CN114702497B

CN114702497B - Crystal form of dihydropyrimidine derivative, preparation method thereof and application thereof in medicines

Info

Publication number: CN114702497B
Application number: CN202210330393.7A
Authority: CN
Inventors: 张珉; 刘辛昌; 时佳佳; 尹丽华; 闫兴国
Original assignee: South China University of Technology SCUT
Current assignee: South China University of Technology SCUT
Priority date: 2021-12-23
Filing date: 2022-03-31
Publication date: 2023-03-21
Anticipated expiration: 2042-03-31
Also published as: CN114702497A

Abstract

The invention discloses a crystal form of a dihydropyrimidine derivative, a preparation method thereof and application thereof in medicines. The crystal form A-1 and the crystal form B-1 have high stability under the conditions of high temperature, high humidity and illumination, and have better pharmacokinetic properties in beagle dogs.

Description

Crystal form of dihydropyrimidine derivative, preparation method thereof and application thereof in medicines

Technical Field

The invention belongs to the technical field of chemical medicines, and particularly relates to a crystal form of a dihydropyrimidine derivative, a preparation method thereof and application thereof in medicines.

Background

Hepatitis b virus belongs to the hepaciviridae family, and it causes acute and/or persistent progressive chronic diseases. Hepatitis b virus also causes many other clinical manifestations in pathological morphology-in particular chronic inflammation of the liver, cirrhosis and canceration of hepatocytes. In addition, co-infection with hepatitis delta can have adverse effects on the progression of the disease.

The conventional drugs licensed for the treatment of chronic hepatitis are interferon and lamivudine (lamivudine). However, interferons have only moderate activity and high toxic side effects; lamivudine (lamivudine) has good activity, but its resistance increases rapidly during treatment and often shows rebound effects after treatment is stopped.

In the related art, heteroaromatic ring substituted dihydropyrimidine (HAP) compounds represented by Bay41-4109 and Bay39-5493 have been able to inhibit HBV replication by preventing the formation of normal nucleocapsid. Research on the action mechanism of the compound discovers that the dihydropyrimidine compound substituted by the heteroaromatic ring changes the included angle between dimers forming the nucleocapsid through the action with 113-143 amino acid residues of the core protein, so that unstable swollen nucleocapsid is formed, and the degradation of the core protein is accelerated.

In addition, WO2015132276 discloses a series of dihydropyrimidine compounds having a better inhibitory effect on HBV viral replication, wherein example 25, i.e., (4- ((S) -7- (((R) -6- (2-chloro-4-fluorophenyl) -5- (methoxycarbonyl) -2- (thiazol-2-yl) -3, 6-dihydropyrimidin-4-yl) methyl) -3-oxohexahydroimidazo [1,5-a ] pyrazin-2 (3H) -yl) benzoic acid (I), has better activity.

However, the inventors found that, in the process of preparing the compound (4- ((S) -7- (((R) -6- (2-chloro-4-fluorophenyl) -5- (methoxycarbonyl) -2- (thiazol-2-yl) -3, 6-dihydropyrimidin-4-yl) methyl) -3-oxohexahydroimidazo [1,5-a ] pyrazin-2 (3H) -yl) benzoic acid (I) and its tautomer (4- ((S) -7- (((R) -6- (2-chloro-4-fluorophenyl) -5- (methoxycarbonyl) -2- (thiazol-2-yl) -1, 6-dihydropyrimidin-4-yl) methyl) -3-oxohexahydroimidazo [1,5-a ] pyrazin-2 (3H) -yl) benzoic acid (Ia), the stability and pharmacokinetic properties of the compound were not satisfactory, and much inconvenience was brought to the development of the formulation at a later stage.

As is well known, different crystal forms, salt forms and salt forms of the same drug may have obvious differences in stability, bioavailability and pharmacokinetic effects, thereby affecting the therapeutic effect of the drug, and therefore, it is of great significance to develop new crystal forms of dihydropyrimidine derivatives which are more beneficial to drug processing and pharmaceutical compositions.

Disclosure of Invention

The present invention is directed to solving at least one of the above problems in the prior art. Therefore, the invention provides a crystal form of the dihydropyrimidine derivative.

The invention also provides a preparation method of the dihydropyrimidine derivative crystal form.

The invention also provides a pharmaceutical composition.

The invention also provides application of the crystal form of the dihydropyrimidine derivative in medicaments.

The invention provides a crystal form of a compound shown as a formula (I) or a formula (Ia), wherein the crystal form is a crystal form A-1 or a crystal form B-1;

wherein the X-ray powder diffraction pattern of the crystal form A-1 comprises diffraction peaks with 2 theta angles of 10.11 +/-0.2 degrees, 10.62 +/-0.2 degrees, 13.12 +/-0.2 degrees, 15.16 +/-0.2 degrees, 18.42 +/-0.2 degrees and 19.97 +/-0.2 degrees;

the X-ray powder diffraction pattern of the crystal form B-1 comprises diffraction peaks with 2 theta angles of 7.95 +/-0.2 degrees, 9.17 +/-0.2 degrees, 9.46 +/-0.2 degrees, 11.23 +/-0.2 degrees, 14.66 +/-0.2 degrees and 17.32 +/-0.2 degrees.

The crystal form has at least the following beneficial effects:

the applicant finds that the crystal form A-1 and the crystal form B-1 have high stability under high temperature, high humidity and illumination conditions, and have better pharmacokinetic properties in beagle dogs.

According to some embodiments of the invention, the X-ray powder diffraction pattern of form a-1 of the present invention comprises diffraction peaks at 2 Θ angles of 10.11 ± 0.2 °, 10.62 ± 0.2 °, 13.12 ± 0.2 °, 15.16 ± 0.2 °, 17.02 ± 0.2 °, 17.87 ± 0.2 °, 18.42 ± 0.2 °, 19.97 ± 0.2 °, 21.26 ± 0.2 °, and 22.44 ± 0.2 °.

According to some embodiments of the present invention, the X-ray powder diffraction pattern of form a-1 of the present invention comprises diffraction peaks ± 2 θ of ± 6.56 ± 0.2 °, 10.11 ± 0.2 °, 10.62 ± 0.2 °, 12.31 ± 0.2 °, 13.12 ± 0.2 °, 14.21 ± 0.2 °, 15.16 ± 0.2 °, 17.02 ± 0.2 °, 17.64 ± 0.2 °, 17.87 ± 0.2 °, 18.42 ± 0.2 °, 19.97 ± 0.2 °, 20.30 ± 0.2 °, 21.26 ± 0.2 °, 21.40 ± 0.2 °, 21.92 ± 0.2 °, 22.44 ± 0.2 °, 22.77 ± 0.2 °, 22.87 ± 0.2 °, 23.51 ± 0.2 °, 23.80 ± 0.2 °, 25.17 ± 0.2 °, 25.34 ± 0.2 °, 25.85 ± 0.85 ± 0.2 °, 22.84 ± 0.32 °,26 ± 0.32 °, 2.32 ± 0.32 °.

According to some embodiments of the invention, the X-ray powder diffraction pattern of form B-1 of the present invention comprises diffraction peaks at 2 Θ angles of 4.60 ± 0.2 °, 7.95 ± 0.2 °, 9.17 ± 0.2 °, 9.46 ± 0.2 °, 11.23 ± 0.2 °, 13.15 ± 0.2 °, 14.66 ± 0.2 °, 15.36 ± 0.2 °, 17.32 ± 0.2 °, 17.92 ± 0.2 °, and 21.67 ± 0.2 °.

According to some embodiments of the present invention, the, the X-ray powder diffraction pattern of the crystal form B-1 comprises diffraction peaks with 2 theta angles of 4.60 +/-0.2 degrees, 7.95 +/-0.2 degrees, 9.17 +/-0.2 degrees, 9.46 +/-0.2 degrees, 11.23 +/-0.2 degrees, 13.15 +/-0.2 degrees, 14.08 +/-0.2 degrees, 14.66 +/-0.2 degrees, 15.36 +/-0.2 degrees, 15.54 +/-0.2 degrees, 17.32 +/-0.2 degrees, 17.92 +/-0.2 degrees, 19.15 +/-0.2 degrees, 19.58 +/-0.2 degrees, 20.10 +/-0.2 degrees, 21.18 +/-0.2 degrees, 21.67 +/-0.2 degrees, 22.27 +/-0.2 degrees, 22.71 +/-0.2 degrees, 23.92 +/-0.2 degrees, 24.18 +/-0.2 degrees, 24.50 +/-0.2 degrees, 25.04 +/-0.2 degrees, 25.79 +/-0.2 degrees, 26.16 +/-0.2 degrees, 23.2 degrees, 23.27.27.0.27 +/-0.0.0.27 degrees, 27.0.0.27 degrees, 27.0.0 degree, 27 +/-0.0.0.27 degrees, 27.0.73 degrees and 31 +/-0.0 degree.

According to some embodiments of the invention, form a-1 of the present invention has an X-ray powder diffraction pattern substantially as shown in figure 7.

According to some embodiments of the invention, form B-1 of the present invention has an X-ray powder diffraction pattern substantially as shown in figure 9.

According to some embodiments of the invention, the differential scanning calorimetry trace of form A-1 of the present invention comprises endothermic peaks at 89.6 ℃ ± 3 ℃ and 180.6 ℃ ± 3 ℃.

According to some embodiments of the invention, the differential scanning calorimetry trace of form B-1 of the invention comprises an endothermic peak at 174.9 ℃ ± 3 ℃.

According to some embodiments of the invention, form a-1 of the present invention has a differential scanning calorimetry trace substantially as shown in figure 8.

According to some embodiments of the invention, form B-1 of the present invention has a differential scanning calorimetry trace substantially as shown in figure 10.

A second aspect of the present invention provides a method for preparing the crystalline form of the present invention, comprising the steps of: stirring the compound shown in the formula (I) and dichloromethane at room temperature, removing dichloromethane, adding anhydrous methanol or ethyl acetate, heating to reflux while stirring, precipitating a solid, cooling to room temperature, stirring at room temperature, and performing liquid-solid separation to obtain the compound.

According to some embodiments of the invention, the liquid-solid separation is filtration, washing and vacuum drying.

The third aspect of the invention provides a pharmaceutical composition, which comprises the crystal form of the dihydropyrimidine derivative and pharmaceutically acceptable auxiliary materials thereof.

According to some embodiments of the invention, the excipient is a carrier, excipient or diluent.

The fourth aspect of the present invention also provides the use of the crystalline form of the present invention or a pharmaceutical composition containing the crystalline form described above in the preparation of a medicament, in particular in the prevention, treatment or alleviation of viral diseases in a patient.

According to some embodiments of the invention, the viral disease is hepatitis b infection or a disease caused by hepatitis b infection.

According to some embodiments of the invention, the disease caused by hepatitis B infection is liver cirrhosis or hepatocellular carcinoma.

Detailed description of the invention

The invention is intended to cover alternatives, modifications and equivalents, which may be included within the scope of the invention as defined by the appended claims. One skilled in the art will recognize that many methods and materials similar or equivalent to those described herein can be used in the practice of the present invention. The present invention is in no way limited to the methods and materials described herein. In the event that one or more of the incorporated documents, patents, and similar materials differ or contradict this application (including but not limited to defined terminology, application of terminology, described techniques, and the like), this application controls.

In the present invention, the crystalline form of the compound represented by formula (I) or formula (Ia) may contain a solvent, and in some cases, the solvent may contribute to the internal stability of the crystalline form of compound (I), compound (Ia), and common solvents include water, ethanol, methanol, isopropanol, acetone, isopropyl ether, diethyl ether, isopropyl acetate, n-heptane, tetrahydrofuran, dichloromethane, ethyl acetate, and the like. The above-mentioned crystal forms with a certain amount of moisture or other solvents should be considered to be included in the scope of the present invention as long as they have any of the characteristics of the crystal forms of the compound represented by formula (I) or formula (Ia) described in the present invention.

It will be further appreciated that certain features of the invention, which are, for clarity, described in the context of separate embodiments, may also be provided in combination in a single embodiment. Conversely, various features of the invention which are, for brevity, described in the context of a single embodiment, may also be provided separately or in any suitable subcombination.

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. All patents and publications referred to herein are incorporated by reference in their entirety. Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, the preferred methods, devices, and materials are described herein.

Definitions and general terms

The term "comprising" is open-ended, i.e. includes the elements indicated in the present invention, but does not exclude other elements.

"room temperature" in the present invention means a temperature of from 10 ℃ to 40 ℃. In some embodiments, "room temperature" refers to a temperature from 20 ℃ to 30 ℃; in other embodiments, "room temperature" refers to a temperature from 25 ℃ to 30 ℃.

The term "pharmaceutically acceptable" as used herein refers to a substance that is acceptable for pharmaceutical use from a toxicological standpoint and does not interact adversely with the active ingredient.

The terms "crystalline form", "crystal form", "polymorph (polymorphs)", "crystal modification" and "polymorph form" as used herein are understood to be synonymous. In the present invention, solid crystalline forms of a compound, a salt of a compound, or a complex are meant, including, but not limited to, single or multicomponent crystals, and/or polymorphs, solvates, hydrates, clathrates, co-crystals, salts, solvates of salts, hydrates of salts of compounds.

Polymorphs can be detected, identified, classified and characterized using well known techniques such as, but not limited to: differential Scanning Calorimetry (DSC), thermogravimetric analysis (TGA), X-ray powder diffraction (XRPD), X-ray single crystal diffraction (XRPD), vibrational spectroscopy, solution calorimetry, solid-state nuclear magnetic resonance (SSNMR), fourier transform infrared spectroscopy (FT-IR spectroscopy), raman spectroscopy (Raman spectroscopy), hot stage optical microscopy, scanning Electron Microscopy (SEM), electron crystallography, and quantitative analysis, particle Size Analysis (PSA), surface region analysis, solubility, and dissolution rate. The skilled person will appreciate that the graphical representation of such data may undergo small changes (e.g. peak relative intensity and peak position) due to factors such as instrument response changes and sample concentration and purity changes, as is well known to the skilled person. Nevertheless, the skilled person is able to compare the graphical data in the figures herein with graphical data generated for unknown crystal forms and can confirm whether the two sets of graphical data represent the same crystal form.

Unless otherwise indicated, when referring to spectra or data presented in graphical form (e.g., XRPD, infrared, raman, and NMR spectra), the term "peak" refers to a peak or other characteristic caused by non-background noise that is recognizable to one of ordinary skill in the art. The term "effective peak" refers to a peak that is at least the median size (e.g., height) of other peaks in the spectrum or data, or at least 1.5, 2, or 2.5 times the median size of other peaks in the spectrum or data.

"XRPD" refers to X-ray powder diffraction.

Information such as change, crystallinity, crystal structure state and the like of the crystal form can be detected by X-ray powder diffraction (XRPD), and the method is a common means for identifying the crystal form. XRPD patterns refer to experimentally observed diffraction patterns or parameters derived therefrom. The powder X-ray diffraction pattern is characterized by the peak position (abscissa) and the peak intensity (ordinate). The peak position depends mainly on the structure of the crystalline form, is relatively insensitive to experimental details, while its relative peak intensity depends on many factors related to sample preparation and instrument geometry. Accordingly, in some embodiments, the crystalline form of the present invention is characterized by an XRPD pattern having certain peak positions, substantially as shown in the XRPD patterns provided in the figures of the present invention. Also, the 2 θ measurement of the XRPD pattern may have experimental error, and the 2 θ measurement of the XRPD pattern may be slightly different from instrument to instrument and from sample to sample, so the 2 θ value cannot be considered absolute. According to the condition of the instrument used in the test, the diffraction peak has error tolerance of +/-0.1 DEG, +/-0.2 DEG, +/-0.3 DEG, +/-0.4 DEG or +/-0.5 DEG; in some embodiments the diffraction peaks have a margin of error of ± 0.2 °.

The term "2 θ value" or "2 θ angle" refers to the position of the peaks in degrees of an experimental setup based on X-ray diffraction experiments and is the common abscissa unit of the diffraction pattern. The experimental setup required that if the reflection was diffracted when the incident beam formed an angle θ (θ) with a certain crystal, the reflected beam was recorded at an angle 2 θ (2 θ). It is to be understood that reference herein to specific 2 θ values for a particular polymorph is intended to refer to the 2 θ values (in degrees) measured using the X-ray diffraction experimental conditions described herein.

In the context of the present invention, the 2 θ values in the X-ray powder diffraction pattern are all in degrees (°).

"relative intensity" refers to the ratio of the intensity of the first strong peak to the intensity of the other peaks when the intensity of the first strong peak is 100% of all the diffraction peaks in an X-ray powder diffraction pattern (XRPD).

Differential Scanning Calorimetry (DSC) is to measure the temperature of a sample and an inert reference substance (usually alpha-Al) by continuously heating or cooling under program control ₂ O ₃ ) The energy difference therebetween varies with temperature. The melting peak height of the DSC curve depends on many factors related to sample preparation and instrument geometry, while the peak position is relatively insensitive to experimental details. Thus, in some embodiments, the crystalline form of the present invention is characterized by a DSC profile with characteristic peak positions substantially as shown in the DSC profiles provided in the figures of the present invention. Meanwhile, the DSC profile may have experimental errors, and the peak position and peak value of the DSC profile may slightly differ between different instruments and different samples, so the peak position or peak value of the DSC endothermic peak cannot be regarded as absolute. Depending on the conditions of the instrument used in this test, the melting peak has a margin of error of + -1 deg.C, + -2 deg.C, + -3 deg.C, + -4 deg.C or + -5 deg.C. In some embodiments the melting peak has a margin of error of ± 3 ℃. Differential Scanning Calorimetry (DSC) can also be used for detecting and analyzing whether the crystal form has crystal transformation or crystal mixing phenomenon.

Solids of the same chemical composition often form isomeric, or referred to as metamorphosis, isomers of different crystal structures under different thermodynamic conditions, and this phenomenon is called polymorphism or homomultiphase phenomenon. When the temperature and pressure conditions are changed, the variants are transformed into each other, and the phenomenon is called crystal transformation. Due to the crystal form transformation, the mechanical, electrical, magnetic and other properties of the crystal can be changed greatly. When the temperature of crystal form transformation is in a measurable range, the transformation process can be observed on a Differential Scanning Calorimetry (DSC) chart, and the DSC chart is characterized in that the DSC chart has an exothermic peak reflecting the transformation process and simultaneously has two or more endothermic peaks which are respectively characteristic endothermic peaks of different crystal forms before and after transformation.

Thermogravimetric analysis (TGA) is a technique for measuring the change in mass of a substance with temperature under program control, and is suitable for examining the loss of a solvent in a crystal or the sublimation and decomposition of a sample, and it can be presumed that the crystal contains crystal water or a crystal solvent. The change in mass shown by the TGA profile depends on many factors such as sample preparation and instrumentation; the mass change of the TGA detection varies slightly from instrument to instrument and from sample to sample. There is a tolerance of + -0.1% for mass change depending on the condition of the instrument used in the test.

"solvent" refers to a substance (typically a liquid) that is capable of completely or partially dissolving another substance (typically a solid). Solvents useful in the practice of the present invention include, but are not limited to: water, acetic acid, diethyl ether, isopropyl ether, petroleum ether, isopropyl acetate, methyl t-butyl ether, N-heptane, acetone, acetonitrile, benzene, chloroform, carbon tetrachloride, methylene chloride, dimethyl sulfoxide, 1, 4-dioxane, ethanol, ethyl acetate, N-butanol, t-butanol, N-dimethylacetamide, N-dimethylformamide, formamide, formic acid, hexane, isopropanol, methanol, methyl ethyl ketone, l-methyl-2-pyrrolidone, mesitylene, nitromethane, polyethylene glycol, N-propanol, 2-acetone, pyridine, tetrahydrofuran, methyl ethyl ketone, toluene, xylene, mixtures thereof, and the like.

By "anti-solvent" is meant a fluid that facilitates precipitation of the product (or product precursor) from the solvent. The anti-solvent may comprise a cold gas, or a fluid that promotes precipitation by a chemical reaction, or a fluid that reduces the solubility of the product in the solvent; it may be the same liquid as the solvent but at a different temperature, or it may be a different liquid than the solvent.

"solvate" means having a solvent on the surface, in the crystal lattice, or on and in the crystal lattice which may be water, acetic acid, acetone, acetonitrile, benzene, chloroform, carbon tetrachloride, dichloro 26353ane, dimethylsulfoxide, 1, 4-dioxane, ethanol, ethyl acetate, butanol, t-butanol, N-dimethylacetamide, N-dimethylformamide, formamide, formic acid, heptane, hexane, isopropanol, methanol, methyl ethyl ketone, methyl pyrrolidone, mesitylene, nitromethane, polyethylene glycol, propanol, 2-propanone, pyridine, tetrahydrofuran, toluene, xylene, mixtures thereof, and the like. A specific example of a solvate is a hydrate, wherein the solvent on the surface, in the crystal lattice or on the surface and in the crystal lattice is water. The hydrates may or may not have other solvents than water on the surface of the substance, in the crystal lattice or both.

The term "equivalent" or its abbreviation "eq", as used herein, is the equivalent amount of the other raw materials required in terms of the equivalent relationship of the chemical reaction, based on the base material used in each step (1 equivalent).

Crystalline forms can be identified by a variety of techniques, such as X-ray powder diffraction (XRPD), infrared absorption spectroscopy (IR), melting point methods, differential Scanning Calorimetry (DSC), thermogravimetric analysis (TGA), nuclear magnetic resonance methods, raman spectroscopy, X-ray single crystal diffraction, dissolution calorimetry, scanning Electron Microscopy (SEM), quantitative analysis, solubility and dissolution rate, and the like.

The term "substantially as shown" means that at least 50%, or at least 60%, or at least 70%, or at least 80%, or at least 90%, or at least 95%, or at least 99% of the peaks in the X-ray powder diffraction pattern or DSC diagram are shown in the diagram.

The term "patient" as used herein refers to humans (including adults and children) or other animals. In some embodiments, "patient" refers to a human.

The term "treating" or "treatment" as used herein refers, in some embodiments, to ameliorating a disease or disorder (i.e., slowing or arresting or reducing the development of the disease or at least one clinical symptom thereof). In other embodiments, "treating" or "treatment" refers to moderating or improving at least one physical parameter, including physical parameters that may not be perceived by the patient. In other embodiments, "treating" or "treatment" refers to modulating the disease or disorder, either physically (e.g., stabilizing a perceptible symptom) or physiologically (e.g., stabilizing a parameter of the body), or both. In other embodiments, "treating" or "treatment" refers to preventing or delaying the onset, occurrence, or worsening of a disease or disorder.

Pharmaceutical compositions of the crystalline forms of the invention

As described herein, pharmaceutically acceptable excipients of the invention, e.g., as used herein, include any solvent, solid excipient, diluent, binder, disintegrant, or other liquid excipient, dispersant, flavoring or suspending agent, surfactant, isotonic agent, thickening agent, emulsifier, preservative, solid binder, glidant or lubricant, and the like, as appropriate for the particular target dosage form. As described in the following documents: in Remington, the Science and Practice of Pharmacy,21st edition,2005, ed.D.B.Troy, lippincott Williams and Wilkins, philadelphia, and Encyclopedia of Pharmaceutical Technology, eds.J.Swarbrick and J.C. Boylan,1988-1999, marcel Dekker, new York, taken together with The disclosure of The literature, indicates that different adjuvants can be used In The preparation of pharmaceutically acceptable compositions and their well-known methods of preparation. Except insofar as any conventional adjuvant is incompatible with the compounds of the invention, e.g., any adverse biological effect produced or interaction in a deleterious manner with any other component of a pharmaceutically acceptable composition, their use is contemplated by the present invention.

Substances that may serve as pharmaceutically acceptable excipients include, but are not limited to, ion exchangers; aluminum; aluminum stearate; lecithin; serum proteins, such as human serum albumin; buffer substances such as phosphates; glycine; sorbic acid; potassium sorbate; partial glyceride mixtures of saturated vegetable fatty acids; water; salts or electrolytes, such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts; colloidal silica; magnesium trisilicate; polyvinylpyrrolidone; polyacrylate esters; a wax; polyethylene-polyoxypropylene-blocking polymers; lanolin; sugars such as lactose, glucose and sucrose; starches such as corn starch and potato starch; cellulose and its derivatives such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate; gum powder; malt; gelatin; talc powder; adjuvants such as cocoa butter and suppository waxes; oils such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil; glycols, such as propylene glycol and polyethylene glycol; esters such as ethyl oleate and ethyl laurate; agar; buffering agents such as magnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water; isotonic salt; ringer's solution; ethanol; phosphoric acid buffer solution; and other non-toxic suitable lubricants such as sodium lauryl sulfate and magnesium stearate; a colorant; a release agent; coating the coating material; a sweetener; a flavoring agent; a fragrance; preservatives and antioxidants.

The crystal form or the pharmaceutical composition is suitable for treating acute and chronic viral infection of infectious hepatitis, particularly effectively inhibiting Hepatitis B Virus (HBV), and is suitable for treating or relieving diseases caused by viruses of patients, particularly acute and chronic persistent HBV viral infection, wherein chronic viral diseases caused by HBV can cause severe pathological changes, and chronic hepatitis B viral infection can cause cirrhosis and/or hepatocellular carcinoma in many cases.

Pharmaceutical compositions of crystalline forms of the compounds of the present invention may be administered in any of the following ways: oral administration, inhalation by spray, topical administration, rectal administration, nasal administration, topical administration, vaginal administration, parenteral administration such as subcutaneous, intravenous, intramuscular, intraperitoneal, intrathecal, intraventricular, intrasternal, or intracranial injection or infusion, or by means of an explanted reservoir. Preferred modes of administration are oral, intramuscular, intraperitoneal or intravenous.

The crystalline forms of the compounds of the present invention or compositions containing them which are pharmaceutically acceptable may be administered in unit dosage form. The administration dosage form can be liquid dosage form or solid dosage form. The liquid dosage forms can be true solutions, colloids, microparticles, and suspensions. Other dosage forms such as tablet, capsule, dripping pill, aerosol, pill, powder, solution, suspension, emulsion, granule, suppository, lyophilized powder for injection, etc.

Oral tablets and capsules may contain excipients such as binding agents, for example syrup, acacia, sorbitol, tragacanth or polyvinylpyrrolidone; fillers, such as lactose, sucrose, corn starch, calcium phosphate, sorbitol, glycine; lubricants, such as magnesium stearate, talc, polyethylene glycol, silica; disintegrants, such as potato starch; or acceptable humectants such as sodium lauryl sulfate. The tablets may be coated by methods known in the art of pharmacy.

Oral liquids may be in the form of suspensions, solutions, emulsions, syrups or elixirs containing hydrated oils, or may be presented as a dry product for constitution with water or other suitable vehicle before use. Such liquid preparations may contain conventional additives such as suspending agents, sorbitol, cellulose methyl ether, glucose syrup, gelatin, hydroxyethyl cellulose, carboxymethyl cellulose, aluminium stearate gelatin, hydrogenated edible fats and oils, emulsifying agents, such as lecithin, sorbitan monooleate, acacia; or a non-aqueous carrier (which may comprise an edible oil), such as almond oil, an oil such as glycerol, ethylene glycol, or ethanol; preservatives, e.g. methyl or propyl p-hydroxybenzoate, sorbic acid. Flavoring or coloring agents may be added if desired.

Suppositories may contain conventional suppository bases such as cocoa butter or other glycerides.

For parenteral administration, the liquid dosage forms are generally prepared from the compound and a sterile vehicle. The carrier is preferably water. The compound can be dissolved in the carrier or made into suspension solution according to the different carrier and drug concentration, when making injection solution, the compound is dissolved in water, filtered and sterilized, and then filled into sealed bottle or ampoule.

When applied topically to the skin, the compounds of the present invention may be formulated in the form of a suitable ointment, lotion, or cream in which the active ingredient is suspended or dissolved in one or more carriers that may be used in ointment formulations including, but not limited to: mineral oil, liquid petrolatum, white petrolatum, propylene glycol, polyethylene oxide, polypropylene oxide, emulsifying wax and water; lotions and creams may employ carriers including, but not limited to: mineral oil, sorbitan monostearate, tween 60, cetyl esters wax, cetearyl alcohol, 2-octyldodecanol, benzyl alcohol and water.

In general, it has proven advantageous, both in human medicine and in veterinary medicine, to administer the active compounds according to the invention in a total amount of from about 0.5 to 500mg, preferably from 1 to 100mg, per kg of body weight per 24 hours, if appropriate in a plurality of single doses, in order to achieve the desired effect. The amount of active compound contained in a single dose is preferably about 1 to 80mg, more preferably 1 to 50mg per kg body weight, but may be varied from the above-mentioned dose, i.e., depending on the kind and body weight of the subject to be treated, the nature and severity of the disease, the type of preparation and the mode of administration of the drug, and the period or interval of administration.

The pharmaceutical composition provided by the invention also comprises an anti-HBV medicament, wherein the anti-HBV medicament is an HBV polymerase inhibitor, an immunomodulator or interferon.

HBV drugs include lamivudine, telbivudine, tenofovir disoproxil, entecavir, adefovir dipivoxil, alfaferone, alloferon, simon interleukin, clavudine, emtricitabine, faprolivir, interferon, poncin CP, intefine, interferon alpha-1 b, interferon alpha-2 a, interferon beta-1 a, interferon alpha-2, interleukin-2, mequitylate, nitazoxanide, peginterferon alpha-2 a, ribavirin, roscovarin-A, sizopyran, euforavac, ritolimod, phosphazid, heplivav, interferon alpha-2 b, levamisole, propafegermanium, and the like.

The crystal form and the application of the pharmaceutical composition thereof

Another aspect of the invention is directed to a crystalline form or pharmaceutical composition of a compound of the invention for use in the preparation of a medicament for the prevention, treatment or amelioration of hepatitis b disease in a patient, comprising administering to the patient a pharmaceutically acceptable effective amount. Hepatitis B disease refers to liver disease caused by hepatitis B virus infection or hepatitis B infection, including acute hepatitis, chronic hepatitis, cirrhosis and hepatocellular carcinoma. Acute hepatitis b virus infection may be asymptomatic or manifest as acute hepatitis symptoms. Patients with chronic viral infections have active disease and can develop cirrhosis and liver cancer.

An "effective amount," "therapeutically effective amount," or "effective dose" of a crystalline form and/or pharmaceutically acceptable pharmaceutical composition of the invention refers to an effective amount to treat or reduce the severity of one or more of the conditions referred to herein. The crystalline forms or pharmaceutically acceptable pharmaceutical compositions of the present invention are effective over a fairly wide dosage range. For example, the daily dosage may be in the range of about 0.1mg to about 1000mg per person, divided into one or more administrations. The crystalline forms and pharmaceutical compositions according to the methods of the present invention can be in any amount administered and by any route of administration effective to treat or reduce the severity of the disease. The exact amount necessary will vary depending on the condition of the patient, depending on age, general condition of the patient, severity of infection, particular factors, mode of administration, and the like. The crystalline forms or pharmaceutical compositions of the invention may be administered in combination with one or more other therapeutic agents, as discussed herein.

Drawings

FIG. 1 is an X-ray powder diffraction (XRPD) pattern of comparative example 2;

FIG. 2 is a Differential Scanning Calorimetry (DSC) chart of comparative example 2;

FIG. 3 is an X-ray powder diffraction (XRPD) pattern of comparative example 3;

FIG. 4 is a Differential Scanning Calorimetry (DSC) chart of comparative example 3;

FIG. 5 is an X-ray powder diffraction (XRPD) pattern of comparative example 4;

FIG. 6 is a Differential Scanning Calorimetry (DSC) chart of comparative example 4;

FIG. 7 is an X-ray powder diffraction (XRPD) pattern of example 1;

FIG. 8 is a Differential Scanning Calorimetry (DSC) profile of example 1;

FIG. 9 is an X-ray powder diffraction (XRPD) pattern for example 2;

FIG. 10 is a Differential Scanning Calorimetry (DSC) chart of example 2;

FIG. 11 is an X-ray powder diffraction (XRPD) pattern of comparative example 2 after standing for 10 days under high humidity conditions;

FIG. 12 is an X-ray powder diffraction (XRPD) pattern of comparative example 1.

Detailed Description

The technical solutions in the embodiments of the present invention will be clearly and completely described below, but the embodiments of the present invention are not limited thereto.

The crystalline form may be prepared by a variety of methods including, but not limited to, for example, crystallization or recrystallization from a suitable solvent mixture; sublimation; solid state conversion from another phase; crystallization from a supercritical fluid; and spraying. Techniques for crystallization or recrystallization of crystalline forms of solvent mixtures include, but are not limited to, for example, solvent evaporation; reducing the temperature of the solvent mixture; seeding (crystal seeding) of a supersaturated solvent mixture of a compound and/or salt thereof; freeze drying the solvent mixture; and an anti-solvent (antisolvent) is added to the solvent mixture. Crystalline forms, including polymorphs, can be prepared using high throughput crystallization techniques.

Crystals (including polymorphs), methods of preparation, and characterization of drug crystals are discussed in Solid-State Chemistry of Drugs, S.R.Byrn, R.R.Pfeiffer, and J.G.Stowell, second edition, SSCI, west Lafayette, indiana (1999).

In crystallization techniques in which a solvent is utilized, the solvent is generally selected based on one or more factors including, but not limited to, for example, the solubility of the compound, the crystallization technique used, and the vapor pressure of the solvent. Combinations of solvents may be utilized. For example, the compound may be solubilized in a first solvent to obtain a solution, and then an anti-solvent is added to reduce the solubility of the compound in the solution and precipitate the crystal formation. An antisolvent is a solvent in which the compound has low solubility.

Seed crystals may be added to any crystallization mixture to promote crystallization. Seeding may be used to control the growth of a particular polymorph, and/or to control the grain size distribution of the crystallized product. Therefore, the calculation of the amount of seeds required depends on the size of the available seeds and the desired size of the average product particles, as described in "Programmed Chemical Batch crystals", J.W.Mullin and J.Nyvlt, chemical Engineering Science,1971,26, 369-377. Small sized seeds are generally required to effectively control crystal growth in the batch. Small size seeds can be produced by large crystals sieving, milling or micronization, or by solution microcrystallization. In crystal milling or micronization, care should be taken to avoid changing crystallinity from the desired crystalline form (i.e., to amorphous or other polymorphic forms).

The cooled crystallization mixture can be filtered under vacuum and the isolated solid product washed with a suitable solvent (e.g., cold recrystallization solvent). After washing, the product can be dried under a nitrogen purge to give the desired crystalline form. The product may be analyzed by suitable spectroscopic or analytical techniques including, but not limited to, for example, differential Scanning Calorimetry (DSC), X-ray powder diffraction (XRPD), and thermogravimetric analysis (TGA) to ensure that a crystalline form of the compound has formed. The resulting crystalline form may be produced in an isolated yield of greater than about 70% by weight, preferably greater than about 90% by weight, based on the weight of the compound initially used in the crystallization process. The product may optionally be de-agglomerated by co-grinding or by passing through a mesh screen.

The features and advantages of the present invention may be more readily understood by those of ordinary skill in the art after reading the following detailed description. It is to be understood that certain features of the invention, which are, for clarity reasons, described above and below in the context of separate embodiments, may also be combined to form a single embodiment. Conversely, various features of the invention which are, for brevity, described in the context of a single embodiment, may also be provided in combination to form a sub-combination thereof. The present disclosure is further illustrated by the following examples, which should not be construed as limiting the scope of the invention or to the specific steps set forth therein.

In the examples described below, all temperatures are given in degrees Celsius (. Degree. C.) unless otherwise indicated. Unless otherwise indicated, reagents were purchased from commercial suppliers such as Aldrich Chemical Company, arco Chemical Company and Alfa Chemical Company and used without further purification. General reagents were purchased from Shantou Wen Long chemical plant, guangdong Guanghua chemical plant, guangzhou chemical plant, shaoshou technology, qingdao Tenglong chemical reagent Co., ltd and Qingdao overseas chemical plant.

NMR spectral data were measured by Bruker Avance 400 NMR spectrometer or Bruker Avance III HD 600 NMR spectrometer, CDC ₁₃ ,DMSO-d6,CD ₃ OD or d 6-propanone was used as a solvent (in ppm) and TMS (0 ppm) or chloroform (7.26 ppm) was used as a reference standard. When multiple peaks occur, the following abbreviations will be used: s (singlets, singlet), s, s (singlets, singlet), d (doublets ), t (triplets, triplets), m (multiplets ), br (broadeneds, broad), dd (doublets of doublets),ddd (doubtlet of triplets), dt (doubtlet of triplets), ddt (doubtlet of triplets, double triplets), td (triplet of triplets), br. Coupling constant J, in Hertz (Hz).

The crystal form prepared by the invention is identified according to the following method:

(1) The X-ray powder diffraction (XRPD) analysis method used in the invention is as follows: the radiation source used in the Empyrean diffractometer was a (Cu, k alpha,

1.540598；

1.544426; k α 2/K α 1 intensity ratio: 0.50 Voltage was set at 45KV and current was set at 40mA. A powdery sample is prepared into a thin layer on a monocrystalline silicon sample holder, and is placed on a rotary sample table to be analyzed in 0.0168 DEG step length within the range of 3-40 deg. Data Collector software was used to collect Data, highScore Plus software processed the Data, and Data Viewer software read the Data.

(2) The Differential Scanning Calorimetry (DSC) analysis method used in the invention comprises the following steps: differential scanning calorimetry was performed using a TA Q2000 module with a thermoanalytical controller. Data were collected and analyzed using TA Instruments Thermal Solutions software. About 1-5mg of the sample was accurately weighed into a specially made aluminum crucible with a lid and the sample analysis was performed from room temperature to about 250 c using a 10 c/min linear heating device. During use, the DSC cell was purged with dry nitrogen at 50 mL/min. The endothermic peak was plotted downward, and the data was analyzed and displayed using TA Universal Analysis.

athers Xbridge-C18 (4.6X 150mm,5 μm). The detection wavelength was 250nm, the flow rate was 1.0mL/min, the column temperature was 35 ℃, and the mobile phase was acetonitrile-water (v/v = 40/60).

Low resolution Mass Spectral (MS) data were measured by an Agilent 6320 series LC-MS spectrometer equipped with a G1312A binary pump and a G1316A TCC (column temperature maintained at 30 ℃), a G1329A autosampler and G1315B DAD detector applied to the analysis, and an ESI source applied to the LC-MS spectrometer.

Both spectrometers are equipped with an Agilent Zorbax SB-C18 column, the specification is 2.1X 30mm,5 μm. The injection volume is determined by the sample concentration; the flow rate is 0.6mL/min; peaks of HPLC were recorded by UV-Vis wavelength at 210nm and 254 nm. The mobile phases were 0.1% formic acid in acetonitrile (phase a) and 0.1% formic acid in ultrapure water (phase B). Gradient elution conditions are shown in table 1:

table 1: gradient elution conditions for low resolution mass spectrometry mobile phase

Time (min)	A(CH ₃ CN,0.1％HCOOH)	B(H ₂ O,0.1％HCOOH)
			0～3	5～100	95～0
3～6	100	0
			6～6.1	100～5	0～95
6.1～8	5	95

The purity of the compounds was assessed by Agilent 1100 series High Performance Liquid Chromatography (HPLC) with UV detection at 210nm and 254nm on a Zorbax SB-C18 column, 2.1X 30mm,4 μm,10 minutes, flow rate 0.6mL/min, 5-95% (0.1% formic acid in acetonitrile) in 0.1% formic acid in water, the column temperature was maintained at 40 ℃.

Chromatographic preparative separation of Compounds was carried out by Agilent 1260 series High Performance Liquid Chromatography (HPLC) with UV detection at 278nm, calesil ODS-120 (4.6X 250mm,120A, 10u) column, flow rate of 1.0mL/min, mobile phase (10 mM ZnSO) ₄ +20mM L-valine buffer): methanol (v/v) =50/50, and the column temperature is maintained at 30 ℃.

The following acronyms are used throughout the invention as shown in Table 2

Table 2: abbreviations and full names

Preparation of Compounds of formula (I)

Step 1) Synthesis of Compound 1-1

Acetone (3138 kg) was charged into the reaction vessel, and 1, 4-bis Boc-2-piperazinecarboxylic acid (198.74 kg, 601.6 mol) was added with stirring, followed by complete dissolution with stirring, and then (S) -1-phenylethylamine (80.0 kg, 660.2mol) was added. After the addition, the reaction was stirred at 30 + -5 deg.C for 18h, centrifuged, the filter cake was washed with acetone (627.8 kg), and vacuum dried at 60 + -5 deg.C for 16h to give compound 1-1 as a white solid (85.32kg, 31.4%). MS (ESI, pos.ion) m/z of 329.3[ m-H ]] ^- 。

Step 2) Synthesis of Compound 1-2

Adding water (853.8 kg), ethyl acetate (923.0 kg) and a compound 1-1 (85.22 kg) into a reaction kettle in sequence, stirring the reaction mixture at 25 +/-5 ℃, dropwise adding concentrated hydrochloric acid to adjust the pH value to 3-4, and standing for layering. The aqueous layer was extracted with ethyl acetate (460.6 kg), the organic layers were combined, washed once with water (426.8 kg), and then the solvent was distilled off under reduced pressure to give the crude product. Adding n-hexane (1472.8 kg) into the crude product for dilution, stirring at 25 + -5 deg.C for 8h, and centrifuging to obtain compound 1-2 wet product. The wet product was dried under vacuum at 60. + -. 5 ℃ for 16h to give compound 1-2 as a white solid (60.33kg, 96.8%). MS (ESI, pos.ion) m/z:329.2[ M-H ] -.

Step 3) Synthesis of Compounds 1-3

DMF (442.4 kg) and Compound 1-2 (60.22kg, 182.3 mol) were added to the reaction vessel. Stirring at 25 +/-5 ℃ until the mixture is completely dissolved, adding N, N-diisopropylethylamine (47.0kg, 363.6 mol) and 2- (7-azabenzotriazole) N, N, N ', N' -tetramethylurea hexafluorophosphate (87.76kg, 230.8 mol), stirring at a controlled temperature for 10min, adding dibenzylamine (43.4kg, 220.0mol), stirring at a high temperature for 8h, slowly adding water (794.60 kg) for dilution, continuously stirring at 25 +/-5 ℃ for 2h, centrifuging, and washing with water (607.80 kg) to obtain a crude compound 1-3. Transferring the crude product of the compound 1-3 into a reaction kettle, adding absolute ethyl alcohol (469.0 kg), heating and refluxing, cooling to 25 +/-5 ℃ after the solid is completely dissolved, keeping the temperature and stirring for 14h, centrifuging, washing the solid with ethanol (120.2 kg), and drying in vacuum at 60 +/-5 ℃ for 16h to obtain a white solid compound 1-3 (68.15kg, 73.3%). MS (ESI, pos.ion) m/z of 454.3[ m ] +H-56] ⁺ 。

Step 4) Synthesis of Compounds 1-4

Tetrahydrofuran (551.0 kg), compound 1-3 (68.14kg, 133.7 mol) and sodium borohydride (15.07kg, 398.4 mol) are sequentially put into a reaction kettle, stirred and dissolved completely, then cooled to minus 15 +/-5 ℃, and a prepared tetrahydrofuran solution of iodine (37.27 kg of iodine simple substance is dissolved by 244.0kg of tetrahydrofuran) is slowly dripped. After dripping, heating to 50 +/-2 ℃ for reaction for 24h, cooling to-15 +/-5 ℃, slowly dripping methanol (162 kg) for quenching reaction, continuing stirring for 2h after dripping is finished, and heating and refluxing until the solid is completely dissolved.

Removing the solvent by reduced pressure evaporation, adding ethyl acetate (1225.5 kg) and sodium hydroxide aqueous solution (45.65 kg sodium hydroxide is dissolved in 454.4kg water) into the residue, stirring and dissolving, layering, stirring and washing the organic layer with water (340.8 kg) for 30min, layering, removing the ethyl acetate by reduced pressure evaporation at 50 +/-5 ℃, adding ethanol (269.8 kg) into the residue for dilution, heating and refluxing, stirring until the solid is completely dissolved, cooling to 25 +/-5 ℃, stirring for 14h, centrifuging, washing the solid with ethanol (26.4 kg), and drying the solid in vacuum at 60 +/-5 ℃ for 16h to obtain white solid compounds 1-4 (57.29kg, 86.4%). MS (ESI, pos.ion) m/z: 496.3[ m + H ]] ⁺ 。

Step 5) Synthesis of Compounds 1-6

Ethyl acetate (677.0 kg) and the compound 1-4 (57.28kg, 115.6 mol) were added to the reaction vessel in this order, and after stirring the mixture well, palladium on charcoal (5.79 kg) was added. Reacting the reaction mixture in a hydrogen atmosphere (about 1.0 MPa) at 50 ℃ for 12h, cooling to 25 +/-5 ℃, filtering, washing with ethyl acetate (258.0 kg), evaporating the filtrate under reduced pressure to remove the solvent, adding tetrahydrofuran (324.6 kg) into the obtained residue for dilution, stirring, adding sodium tert-butoxide (22.23kg, 231.3 mol), heating to reflux, reacting for 14h under heat preservation, cooling to 30 +/-5 ℃, adding water (18.23 kg) for quenching reaction. The reaction mixture was evaporated under reduced pressure to remove the solvent, and water (109.4 kg) was added to the residue, stirred at 25. + -. 5 ℃ for 1 hour, centrifuged, and washed with water (72.6 kg) to give crude compounds 1-6. Adding methyl tert-butyl ether (161.2 kg) into the crude product of compound 1-6, stirring at 25 + -5 deg.C for 2 hr, centrifugingThe solid was washed with methyl tert-butyl ether (26.8 kg) and then dried under vacuum at 60 + -5 deg.C for 16h to give compound 1-6 as a white solid (25.45kg, 91%). MS (ESI, pos.ion) m/z:483.3[2 ] M + H]+； ¹ H NMR(400MHz, CH ₃ OH-d ₄ )δ4.12(d,J＝10.6Hz,1H),4.04(d,J＝11.0Hz,1H),3.77–3.64(m,2H),3.55 (t,J＝8.9Hz,1H),3.08(dd,J＝9.3,5.2Hz,1H),2.87(td,J＝12.2,2.8Hz,1H),2.82–2.68 (m,2H),1.49(s,9H).

Step 6) Synthesis of Compounds 1-7

N ₂ A mixture of compounds 1-6 (26.76g, 110.9mmol), methyl 4-bromo-benzoate (26.23g, 121.99 mmol), palladium acetate (1.24g, 5.55mmol), tBu-XPhos (4.70g, 11.07mmol), cesium carbonate (54.19 g, 166.31 mmol) and 1, 4-dioxane (400 mL) was reacted at 110 ℃ for 5h with protection, suction filtered over hot pad celite, and the filter cake was washed with dichloromethane (2680 mL). The filtrate was concentrated, and ethanol (168 mL) was added to the residue, followed by stirring at room temperature overnight. The filter cake was filtered with suction and washed successively with ethanol (168 mL) and water (2680 mL) to give a white solid (35.65 g, 85.63%). MS (ESI, pos.ion) m/z 320.1[ m ] +H-56] ⁺ 。

Step 7) Synthesis of Compounds 1 to 8

After compounds 1 to 7 (48.17g, 128.31mmol), tetrahydrofuran (232 mL) and methanol (93 mL) were added in this order to a dry reaction flask, they were completely dissolved by stirring at room temperature, and then a solution of lithium hydroxide monohydrate (16.15g, 384.93 mmol) in water (93 mL) was added. The reaction mixture was warmed to 50 ℃ and stirred for 40min, the heating was turned off and the temperature was reduced to room temperature. Water (232 mL) and petroleum ether (232 mL) were added to the reaction system, the layers were separated by extraction, the organic layer was discarded, and the aqueous layer was extracted with petroleum ether (232 mL. Times.2). Diluting the water phase with ice water (926 mL), adjusting the pH to 3-4 with concentrated hydrochloric acid while stirring, precipitating a large amount of solid, continuing to stir at room temperature for about 2h, and pumpingFiltration and vacuum drying of the filter cake gave compounds 1-8 as white solids (43.97 g, 94.82%). MS (ESI, pos.ion) m/z 306.2[ M + H-56 ]] ⁺ 。

Step 8) Synthesis of Compounds 1-9

After adding the compounds 1 to 8 (43.97g, 121.67mmol) and dichloromethane (152 mL) in this order to a dry reaction flask and stirring them well at room temperature, trifluoroacetic acid (152 mL) was added and the mixture was stirred at room temperature for about 12 hours. The solvent was distilled off under reduced pressure, methylene chloride (152 mL) was added to the residue and the solvent was distilled off under reduced pressure twice. The residue was diluted with ethyl acetate (253 mL) and stirred at room temperature for about 3h to precipitate a large amount of solid. Filtration gave compounds 1-9 as white solids (43.05g, 94.28%). MS (ESI, pos.ion) m/z:262.1[ 2 ], [ M + H ]] ⁺ 。

Step 9) Synthesis of Compound represented by formula (I)

To a dry reaction flask were added methyl (R) -6- (bromomethyl) -4- (2-chloro-4-fluorophenyl) -2- (thiazol-2-yl) -1, 4-dihydropyrimidine-5-carboxylate (51.01g, 114.70mmol), compound 1-9 (43.05g, 114.70mmol), anhydrous potassium carbonate (31.71g, 229.4 mmol), and anhydrous ethanol (500 mL) in this order, the reaction mixture was reacted at 50 ℃ for 16h, filtered, the filter cake was washed with dichloromethane (510 mL) and anhydrous ethanol (510 mL) in this order, and the solvent was distilled off from the filtrate under reduced pressure. To the concentrated residue were added water (510 mL) and ethyl acetate (510 mL) to dilute the mixture, the layers were separated, the aqueous layer was extracted with ethyl acetate (400 mL. Times.3), finally ethyl acetate (510 mL) was added to the aqueous layer, the pH was adjusted to 6 to 7 with stirring with concentrated hydrochloric acid, a large amount of solid was precipitated, the solid was filtered, the solid was dissolved in dichloromethane (400 mL), the organic layer was washed with water (200 mL. Times.3), and the solvent was evaporated under reduced pressure to give a compound represented by formula (I) (56g, 78.1%). MS (ESI, pos.ion) M/z 625.3[ M + H ]] ⁺ ； ¹ H NMR(400MHz,CDCl ₃ )δ9.62(s,1H),8.09(d,J＝8.8Hz,2H),7.87(d,J＝3.1 Hz,1H),7.67(d,J＝8.9Hz,2H),7.49(d,J＝3.1Hz,1H),7.34–7.29(m,1H),7.16(dd,J＝8.5,2.5Hz,1H),6.95(td,J＝8.4,2.4Hz,1H),6.23(s,1H),4.20–4.05(m,3H),4.03–3.88 (m,2H),3.62(s,3H),3.52–3.46(m,1H),3.38–3.25(m,1H),3.01–2.88(m,2H),2.60– 2.48(m,1H),2.35–2.22(m,1H).

Example 1

Example 1 is crystalline form a-1 of (4- ((S) -7- (((R) -6- (2-chloro-4-fluorophenyl) -5- (methoxycarbonyl) -2- (thiazol-2-yl) -3, 6-dihydropyrimidin-4-yl) methyl) -3-oxohexahydroimidazo [1,5-a ] pyrazin-2 (3H) -yl) benzoic acid, which is prepared as follows:

the compound represented by the formula (I) prepared above (6 g,11.4 mmol) and methylene chloride (100 mL) were sequentially added to a dry reaction flask, stirred at room temperature until completely dissolved, and the solvent was distilled off under reduced pressure to obtain a foamy solid. Adding anhydrous methanol (120 mL) into the foamy solid, completely dissolving the solid under stirring, heating to reflux, gradually separating out the solid from the system, keeping the temperature and stirring for 30min, turning off the heating, naturally cooling to room temperature, then keeping the temperature and stirring for 12h, filtering, washing with anhydrous methanol (20 mL), and vacuum-drying the solid at 50 ℃ for 8h to obtain a yellowish solid (5.2g, 87%).

And (4) result identification:

(1) LC-MS and Nuclear magnetic: MS (ESI, pos.ion) m/z 625.3[ m + H ]] ⁺ ； ¹ H NMR(400MHz,CDCl ₃ ) δ(ppm)9.62(s,1H),8.09(d,J＝8.8Hz,2H),7.87(d,J＝3.1Hz,1H),7.67(d,J＝8.9Hz, 2H),7.49(d,J＝3.1Hz,1H),7.34–7.29(m,1H),7.16(dd,J＝8.5,2.5Hz,1H),6.95(td,J＝ 8.4,2.4Hz,1H),6.23(s,1H),4.20–4.05(m,3H),4.03–3.88(m,2H),3.62(s,3H),3.52– 3.46(m,1H),3.38–3.25(m,1H),3.01–2.88(m,2H),2.60–2.48(m,1H),2.35–2.22(m, 1H).

(2) Identified by Empyrean X-ray powder diffraction (XRPD) analysis: the obtained XRPD spectrum is shown in figure 7, an X-ray powder diffraction pattern of the crystal form A-1 comprises diffraction peaks with 2 theta angles of 6.56 degrees, 10.11 degrees, 10.62 degrees, 12.31 degrees, 13.12 degrees, 14.21 degrees, 15.16 degrees, 17.02 degrees, 17.64 degrees, 17.87 degrees, 18.42 degrees, 19.97 degrees, 20.30 degrees, 21.26 degrees, 21.40 degrees, 21.92 degrees, 22.44 degrees, 22.77 degrees, 22.87 degrees, 23.51 degrees, 23.80 degrees, 25.17 degrees, 25.34 degrees, 25.85 degrees, 26.46 degrees, 26.84 degrees, 27.92 degrees, 29.25 degrees, 32.38 degrees and 36.35 degrees, and the diffraction peak positions can have error tolerance of +/-0.2 degrees.

(3) Identification by TA Q2000 Differential Scanning Calorimetry (DSC) analysis: the scanning speed was 10 ℃/min and the resulting DSC curve, as shown in figure 8, contained endothermic peaks at 89.6 ℃ and 180.6 ℃, with a margin of error of ± 3 ℃ being possible.

Example 2

Example 2 is crystalline form B-1 of (4- ((S) -7- (((R) -6- (2-chloro-4-fluorophenyl) -5- (methoxycarbonyl) -2- (thiazol-2-yl) -3, 6-dihydropyrimidin-4-yl) methyl) -3-oxohexahydroimidazo [1,5-a ] pyrazin-2 (3H) -yl) benzoic acid, prepared as follows:

the compound represented by the formula (I) (0.3 g,0.5 mmol) prepared above and methylene chloride (5 mL) were sequentially added to a dry reaction flask, stirred at room temperature and dissolved completely, and the solvent was evaporated under reduced pressure to obtain a foamy solid. Adding ethyl acetate (5 mL) into the foamy solid, heating to 70 ℃, keeping the temperature and stirring for 30min, turning off the heating, and naturally cooling to room temperature. Stirring was continued for 12h, filtration was carried out, and the filter cake was washed with ethyl acetate (1.5 mL) and then dried under vacuum at 60 ℃ for 12h to give a yellowish solid (0.27g, 90%).

And (4) result identification:

(1)LC-MS：MS(ESI,pos.ion)m/z:625.3[M+H] ⁺ 。

(2) Identified by Empyrean X-ray powder diffraction (XRPD) analysis: the obtained XRPD spectrum is shown in figure 9, an X-ray powder diffraction pattern of the crystal form B-1 comprises diffraction peaks with 2 theta angles of 4.60 degrees, 7.95 degrees, 9.17 degrees, 9.46 degrees, 11.23 degrees, 13.15 degrees, 14.08 degrees, 14.66 degrees, 15.36 degrees, 15.54 degrees, 17.32 degrees, 17.92 degrees, 19.15 degrees, 19.58 degrees, 20.10 degrees, 21.18 degrees, 21.67 degrees, 22.27 degrees, 22.71 degrees, 23.92 degrees, 24.18 degrees, 24.50 degrees, 25.04 degrees, 25.79 degrees, 26.16 degrees, 26.80 degrees, 27.73 degrees, 28.43 degrees, 30.75 degrees and 37.79 degrees, and the diffraction peak positions can have error tolerance of +/-0.2 degrees.

(3) Identification by TA Q2000 Differential Scanning Calorimetry (DSC) analysis: the scanning speed was 10 ℃/min, the obtained DSC curve is shown in figure 10, and comprises an endothermic peak at 174.9 ℃, and a margin of error of ± 3 ℃ can be present.

Comparative example 1

Comparative example 1 was (4- ((S) -7- (((R) -6- (2-chloro-4-fluorophenyl) -5- (methoxycarbonyl) -2- (thiazol-2-yl) -3, 6-dihydropyrimidin-4-yl) methyl) -3-oxohexahydroimidazo [1,5-a ] pyrazin-2 (3H) -yl) benzoic acid, a solid made according to the synthetic method of example 25 in PCT application WO2015132276, identified by Empyrean X-ray powder diffraction (XRPD) analysis, and the resulting XRPD spectrum is shown in figure 12.

Comparative example 2

Comparative example 2 is (4- ((S) -7- (((R) -6- (2-chloro-4-fluorophenyl) -5- (methoxycarbonyl) -2- (thiazol-2-yl) -1, 6-dihydropyrimidin-4-yl) methyl) -3-oxohexahydroimidazo [1,5-a ] pyrazin-2 (3H) -yl) benzoic acid phosphate crystalline form a, prepared as follows:

adding the compound (0.3 g,0.5 mmol) shown in the formula (I) and acetone (4.5 mL) into a dry reaction bottle in sequence, heating to 50 ℃, completely dissolving the solid, diluting phosphoric acid (0.12g, 85%) with water (0.15 mL), adding into a reaction system, clarifying the solution after adding, keeping the temperature and stirring for 30min, closing the heating, naturally cooling to room temperature, and gradually precipitating the solid. Stirring was continued for 22h at room temperature, filtered, and the filter cake was washed with acetone (2 mL) and then dried under vacuum at 60 ℃ for 12h to give a yellow solid (0.33g, 94%).

And (4) result identification:

(1)LC-MS：MS(ESI,pos.ion)m/z:625.3[M+H] ⁺ 。

(2) Identified by Empyrean X-ray powder diffraction (XRPD) analysis: the obtained XRPD spectrum is shown in figure 1, the X-ray powder diffraction pattern of the phosphate crystal form A comprises diffraction peaks with 2 theta angles of 5.81 degrees, 6.46 degrees, 11.85 degrees, 12.85 degrees, 14.10 degrees, 15.53 degrees, 16.90 degrees, 17.35 degrees, 18.22 degrees, 19.32 degrees, 20.79 degrees, 23.20 degrees, 23.79 degrees, 24.80 degrees, 25.64 degrees, 27.33 degrees, 28.26 degrees, 28.97 degrees, 29.62 degrees and 30.38 degrees, and the positions of the diffraction peaks can have error tolerance of +/-0.2 degrees.

(3) Identification by TA Q2000 Differential Scanning Calorimetry (DSC) analysis: the scan rate was 10 ℃/min and the resulting DSC curve is shown in figure 2, containing an endothermic peak at 207.2 ℃, with a tolerance of ± 3 ℃ being possible.

(4) Ion chromatography:

salt formation ratio of the compound of formula (Ia) to phosphoric acid in (4- ((S) -7- (((R) -6- (2-chloro-4-fluorophenyl) -5- (methoxycarbonyl) -2- (thiazol-2-yl) -1, 6-dihydropyrimidin-4-yl) methyl) -3-oxohexahydroimidazo [1,5-a ] pyrazin-2 (3H) -yl) benzoic acid phosphate form a was determined by ion chromatography (TI-00586), with the method parameters as shown in table 3.

Table 3 ion chromatograph parameter settings

The test result shows that the salifying molar ratio of the compound shown in the formula (Ia) in the (4- ((S) -7- (((R) -6- (2-chloro-4-fluorophenyl) -5- (methoxycarbonyl) -2- (thiazole-2-yl) -1, 6-dihydropyrimidin-4-yl) methyl) -3-oxohexahydroimidazo [1,5-a ] pyrazine-2 (3H) -yl) benzoic acid phosphate crystal form A to phosphoric acid is 1.

Comparative example 3

Comparative example 3 was (4- ((S) -7- (((R) -6- (2-chloro-4-fluorophenyl) -5- (methoxycarbonyl) -2- (thiazol-2-yl) -1, 6-dihydropyrimidin-4-yl) methyl) -3-oxohexahydroimidazo [1,5-a ] pyrazin-2 (3H) -yl) benzoic acid mesylate crystalline form a, prepared as follows:

adding a compound (6 g,9.6 mmol) shown in the formula (I) and acetone (90 mL) into a dry reaction bottle in sequence, heating to 50 ℃, completely dissolving the solid, diluting methanesulfonic acid (1.01g, 10.56mmol) with water (1.8 mL), adding into a reaction system, after the addition is finished, keeping the temperature and stirring for 30min, separating out a large amount of solid in the process of keeping the temperature and stirring, closing the heating after the temperature is finished, and naturally cooling to room temperature. Stirring was continued for 19h at room temperature, filtered, washed with acetone (30 mL) and the solid dried under vacuum at 60 ℃ for 12h to give a yellow solid (5.73g, 82.8%).

And (4) result identification:

(1) LC-MS and Nuclear magnetic: MS (ESI, pos.ion) m/z of 625.3[ m ] +H] ⁺ ； ¹ H NMR(400MHz, CH ₃ OH-d ₄ )δ(ppm)8.06–8.01(m,3H),7.93(d,J＝3.1Hz,1H),7.73(d,J＝8.9Hz,2H), 7.56(dd,J＝8.7,6.0Hz,1H),7.30(dd,J＝8.6,2.5Hz,1H),7.14(td,J＝8.4,2.5Hz,1H), 6.21(s,1H),4.78(d,J＝16.1Hz,1H),4.60(d,J＝16.0Hz,1H),4.43–4.32(m,1H),4.26 (dd,J＝14.8,3.2Hz,1H),4.18(t,J＝9.4Hz,1H),3.90–3.73(m,3H),3.65(s,3H),3.62– 3.51(m,1H),3.39–3.35(m,1H),3.31–3.23(m,1H),2.70(s,3H).

(2) Identified by Empyrean X-ray powder diffraction (XRPD) analysis: the obtained XRPD spectrum is shown in fig. 3, the X-ray powder diffraction pattern of mesylate crystal form a includes diffraction peaks with 2 θ angles of 3.98 °, 7.91 °, 9.69 °, 10.21 °, 11.51 °, 11.86 °, 12.11 °, 13.38 °, 14.86 °, 15.21 °, 15.80 °, 16.82 °, 18.12 °, 18.38 °, 19.16 °, 19.62 °, 21.06 °, 21.66 °, 22.06 °, 22.57 °, 23.35 °, 23.81 °, 24.20 °, 24.90 °, 25.17 °, 26.35 °, 26.92 °, 28.42 °, 29.03 °, 29.89 °, 30.89 °, 31.52 °, 31.90 °, 32.92 °, 34.94 °, 35.78 °, 36.70 °, 38.57 °, 39.59 °, 40.20 °, 40.82 °, 41.67 °, 42.35 °, 43.75 °, 44.98 °, 46.66 ° and 49.02 °, and diffraction peak position tolerance of the diffraction peak error of 0.02 ° exists.

(3) Identification by TA Q2000 Differential Scanning Calorimetry (DSC) analysis: the scanning speed was 10 ℃/min and the resulting DSC curve, as shown in figure 4, contained an endothermic peak at 217.0 ℃ with a margin of error of ± 3 ℃.

Comparative example 4

Comparative example 4 was crystalline form B of (4- ((S) -7- (((R) -6- (2-chloro-4-fluorophenyl) -5- (methoxycarbonyl) -2- (thiazol-2-yl) -1, 6-dihydropyrimidin-4-yl) methyl) -3-oxohexahydroimidazo [1,5-a ] pyrazin-2 (3H) -yl) benzoic acid mesylate, prepared as follows:

adding a compound (0.3 g,0.5 mmol) shown in the formula (I) and acetone (4.5 mL) into a dry reaction bottle in sequence, heating to 50 ℃, completely dissolving the solid, diluting methanesulfonic acid (51 mg) with water (0.3 mL), adding into a reaction system, clarifying the solution after adding, keeping the temperature, stirring for 30min, closing and heating, naturally cooling to room temperature, and gradually precipitating the solid. Stirring was continued for 12h at room temperature, filtered, washed with acetone (2 mL) and the solid dried under vacuum at 60 ℃ for 12h to give a yellow solid (0.25 g, 70%).

And (4) result identification:

(1) LC-MS and Nuclear magnetic: MS (ESI, pos.ion) m/z of 625.3[ m ] +H] ⁺ 。 ¹ H NMR(400MHz, CH ₃ OH-d ₄ )δ(ppm)8.06–8.01(m,3H),7.93(d,J＝3.1Hz,1H),7.73(d,J＝8.9Hz,2H), 7.56(dd,J＝8.7,6.0Hz,1H),7.30(dd,J＝8.6,2.5Hz,1H),7.14(td,J＝8.4,2.5Hz,1H), 6.21(s,1H),4.78(d,J＝16.1Hz,1H),4.60(d,J＝16.0Hz,1H),4.43–4.32(m,1H),4.26 (dd,J＝14.8,3.2Hz,1H),4.18(t,J＝9.4Hz,1H),3.90–3.73(m,3H),3.65(s,3H),3.62– 3.51(m,1H),3.39–3.35(m,1H),3.31–3.23(m,1H),2.70(s,3H).

(2) Identified by Empyrean X-ray powder diffraction (XRPD) analysis: the obtained XRPD spectrum is shown in fig. 5, the X-ray powder diffraction pattern of mesylate form B comprises diffraction peaks with 2 θ angles of 5.53 °, 6.69 °, 12.15 °, 12.46 °, 12.98 °, 14.34 °, 15.10 °, 16.35 °, 16.50 °, 17.59 °, 17.80 °, 18.41 °, 19.05 °, 19.43 °, 19.68 °, 20.04 °, 20.52 °, 20.79 °, 21.72 °, 22.29 °, 22.44 °, 22.83 °, 22.94 °, 23.74 °, 24.54 °, 25.52 °, 26.22 °, 27.66 °, 28.39 °, 28.74 °, 29.56 °, 30.46 °, and 31.03 °, and the diffraction peak positions can have an error tolerance of ± 0.2 °.

(3) Identification by TA Q2000 Differential Scanning Calorimetry (DSC) analysis: the scan rate was 10 ℃/min and the resulting DSC curve, as shown in figure 6, contained 197.0 ℃ and a margin of error of ± 3 ℃ may be present.

Performance testing

High-temperature test: taking a proper amount of a test product, placing the test product into a flat weighing bottle, spreading the test product into a thin layer with the thickness less than or equal to 5mm, placing the thin layer at the temperature of 60 ℃ for 10 days, and sampling and detecting the appearance, related substances and purity on the 5 th day and the 10 th day. If the tested sample is changed obviously, the test is carried out at 40 ℃. If there is no significant change at 60 ℃, then the 40 ℃ test is not necessary.

High humidity test: taking a proper amount of a test article, placing the test article into a flat weighing bottle, spreading the test article into a thin layer with the thickness of less than or equal to 5mm, placing the test article for 10 days at the temperature of 25 ℃ and under the condition that the relative humidity is 90% +/-5%, sampling and detecting the appearance, related substances and purity on the 5 th day and the 10 th day, and simultaneously accurately weighing the weight of the test article before and after the test so as to examine the moisture absorption and deliquescence performance of the test article. If the moisture absorption weight gain is more than 5%, performing the test by the same method under the conditions of 25 ℃ and 75% +/-5% of relative humidity; if the moisture absorption weight gain is less than 5 percent and other expedition items meet the requirements, the test is not carried out. ( Note: before the high-humidity test, the flat weighing bottle is put into a constant-humidity box (or a dryer containing a saturated potassium nitrate solution) for presaturation for one day, then the sample and the flat weighing bottle are weighed together, and the mass of the flat weighing bottle and the mass of the sample are recorded. )

And (3) illumination test: placing a proper amount of the test product into a flat weighing bottle, spreading into a thin layer with thickness of less than or equal to 5mm, placing in a light box (with ultraviolet) with an opening at the illumination of 4500 + -500 lx and the ultraviolet light of more than or equal to 0.7w/m ² The sample was taken at 5 and 10 days to examine appearance, related substances and purity. The test results are shown in table 4 below:

in the tests of high temperature test, high humidity test and illumination test, whether the tested crystal form is subjected to crystal transformation or not is identified through Empyrean X-ray powder diffraction (XRPD) analysis.

Table 4: chemical stability influencing factor data of the crystal forms or salts according to the invention

N/A represents not measured

And (4) conclusion: the inventive examples 1 and 2 were stable under high temperature, high humidity and light conditions, wherein the comparative example 2 was unstable under high temperature conditions, and after 10 days of high humidity, XRD detection occurred with crystal transformation, see fig. 11, and thus, was also unstable under high humidity conditions; of these, comparative example 1 is unstable under light conditions.

Pharmacokinetic evaluation of test animals following oral dosing of test samples

The experimental method comprises the following steps:

beagle dogs were orally administered 5mg/kg of the test capsule formulation, and blood was collected from the anterior limb of beagle dogs at various time points (0.083, 0.25, 0.5,1,2,4,6,8, 10 and 24 hours) after administration, collected in an anticoagulant tube of EDTA-K2, and centrifuged to prepare plasma. Plasma samples were pre-processed and then quantitatively analyzed on a triple quadrupole tandem mass spectrometer in a multiplex reactive ion monitoring (MRM) mode. Pharmacokinetic parameters were calculated using a non-compartmental model using WinNonlin 6.1 software.

Table 5: in vivo metabolism test data of test sample in beagle dog

The experimental results show that the exposure of the example 1, the example 2 and the comparative example 3 of the invention in beagle dogs is better and is obviously better than the exposure of the comparative example 4 and the comparative example 1 in beagle dogs, which shows that the active ingredients have good absorption in beagle dogs and have good application prospect in the aspect of anti-HBV.

It should be understood that the above-described embodiments of the present invention are merely examples for clearly illustrating the present invention, and are not intended to limit the embodiments of the present invention. Other variations and modifications will be apparent to persons skilled in the art in light of the above description. And are neither required nor exhaustive of all embodiments. Any modification, equivalent replacement, and improvement made within the spirit and principle of the present invention should be included in the protection scope of the claims of the present invention.

Claims

1. A crystal form of a compound shown as a formula (I) or a formula (Ia), which is characterized in that the crystal form is a crystal form A-1 or a crystal form B-1;

(I) Or

(Ia)

2. A crystalline form according to claim 1, characterized in that the form a-1 has an X-ray powder diffraction pattern comprising diffraction peaks at 2 Θ angles of 10.11 ± 0.2 °, 10.62 ± 0.2 °, 13.12 ± 0.2 °, 15.16 ± 0.2 °, 17.02 ± 0.2 °, 17.87 ± 0.2 °, 18.42 ± 0.2 °, 19.97 ± 0.2 °, 21.26 ± 0.2 ° and 22.44 ± 0.2 °.

3. A crystalline form according to claim 1, characterized in that the X-ray powder diffraction pattern of crystalline form a-1 comprises diffraction peaks at 2 Θ angles of 6.56 ± 0.2 °, 10.11 ± 0.2 °, 10.62 ± 0.2 °, 12.31 ± 0.2 °, 13.12 ± 0.2 °, 14.21 ± 0.2 °, 15.16 ± 0.2 °, 17.02 ± 0.2 °, 17.64 ± 0.2 °, 17.87 ± 0.2 °, 18.42 ± 0.2 °, 19.97 ± 0.2 °, 20.30 ± 0.2 °, 21.26 ± 0.2 °, 21.40 ± 0.2 °, 21.92 ± 0.2 °, 22.44 ± 0.2 °, 22.77 ± 0.2 °, 22.87 ± 0.2 °, 23.51 ± 0.2 °, 23.80 ± 0.2 °, 25.17 ± 0.2 °, 25.34 ± 0.2 °, 25.25.85 ± 0.85 ± 0.2 °,22 ± 0.26 ± 0.32 °, 2.32 °,26 ± 0.32 °.

4. A crystalline form according to claim 1 or 2, characterized in that the form B-1 has an X-ray powder diffraction pattern comprising diffraction peaks at 2 Θ angles of 4.60 ± 0.2 °, 7.95 ± 0.2 °, 9.17 ± 0.2 °, 9.46 ± 0.2 °, 11.23 ± 0.2 °, 13.15 ± 0.2 °, 14.66 ± 0.2 °, 15.36 ± 0.2 °, 17.32 ± 0.2 °, 17.92 ± 0.2 ° and 21.67 ± 0.2 °.

5. The crystalline form according to claim 1, the X-ray powder diffraction pattern of the crystal form B-1 comprises diffraction peaks with 2 theta angles of 4.60 +/-0.2 degrees, 7.95 +/-0.2 degrees, 9.17 +/-0.2 degrees, 9.46 +/-0.2 degrees, 11.23 +/-0.2 degrees, 13.15 +/-0.2 degrees, 14.08 +/-0.2 degrees, 14.66 +/-0.2 degrees, 15.36 +/-0.2 degrees, 15.54 +/-0.2 degrees, 17.32 +/-0.2 degrees, 17.92 +/-0.2 degrees, 19.15 +/-0.2 degrees, 19.58 +/-0.2 degrees, 20.10 +/-0.2 degrees, 21.18 +/-0.2 degrees, 21.67 +/-0.2 degrees, 22.27 +/-0.2 degrees, 22.71 +/-0.2 degrees, 23.92 +/-0.2 degrees, 24.18 +/-0.2 degrees, 24.50 +/-0.2 degrees, 25.04 +/-0.2 degrees, 25.79 +/-0.2 degrees, 26.16 +/-0.2 degrees, 23.2 degrees, 27.27.0.27 +/-0.0.27.0.0.27 degrees, 27.0.0 degree, 27.0.73 +/-0.0 degrees, 27 degrees and 31 +/-0.0 degree.

6. A crystalline form according to claim 4 or 5, characterized in that the form A-1 has an X-ray powder diffraction pattern substantially as shown in figure 7; or said form B-1 has an X-ray powder diffraction pattern substantially as shown in figure 9.

7. A crystalline form according to claim 1 or 2, characterized in that said differential scanning calorimetry trace of form a-1 comprises endothermic peaks at 89.6 ℃ ± 3 ℃ and 180.6 ℃ ± 3 ℃; or said form B-1 comprises an endothermic peak at 174.9 ℃. + -. 3 ℃ in a differential scanning calorimetry trace.

8. A crystalline form according to claim 7, characterized in that form A-1 has a differential scanning calorimetry pattern substantially as shown in figure 8.

9. A crystalline form according to claim 7, characterized in that said form B-1 has a differential scanning calorimetry pattern substantially as shown in figure 10.

10. A method for preparing a crystalline form according to any one of claims 1 to 9, characterized in that the method for preparing the crystalline form comprises the following steps: stirring the compound shown in the formula (I) and dichloromethane at room temperature, removing dichloromethane, adding anhydrous methanol or ethyl acetate, heating to reflux while stirring, precipitating a solid, cooling to room temperature, stirring at room temperature, and performing liquid-solid separation to obtain the compound.

11. A pharmaceutical composition, which is characterized by comprising the crystal form of any one of claims 1 to 9 and a pharmaceutically acceptable excipient.

12. Use of the crystalline form according to any one of claims 1 to 9 or the pharmaceutical composition of claim 11 in the manufacture of a medicament for the prevention, treatment or alleviation of a viral disease in a patient; the viral diseases refer to hepatitis B infection or diseases caused by hepatitis B infection.

13. The use of claim 12, wherein the disease caused by hepatitis B infection is liver cirrhosis or hepatocellular carcinoma.