appb-000045
步骤a:
40℃下，将三氯氧膦(2.3g,15mmol)加入到1(0.464g，1mmol)的磷酸三乙酯(10mL)溶液中，搅拌1h。加入三乙胺/碳酸氢铵缓冲溶液(1M,pH 7)，减压浓缩至干，残留物加入二氯甲烷和水，水相用二氯甲烷萃取3次，合并有机相，干燥，过滤，除去溶剂得粗品，粗品 溶于少量二氯甲烷，加入大量冷乙醚，析出固体，过滤，乙醚洗涤，得产物化合物4(0.45g83％)。
步骤b:
L-丙氨酸异丙酯(0.106g,0.81mmol)、二环已基碳二亚胺(0.17g,0.81mmol)分别加到化合物4(0.4g 0.735mmol)的叔丁醇-水(1/1 50mL)溶液中，加完后回流反应1小时。反应结束后，减压蒸去溶剂，残余物用二氯甲烷溶解，然后依次用水、饱和食盐水洗，收集二氯甲烷层，无水硫酸钠干燥，滤液蒸干，得粗产物，粗品用硅胶柱层析(二氯甲烷/甲醇85/15)纯化，得白色固体化合物I-42(0.29g,60％)。
Figure PCTCN2020097944-appb-000046
实施例8：
Figure PCTCN2020097944-appb-000047
将化合物4(0.5g,0.92mmol)、2-(十六烷氧基)乙醇(0.263g,0.92mmol)、二环已基碳二亚胺(0.22g,1.07mmol)以及三乙基胺(0.181g,1.8mmol)，分别加到二氯甲烷(20mL)中，室温反应24小时。反应结束后，过滤，滤液加入大量水，二氯甲烷萃取3次，有机相用水洗，收集二氯甲烷层，干燥，抽滤，减压蒸去滤液中的二氯甲烷，得粗产物，粗品用硅胶柱层析 (二氯甲烷/甲醇85/15)纯化，得白色固体化合物I-43(0.28g,50％)。
Figure PCTCN2020097944-appb-000048
实施例9：
Figure PCTCN2020097944-appb-000049
步骤a：
-20℃，氮气保护下，将新蒸的三氯化膦PCl 3(0.85mL,1.3g,9.5mmol)滴加到邻羟基苄醇(1.01g,8.10mmol)的乙醚(30mL)溶液中，10分钟后,反应体系中滴加三乙胺(1.89g,2.60mL,18.7mmol)的乙醚溶液(约2-3h)，加毕，升温至室温，继续搅拌1h，将反应体系置于-20℃过夜，过滤除去三乙胺的盐酸盐，滤液浓缩得黄色油状物化合物5(1.30g)，该粗品化合物5直接用于下一步反应。
步骤b：
氮气保护下，二异丙胺(866mg,1.20mL,8.56mmol)的乙醚(4mL)溶液滴加到化合物5的乙醚溶液(9mL)中，室温搅拌3小时，过滤，滤液减压浓缩得粗品化合物6(1.04g)，直接用于下一步反应。
步骤c：
0℃，氮气保护下，将化合物1(172mg,0.37mmol))、吡啶盐酸盐(179mg,1.51mmol)溶于乙腈(10mL)中,将粗品化合物6(241mg)的乙腈(5mL)溶液加入到上述溶液，反应液在0℃下搅拌3小时。体系中加入叔丁基过氧化氢的正癸烷溶液(0.23mL,1.3mmol，C 5.5M)，升至室温继续反应1小时，减压蒸除溶剂，残余物经硅胶柱层析(二氯甲烷/甲醇95/5)纯化，得无色泡沫化合物I-44(0.12g,51％)。
Figure PCTCN2020097944-appb-000050
Figure PCTCN2020097944-appb-000051
实施例10：
Figure PCTCN2020097944-appb-000052
准确称取I-2(300mg)于茄形瓶中，加入30mL乙腈溶剂并在60℃条件下不断搅拌，而后在60℃条件下缓慢加入1.2倍当量的盐酸溶液(约55ul浓盐酸溶用10mL乙腈稀释)。将反应相在60℃条件下混悬1小时后，冷却至室温，静置12小时。将析出的固体过滤，用少量乙腈洗涤3次。之后将获得的固体在真空干燥箱干燥得到I-46，浅黄色固体310毫克(97％)。
生物活性检测实施例
测试实施例1：实施例中制得的化合物对乙型肝炎病毒DNA复制能力的影响
1、实验材料
1.1筛选体系
稳定转染全长HBV的人肝癌细胞HepG2.2.15细胞株(中科院上海药物研究所提供)
1.2实验仪器
培养箱(ThermoForma3111)；酶标仪(Molecular Devices Spectra Max 190)；电子天平；显微镜；生物安全柜(Heal Force safe15)；离心机(Eppendorf Centrifuge 5810R)；Real-Time PCR(FASTA GEN-DNA fast2000)
1.3实验药物和试剂
阳性药及配置：拉米夫定(3TC)，由中科院上海药物研究所药物化学组合成，用DMEM/High Glucose培养液(Dulbecco’s modified Eagle’s medium，Hyclone公司)配置成40mM储液待用。
其他溶液及配置：
DMEM/High Glucose培养液：Dulbecco’s modified Eagle’s medium 1×(Hyclone公司)磷酸盐缓冲液(PBS，pH7.3，1L)：NaCl，8.0g；Na 2HPO 4，1.16g；KH 2PO 4，0.2g；KCl：0.2g；
MTT溶液：MTT(Sigma公司)，用PBS配制成5mg/ml；
DNA抽提试剂盒：
Figure PCTCN2020097944-appb-000053
Blood & Tissue(Qiagen公司)
2、实验方法
2.1细胞培养
HepG2.2.15细胞按常规方法培养传代。利用的培养基为DMEM，内含有10％(v/v)牛血清及选择抗生素G418，在37℃，5％CO 2的培养箱中培养8天(第4天换液)。
2.2待测化合物及阳性药的配制
待测化合物用DMSO配置成40mM的储备液，含10％Hyclone TM Fetal Bovine Serum的DMEM培养液配成指定最高浓度的溶液并稀释；阳性药为拉米夫定，同样以含10％Hyclone TM Fetal Bovine Serum的DMEM培养液配成指定浓度。
2.3 MTT测定细胞毒性
HepG2.2.15细胞以5×10 3细胞/孔接种于96孔板，按上述方法在药物作用下培养八天，取出上清200μl后加入MTT溶液，培养4h后加入裂解液，培养12h后用酶标仪测定OD 570，与对照孔的吸光度进行比较，计算存活细胞百分比，并计算出致半数细胞毒性所需浓度CC 50。
2.4细胞培养上清中HBV DNA含量的测定
HepG2.2.15细胞经不同浓度的化合物作用8天(第4天换液)后，吸取培养上清，采用实时PCR(Real-time PCR)法定量检测上清中成熟病毒粒子内含有的HBV DNA。
柱提HepG2.2.15细胞上清DNA(Qiagen，
Figure PCTCN2020097944-appb-000054
Blood & Tissue Handbook)
1)收96孔板上清DNA，200μl每孔，将复孔均收集至同一个EP管中，4000rcf*5min离心，取上清；
2)取200μl上清，加入1.5ml EP管中，加入20μl蛋白酶K和200μl Buffer AL(Qiagen，
Figure PCTCN2020097944-appb-000055
Blood & Tissue Kit)，涡旋使其完全混匀，56℃孵育10min；
3)加入200μl乙醇，涡旋使其完全混匀；
4)将第3)步的液体全部加入置于2ml废液收集管中的DNeasy Mini spin column(Qiagen，
Figure PCTCN2020097944-appb-000056
Blood & Tissue Kit)，6000rcf*1min离心，弃去上清；
5)将DNeasy Mini spin column置于新的2ml废液收集管，加入500μl Buffer AW1(Qiagen，
Figure PCTCN2020097944-appb-000057
Blood & Tissue Kit)，6000rcf*1min离心，弃去上清；
6)将DNeasy Mini spin column置于新的2ml废液收集管，加入500μl Buffer AW2，(Qiagen，
Figure PCTCN2020097944-appb-000058
Blood & Tissue Kit)20000rcf*3min离心，弃去上清；
7)将DNeasy Mini spin column置于新的1.5ml EP管内，吸取50μl Buffer AE(Qiagen，
Figure PCTCN2020097944-appb-000059
Blood & Tissue Kit)直接加入DNeasy Mini spin column的膜上，在室温放置5min， 6000rcf*1min离心洗脱膜上的DNA，弃去DNeasy Mini spin column，收集DNA样品至-20℃。
实时PCR检测上清HBV DNA(乙型肝炎病毒核酸定量检测试剂盒，达安基因)
1)标准曲线：1e 7-1e 4IU/ml，1μl上样，设置一个空白孔，以检测反应体系是否污染；
2)按顺序加DNA样品1μl；
3)加入酶及反应缓冲液：先将两管反应液加入酶中，混匀之后稍微离心置于冰上，在上样后加入酶反应液19μl，加入时确保不触碰到DNA样品造成污染；
4)贴上封膜，离心；
5)PCR反应：
第一阶段:93℃，2min
第二阶段:10个循环
第一步：93℃，45s
第二步：55℃，1min
第三阶段：45个循环
第一步：93℃，30s
第二步：55℃，45s
样品：20μl
检测：在完成第三阶段的第二步(55℃，45s)后收集数据。
2.8数据处理
采用Origin软件对实验数据进行统计，计算IC 50
实验结果：实验结果如表1所示。
表1：实施例的式I化合物对HepG2.2.15细胞的毒性和抑制HBV DNA的活性
化合物 CC 50(μM) IC 50(μM) 化合物 CC 50(μM) IC 50(μM)
I-1 >100 0.0091 I-24 67.25 0.44
I-2 63.5 0.0057 I-25 71.88 0.26
I-3 >100 0.049 I-26 >100 1.22
I-4 24 0.029 I-27 >100 0.11
I-5 >100 0.68 I-28 >100 1.2
I-6 87.87 0.12 I-29 >100 0.33
I-7 >100 0.45 I-30 >100 0.0081
I-8 >100 0.8 I-31 >100 0.20
I-9 >100 2.1 I-32 63.5 0.061
I-10 28.02 0.27 I-33 94.4 0.3
I-11 >100 0.88 I-34 >100 0.015
I-12 >100 0.43 I-35 >100 0.042
I-13 87.87 0.014 I-36 24.4 0.035
I-14 >100 3.71 I-37 >100 0.8
I-15 >100 0.32 I-38 65.21 1.69
I-16 71.88 0.2 I-39 70.36 0.34
I-17 82.98 0.29 I-40 >100 0.34
I-18 71.88 0.19 I-41 71.88 0.10
I-19 82.84 <0.098 I-42 >100 <0.098
I-20 >100 0.068 I-43 70.36 0.8
I-21 >100 0.061 I-44 67.25 1.7
I-22 >100 0.21 I-45 >100 0.080
I-23 >100 0.46      
注：CC 50为实施例的化合物对HepG2.2.15细胞的生长的影响，半数(50％)致死浓度。IC 50为实施例的化合物对乙肝病毒DNA复制的抑制达半数(50％)时的浓度。
从测试结果可以看出，实施例的化合物的IC 50均小于10μM，在细胞水平上都具有高的抑制HBV DNA复制的活性，且对HepG2.2.15细胞的生长毒性较小。
测试实施例2：药代动力学研究
取ICR(CD-1)雄性小鼠(体重：～20g)，每组每个时间点动物数为3，分别灌胃给药20mg/kg剂量给予化合物I-2、化合物I-36和对比化合物1，采血时间点安排为口服组0min、15min、30min、1h、2h、4h、8h及24h。采血后离心制备血浆，并按20μL分装，于–70℃保存直至分析。药动学参数用Kinetica 5.0软件(美国)非房室模型分析处理。
通过LC-MS分别测定化合物1和化合物1的活性代谢产物1-M在血浆中的浓度，结果汇总在表2中。
Figure PCTCN2020097944-appb-000060
表2：口服实施例化合物I-2、实施例化合物I-36和对比化合物1小鼠药代动力学参数(给药方式：口服20mg/kg)
Figure PCTCN2020097944-appb-000061
备注：表中“-”表示未测量到。
表2中化合物1的PK数据表明，在口服化合物1之后的血浆中检测到非常有限的药物相关化合物(包括化合物1和活性代谢产物1-M)。式I-2化合物、式I-36化合物可以在口服后将药物有效地传递到血浆中，所述药物包括化合物1和活性代谢化合物1-M。表2的PK数据表明，本发明式I所示化合物是可以适用于口服的非核苷类小分子HBV病毒抑制药物。
测试实施例3：
取6～8周的Balb/c雄性小鼠(体重：～20g)，将重组病毒AAV8-HBV病毒液以1×10 11vg/只(Viral genome,vg)的剂量尾静脉注射，构建野生型HBV慢性感染小鼠模型。在感染4周后，将血清中的病毒含量持续稳定的感染小鼠随机分为三组，每组六只，分别为模型组(0.5％CMC-Na溶剂对照组)，化合物I-2 100mg/kg剂量组和恩替卡韦(ETV)1mg/kg剂量组(阳性药组)。化合物I-2组每天口服给药两次(Bid)，模型组和阳性药组每天口服给药一次(Qd)。连续给药4周，每周眼眶取血，分别用qPCR和免疫荧光法检测外周血中病毒的含量和病毒相关分泌蛋白(HBeAg和HBsAg)含量；完成4周药物治疗后，安乐死小鼠取肝脏，免疫组化检测肝脏中的HBcAg水平。
结果显示口服化合物I-2给药在给药一周后，外周血中的病毒HBV DNA含量(图1)和HBsAg(图2)与HBeAg(图3)水平与模型组相比均有显著性下降(P<0.001，Two-way ANOVA)，而恩替卡韦(ETV)治疗只降低外周血中的病毒HBV DNA含量，对HBsAg与HBeAg无抑制作用。
药物治疗4周后用特异性抗HBcAg抗体检测肝脏中的HBcAg水平。免疫组化结果显示，在肝组织切片中，核心蛋白阳性的细胞呈现深黑色，阴性细胞呈现浅黑色，如图4，化合物I-2治疗组小鼠肝脏中呈现核心蛋白阳性的肝细胞明显减少，而ETV给药组与模型组无差异。
上述化合物I-2体内药效学结果表明，化合物I-2不仅可以降低感染动物体内HBV DNA水平，还能减少体内HBcAg、HBsAg和HBeAg抗原水平。而HBsAg和HBeAg作为临床上用来判断乙肝病毒感染状态和功能性治愈的指标，具有重大临床意义。
本发明化合物尤其适用于预防或治疗肝病，所述肝病包括例如肝癌、肝脏感染，包括肝炎感染。
以上详细描述了本发明的优选实施方式，但是，本发明并不限于上述实施方式中的具体细节，在本发明的技术构思范围内，可以对本发明的技术方案进行多种简单变型，这种简单变型均属于本发明的保护范围。

Claims (11)

  1. 一种式I化合物或其药学上可接受的盐：
    Figure PCTCN2020097944-appb-100001
    其中，
    R A选自卤素或氰基；
    R B选自卤素；
    R C选自H、烷基、环烷基；
    L为连接基团，选自亚烷基、亚烷基的任意碳原子被选自-O-、-CO-中至少一种基团置换所得的基团、亚环烷基；
    R D选自
    Figure PCTCN2020097944-appb-100002
    其中，
    R 1为H，烷基，芳基，被选自羟基、巯基、烷硫基、氨基烷酰基、羧基、氨基、胍基、脲基、芳基、羟基芳基、杂芳基的基团取代的烷基；
    R 2为氢或烷基；
    R 3为氢、叔丁氧羰基；
    W选自烷基、卤代烷基、环烷基、杂环基、杂链烃基、芳基、杂芳基、烷氧基、氨基；其中，所述烷基、环烷基、杂环基、杂链烃基、芳基、烷氧基、氨基任选各自独立地被1、2、3或4个选自烷基、羧基、烷氧基、杂环基、芳基、杂芳基的基团取代；
    X为单键或亚烷基；
    R 5，R 6各自独立地选自-OH、芳氧基、烷基的任意碳原子被选自-O-、-CO-、-S-中至少一种基团置换所得的基团、氨基酸残基，此处所述氨基酸残基是指氨基酸的α-氨基除去一个氢所得到的基团，所述氨基酸残基中羧基中的羟基任选被烷氧基、芳氧基、杂芳氧基或苄氧基的基团置换；此处所述氨基酸选自L构型、D构型、或L和D混合构型的下列氨基酸：甘氨酸、脯氨酸、丙氨酸、缬氨酸、正缬氨酸、亮氨酸、异亮氨酸、正亮氨酸、2-叔丁基甘氨酸、2-苯基甘氨酸、苯基丙氨酸、酪氨酸、色氨酸、组氨酸、丝氨酸、高丝氨酸、苏氨酸、半胱氨酸、S-甲基半胱氨酸、高半胱氨酸、蛋氨酸、天冬酰胺、谷氨酰胺、天冬氨酸、谷氨 酸、赖氨酸、羟基赖氨酸、鸟氨酸、2,4-二氨基丁酸、2,3-二氨基丙酸、精氨酸、瓜氨酸；或者R 5、R 6与P一起形成杂环基，所述杂环基任选被卤代芳基取代，所述杂环基任选与芳基稠合。
  2. 根据权利要求1所述的式I化合物或其药学上可接受的盐，其特征在于，
    可选地，所述“卤素”选自F、Cl、Br或I；
    可选地，所述“烷基”、“卤代烷基”、“烷氧基”、“烷硫基”等任意基团中所具有的烷基各自独立地为C 1-C 20直链或支链烷基，可选地为C 1-C 15直链或支链烷基，可选地为C 1-C 10直链或支链烷基，可选地为C 1-C 7直链或支链烷基，可选地为C 1-C 6直链或支链烷基，可选地为C 1-C 5直链或支链烷基，可选地为C 1-C 4直链或支链烷基，可选地，选自甲基、乙基、正丙基、异丙基、正丁基、异丁基、叔丁基、仲丁基、正戊基，1-甲基丁基、2-甲基丁基、3-甲基丁基、异戊基、1-乙基丙基、新戊基、正己基、1-甲基戊基、2-甲基戊基、3-甲基戊基、异己基、1,1-二甲基丁基、2,2-二甲基丁基、3,3-二甲基丁基、1,2-二甲基丁基、1,3-二甲基丁基、2,3-二甲基丁基、2-乙基丁基、正庚基、2-甲基己基、3-甲基己基、2,2-二甲基戊基、3,3-二甲基戊基、2,3-二甲基戊基、2,4-二甲基戊基、3-乙基戊基或2,2,3-三甲基丁基；可选地，选自甲基、乙基、丙基、异丙基、丁基或异丁基；
    可选地，所述“亚烷基”、“亚杂环基烷基”、“氨基烷酰基”等任意基团中所具有的亚烷基各自独立地选自C 1-C 20直链或支链亚烷基，可选地为C 1-C 15直链或支链亚烷基，可选地为C 1-C 10直链或支链亚烷基，可选地为C 1-C 8直链或支链亚烷基，可选地为C 1-C 6直链或支链亚烷基，可选地为C 1-C 4直链或支链亚烷基，可选地为C 2-C 6直链或支链亚烷基，可选地为C 3-C 6直链或支链亚烷基；
    可选地，所述“杂链烃基”为链上含1个、2个、3个或4个选自N、O、S的杂原子的直链或支链饱和或不饱和C 1-C 20杂链烃结构，可选地，为链上含1个、2个、3个或4个N的直链或支链饱和或不饱和C 1-C 20杂链烃结构，可选地，为链上含1个、2个、3个或4个N的直链或支链饱和或不饱和C 1-C 10杂链烃结构；
    可选地，所述“环烷基”为C 3-C 10单环或双环环烷基，可选地，为C 3-C 7单环环烷基，可选地，为环丙基、环丁基、环戊基、环已基或环庚基；
    可选地，所述“亚环烷基”为C 3-C 10单环或双环亚环烷基，可选地，为C 3-C 8单环亚环烷基，可选地，为亚环丙基、亚环丁基、亚环戊基、亚环己基、亚环庚基及亚环辛基；
    可选地，所述“亚杂环基”、“杂环基”、“亚杂环基烷基”等任意基团中所具有的杂环为环上含有1个、2个或3个选自N、O、S的杂原子的3-10元非芳香杂环，可选地，所述杂 环为环上含有1个或2个选自N、O的杂原子的3-10元非芳香环；所述杂环为环上含有1个或2个选自N、O的杂原子的3-6元非芳香环；可选地，所述杂环为环上含有1个或2个选自N、S的杂原子的3-10元非芳香环；所述杂环为环上含有1个或2个选自N、S的杂原子的3-6元非芳香环；
    可选地，所述“芳基”、“羟基芳基”、“卤代芳基”、“芳氧基”等任意基团中所具有的芳基为6-10元芳基；可选为苯基或萘基，可选为苯基、1-萘基、2-萘基；
    可选地，所述“杂芳氧基”、“杂芳基”等、“芳氧基”中任意基团中所含的杂芳基的环为含有1-3个选自N、O和S中的杂原子的5-10元杂芳环；可选地，为含有1-2个选自N、O和S中的杂原子的5-10元杂芳环；可选地，所述杂芳环选自吡啶环、吡咯环、嘧啶环、吡嗪环、哒嗪环、噻吩环、呋喃环；可选地，所述杂芳基为选自吡啶-2-基、吡啶-3-基、吡啶-4-基、哒嗪-3-基、哒嗪-4-基、嘧啶-2-基、嘧啶-4-基、嘧啶-5-基、吡嗪-2-基、吡嗪-3-基、吲哚基、异吲哚基、吲唑基、吲嗪基、嘌呤基、喹嗪基、喹啉基、异喹啉基、噌啉基、酞嗪基、萘啶基、喹唑啉基、喹喔啉基、噻吩并[2,3-b]呋喃基、呋喃并[3,2-b]-吡喃基、吡啶并[2,3-d]噁嗪基、吡唑并[4,3-d]噁唑基、咪唑并[4,5-d]噻唑基、吡嗪并[2,3-d]哒嗪基、咪唑并[2,1-b]噻唑基、咪唑并[1,2-b][l,2,4]三嗪基、苯并噻吩基、苯并噁唑基、苯并咪唑基、苯并噻唑基、苯并噁庚因基、苯并噁嗪基、苯并呋喃基、苯并三唑基、吡咯并[2,3-b]吡啶基、吡咯并[3,2-c]吡啶基、吡咯并[3,2-b]吡啶基、咪唑并[4,5-b]吡啶基、咪唑并[4,5-c]吡啶基、吡唑并[4,3-d]吡啶基、吡唑并[4,3-c]吡啶基、吡唑并[3,4-c]吡啶基、吡唑并[3,4-d]吡啶基、吡唑并[3,4-b]吡啶基、咪唑并[1,2-a]吡啶基、吡唑并[1,5-a]吡啶基、吡咯并[1,2-b]哒嗪基、咪唑并[1,2-c]嘧啶基、吡啶并[3,2-d]嘧啶基、吡啶并[4,3-d]嘧啶基、吡啶并[3,4-d]嘧啶基、吡啶并[2,3-d]嘧啶基、吡啶并[2,3-b]吡嗪基、吡啶并[3,4-b]吡嗪基、嘧啶并[5,4-d]嘧啶基、吡唑并[2,3-b]吡嗪基、嘧啶并[4,5-d]嘧啶基，可选为吡啶-2-基、吡啶-3-基、吡啶-4-基、嘧啶-2-基、嘧啶-4-基或嘧啶-5-基；
    可选地，所述“烷基”、“亚烷基”、“亚烷基的任意碳原子被选自-O-、-CO-中至少一种基团置换所得的基团”、“芳基”、“杂芳基”或“苄基”各自独立地被选自下列基团的取代基取代：烷基(如甲基、乙基、正丙基、异丙基、正丁基、异丁基、叔丁基)、烯基(如乙烯基、丙烯基)、炔基(如乙炔基、丙炔基)、羟基、烷氧基(如甲氧基、乙氧基、正丙氧基、异丙氧基、叔丁氧基)、酰氧基(如甲酰氧基、乙酰氧基、丙酰氧基、丁酰氧基、异丙基羰基氧基、叔丁基羰基氧基)、硫基、烷硫基(如甲硫基、乙硫基、正丙硫基、异丙硫基、叔丁硫基)、氨基、烷氨基(如甲氨基、乙氨基、二甲氨基、二乙氨基、正丙氨基、异丙氨 基、叔丁氨基)、酰氨基(如甲酰氨基、乙酰氨基、丙酰氨基、丁酰氨基、异丙基羰基氨基、叔丁基羰基氨基)、卤素(如氟、氯、溴、碘)、氰基、硝基、酰基(如甲酰基、乙酰基、羧基、甲氧碳基、乙氧羰基、异丙氧羰基、叔丁氧羰基、氨基羰基、N,N-二甲基氨基羰基)；可选为甲基、乙基、羟基、甲氧基、乙氧基、乙酰氧基、氨基、二甲氨基、乙酰氨基、氟、氯、氰基、乙酰基、甲氧羰基、乙氧羰基、氨基羰基或N,N-二甲基氨基羰基。
  3. 根据权利要求1或2所述的式I化合物或其药学上可接受的盐，
    R A选自氯、氰基；
    R B选自氯；
    R C选自H、甲基、环丙基；
    L选自-CH 2CH 2-、-CH 2CH 2CH 2-、-CH 2CH 2CH 2CH 2-、CH 2CH 2OCH 2CH 2-、
    Figure PCTCN2020097944-appb-100003
    Figure PCTCN2020097944-appb-100004
    R D选自氨基酸残基、
    Figure PCTCN2020097944-appb-100005
    其中，此处所述氨基酸残基是指氨基酸中的羧基除去羟基所得的基团，所述氨基酸残基为选自L构型、D构型、或L和D混合构型的下述氨基酸的残基：甘氨酸、脯氨酸、丙氨酸、缬氨酸、正缬氨酸、亮氨酸、异亮氨酸、正亮氨酸、2-叔丁基甘氨酸、2-苯基甘氨酸、苯丙氨酸、酪氨酸、苯乙氨酸、色氨酸、组氨酸、丝氨酸、高丝氨酸、苏氨酸、半胱氨酸、S-甲基半胱氨酸、高半胱氨酸、蛋氨酸、天冬酰胺、谷氨酰胺、天冬氨酸、谷氨酸、赖氨酸、羟基赖氨酸、鸟氨酸、2,4-二氨基丁酸、2,3-二氨基丙酸、精氨酸、瓜氨酸；所述氨基酸中α氨基上的氢原子任选被选自叔丁氧羰基的基团置换，所述氨基酸中α碳上的氢原子任选被烷基置换；
    W选自C 1-6烷基、羧基C 1-6烷基、C 5-10杂环基C 1-6烷基、C 1-6烷氧基C 6-10芳基、C 1-6烷氧基、一个或两个C 1-6烷基取代的氨基、C 5-10杂环基；
    X为单键、-CH 2-、或-CH 2(CH 3)-；
    R 5，R 6各自独立地选自-OH、C 6-10芳氧基、C 1-6烷基-O(C＝O)O-C 1-6亚烷氧基、C 1-6烷基(C＝O)O-C 1-6亚烷氧基、C 1-16烷基-O-C 1-6亚烷氧基、氨基酸残基，此处所述氨基酸残基是指氨基酸的α-氨基除去一个氢所得到的基团，所述氨基酸残基中羧基中的羟基任选被C 1-6烷氧基置换；此处所述氨基酸选自L构型、D构型、或L和D混合构型的下列氨基酸：丙 氨酸、缬氨酸、正缬氨酸、亮氨酸、异亮氨酸、正亮氨酸、2-叔丁基甘氨酸、2-苯基甘氨酸、苯基丙氨酸、酪氨酸、色氨酸、组氨酸、丝氨酸、高丝氨酸、苏氨酸、半胱氨酸、S-甲基半胱氨酸、高半胱氨酸、蛋氨酸、天冬酰胺、谷氨酰胺、天冬氨酸、谷氨酸、赖氨酸、羟基赖氨酸、鸟氨酸、2,4-二氨基丁酸、2,3-二氨基丙酸、精氨酸、瓜氨酸，优选丙氨酸；
    或者R 5、R 6与P一起形成C 5-10杂环基，所述C 5-10杂环基任选被卤代苯基取代，所述C 5-10杂环基任选与苯基稠合。
  4. 根据权利要求1-3任一项所述的式I化合物或其药学上可接受的盐，其中所述式I化合物选自下列化合物：
    Figure PCTCN2020097944-appb-100006
    Figure PCTCN2020097944-appb-100007
    Figure PCTCN2020097944-appb-100008
    Figure PCTCN2020097944-appb-100009
    Figure PCTCN2020097944-appb-100010
    Figure PCTCN2020097944-appb-100011
    Figure PCTCN2020097944-appb-100012
  5. 根据权利要求1-4任一项所述的式I化合物或其药学上可接受的盐，其特征在于，所述药学上可接受的盐包括式I化合物的阴离子盐和阳离子盐；
    可选地，所述药学上可接受的盐包括式I化合物的碱金属的盐、碱土金属的盐、铵盐；
    可选地，所述碱金属包括钠、钾、锂、铯，所述碱土金属包括镁、钙、锶；
    可选地，所述药学上可接受的盐包括式I化合物与有机碱形成的盐；
    可选地，所述有机碱包括三烷基胺、吡啶、喹啉、哌啶、咪唑、甲基吡啶、二甲氨基吡啶、二甲基苯胺、N-烷基吗啉、1,5-二氮杂双环[4.3.0]壬烯-5、1,8-二氮杂双环[5.4.0]十一碳烯-7、1,4-二氮杂双环[2.2.2]辛烷；可选地，所述三烷基胺包括三甲胺、三乙胺、N-乙基二异丙胺；可选地，所述N-烷基吗啉包括N-甲基吗啉；
    可选地，所述药学上可接受的盐包括式I化合物与酸形成的盐；
    可选地，所述酸包括无机酸、有机酸；可选地，所述无机酸包括盐酸、氢溴酸、氢碘酸、硫酸、硝酸、磷酸、碳酸；可选地，所述有机酸包括甲酸、乙酸、丙酸、草酸、丙二酸、琥珀酸、富马酸、马来酸、乳酸、苹果酸、柠檬酸、枸橼酸、酒石酸、碳酸、苦味酸、甲磺酸、乙磺酸、对甲苯磺酸、谷氨酸、双羟萘酸。
  6. 权利要求1-5任一项所述的式I化合物或其药学上可接受的盐的制备方法，该方法通过以下反应路线1实现：
    Figure PCTCN2020097944-appb-100013
    该反应路线包括：将Hu1化合物与R DOH进行缩合成酯反应，得到式I化合物，其中，式Hu1和式I中，R A、R B、L的定义如权利要求1-5任一项所述，R C为H；式I中，R D定义如权利要求1-5任一项所述；
    或者，该方法通过以下反应路线2实现：
    Figure PCTCN2020097944-appb-100014
    该反应路线包括：将Hu1-1化合物与R DOH进行缩合成酯反应，得到式Hu1-2化合物，其Hu1-2化合物中的R D为权利要求1-5任一项所定义的含有保护基的基团，然后将式Hu1-2化合物与卤化物反应引入R C，任选地脱去R D中的保护基团，得到式I化合物，其中，式Hu1-1、式Hu1-2和式I中，R A、R B、L的定义如权利要求1-5任一项所述；式I中，R D的定义如权利要求1-5任一项所述；式I中，R C为权利要求1-5任一项所定义的不为氢的基团；
    任选地，上述反应路线1和反应路线2中，采用保护基团对式Hu1化合物、式Hu1-1和式R DOH化合物进行保护，进行缩合成酯反应后，任选地进行脱保护；
    R DOH为式HOOCC(R 1)(R 2)NHR 3、式WCOOH、式PG-(P＝O)R 5R 6化合物，其中R 1、R 2、R 3、W定义如权利要求1-5任一项所述；PG为保护基团，PG优选为五氟苯氧基；R 5，R 6各自独立地选自-OH、芳氧基、氨基酸残基，此处所述氨基酸残基是指氨基酸的α-氨基除去一个氢所得到的基团，所述氨基酸残基中羧基中的羟基任选被烷氧基、芳氧基、杂芳氧基或苄氧基的基团置换；此处所述氨基酸选自L构型、D构型、或L和D混合构型的下列氨基酸：甘氨酸、脯氨酸、丙氨酸、缬氨酸、正缬氨酸、亮氨酸、异亮氨酸、正亮氨酸、2-叔丁基甘氨酸、2-苯基甘氨酸、苯基丙氨酸、酪氨酸、色氨酸、组氨酸、丝氨酸、高丝氨酸、苏氨酸、半胱氨酸、S-甲基半胱氨酸、高半胱氨酸、蛋氨酸、天冬酰胺、谷氨酰胺、天冬氨酸、谷氨酸、赖氨酸、羟基赖氨酸、鸟氨酸、2,4-二氨基丁酸、2,3-二氨基丙酸、精氨酸、瓜氨酸；或者R 5、R 6与P一起形成杂环基，所述杂环基任选被卤代芳基取代，所述杂环基任选与芳基稠合；
    可选地，所述缩合成酯反应中，所述缩合剂选自碳二亚胺例如N,N’-二环己基碳二亚胺(DCC)或N-(3-二甲基氨基异丙基)-N’-乙基碳二亚胺盐酸盐(EDC)；
    可选地，所述缩合成酯反应在碱的存在下进行；可选地，所述碱选自碱金属碳酸盐或有机碱，可选地，所述碱金属碳酸盐选自碳酸钠、碳酸钾，可选地，所述有机碱选自三乙胺、N-甲基吗啉、N-甲基哌啶、N,N-二异丙基乙胺、4-N,N-二甲基氨基吡啶；
    可选地，所述缩合成酯反应的温度为0℃至+60℃，优选+20℃至+40℃；
    可选地，所述缩合成酯反应在常压进行；
    或者，上述式I化合物或其药学上可接受的盐采用以下方法制备：
    将式Hu1化合物与式Hu2化合物反应，得到式Hu3化合物，再引入R 5和R 6基团，得到式I化合物；
    Figure PCTCN2020097944-appb-100015
    式I中，
    R A、R B、R C、L如权利要求1-5任一项所述；R D为
    Figure PCTCN2020097944-appb-100016
    其中，X如权利要求1-5任一项所述，R 5，R 6各自独立地选自-OH、芳氧基、烷基的任意碳原子被选自-O-、-CO-、-S-中至少一种基团置换所得的基团、氨基酸残基，此处所述氨基酸残基是指氨基酸的α-氨基除去一个氢所得到的基团，所述氨基酸残基中羧基中的羟基任选被烷氧基、芳氧基、杂芳氧基或苄氧基的基团置换；此处所述氨基酸选自L构型、D构型、或L和D混合构型的下列氨基酸：甘氨酸、脯氨酸、丙氨酸、缬氨酸、正缬氨酸、亮氨酸、异亮氨酸、正亮氨酸、2-叔丁基甘氨酸、2-苯基甘氨酸、苯基丙氨酸、酪氨酸、色氨酸、组氨酸、丝氨酸、高丝氨 酸、苏氨酸、半胱氨酸、S-甲基半胱氨酸、高半胱氨酸、蛋氨酸、天冬酰胺、谷氨酰胺、天冬氨酸、谷氨酸、赖氨酸、羟基赖氨酸、鸟氨酸、2,4-二氨基丁酸、2,3-二氨基丙酸、精氨酸、瓜氨酸；
    式Hu2中，X如权利要求1-5任一项所述，Q 1、Q 2各自独立地为H、烷基；
    式Hu3中，R A、R B、R C、L、X如权利要求1-5任一项所述；
    R 5和R 6基团的引入方法包括，将式Hu3化合物与相应的氨基酸进行磷酰胺化反应制得相应的磷酰胺式I化合物，或者与相应的卤代物或醇进行反应制得磷酸酯式I化合物；
    可选地，所述式I化合物的药学上可接受的盐，通过将式I化合物溶于相应的酸饱和的醇溶液中进行反应而制备。
  7. 一种药物组合物，其包含治疗有效量的上述式I化合物或其药学上可接受的盐中的一种或多种以及任选存在的药学上可接受的载体。
  8. 根据权利要求7所述的药物组合物，其特征在于，所述药物组合物的剂型包括口服制剂、直肠给药制剂、肠外给药制剂；
    可选地，所述口服制剂包括固体制剂、液体制剂，
    可选地，所述固体制剂包括片剂、粉剂、粒剂、胶囊；
    可选地，所述液体制剂包括水或油悬浮剂、糖浆；
    可选地，所述用于肠外给药制剂包括注射用的溶液、水或油性悬浮剂。
  9. 权利要求1-5任一项所述的式I化合物或其药学上可接受的盐，或者权利要求7或8所述的药物组合物在制备预防和/或治疗乙型肝炎疾病的药物中的用途。
  10. 权利要求1-5任一项所述的式I化合物或其药学上可接受的盐，或者权利要求7或8所述的药物组合物在制备乙型肝炎病毒抑制剂中的用途。
  11. 权利要求1-5任一项所述的式I化合物或其药学上可接受的盐，或者权利要求7或8所述的药物组合物在制备抑制病毒复制的药物中的用途，所述病毒包括甲肝病毒、乙肝病毒、丙肝病毒、流感病毒、腺病毒、艾滋病毒、疱疹病毒、人乳头瘤病毒。
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Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN104887680A (zh) * 2015-05-08 2015-09-09 胡继承 多聚adp核糖聚合酶抑制剂治疗乙肝病毒相关疾病的新用途
WO2017028798A1 (zh) * 2015-08-19 2017-02-23 中国科学院上海药物研究所 哒嗪酮类化合物、其制备方法、药物组合物及用途
WO2019029656A1 (zh) * 2017-08-10 2019-02-14 中国科学院上海药物研究所 酞嗪酮类化合物、其制备方法、药物组合物及用途
CN109384727A (zh) * 2017-08-10 2019-02-26 中国科学院上海药物研究所 酞嗪酮类化合物、其制备方法、药物组合物及用途

Family Cites Families (1)

* Cited by examiner, † Cited by third party
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WO2011055270A1 (en) * 2009-11-04 2011-05-12 Wyeth Llc Indole based receptor crth2 antagonists

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN104887680A (zh) * 2015-05-08 2015-09-09 胡继承 多聚adp核糖聚合酶抑制剂治疗乙肝病毒相关疾病的新用途
WO2017028798A1 (zh) * 2015-08-19 2017-02-23 中国科学院上海药物研究所 哒嗪酮类化合物、其制备方法、药物组合物及用途
WO2019029656A1 (zh) * 2017-08-10 2019-02-14 中国科学院上海药物研究所 酞嗪酮类化合物、其制备方法、药物组合物及用途
CN109384727A (zh) * 2017-08-10 2019-02-26 中国科学院上海药物研究所 酞嗪酮类化合物、其制备方法、药物组合物及用途

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