Cyano-substituted polypeptide compound crystals and method for producing the same

Abstract

The present invention discloses crystals of polypeptide compounds containing cyano substituents and a method for preparing the same, specifically, crystals of the compound represented by formula (I) and a method for preparing the same. [Formula 1] JPEG2025533996000028.jpg42170

Description

[Technical Field]

[0001] This application claims priority to Chinese Patent Application No. 2022112639720, filed on 14 October 2022. This application incorporates the full text of the aforementioned Chinese Patent Application.

[0002] The present invention relates to crystals of cyano-substituted polypeptide compounds and methods for producing the same, and more specifically to crystals of a compound represented by formula (I) and methods for producing the same. [Background technology]

[0003] In December 2019, COVID-19 (Coronavirus 2019) emerged, rapidly developing into a pandemic and having a significant impact on global public health systems and the world economy. The pathogen, Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), has a very high RNA genome similarity (approximately 80%) to SARS-CoV-1, which led to the 2003 SARS outbreak. In response to the spread and severity of SARS-CoV-2 infection, many countries and organizations have made tremendous efforts to develop prevention and treatments, including vaccine development, large-scale vaccination programs, and the development and use of several therapeutic drugs.

[0004] Coronaviruses are enveloped, single-stranded positive-sense RNA viruses that encode structural and non-structural proteins that facilitate viral entry into and replication within a host. In the viral replication cycle, the non-structural protein 3CL (3-chymotrypsin-like protease) protease plays a crucial role, primarily by hydrolyzing two polyproteins expressed by the virus. Sequence analysis has indicated that 3CL protease may be a key target for drug design.

[0005] The currently available 3CL protease is Pfizer's nilmatrelvir tablets, which can reduce hospitalization and / or mortality rates by 89% and 70% respectively in high-risk and low-risk groups, and has been approved for sale or emergency use in the United States and many other countries. Currently, numerous 3CL proteases are in various stages of clinical trials both domestically and internationally. Therefore, the development of 3CL protease inhibitors is particularly important for the treatment of COVID-19 or other coronavirus infections. [Overview of the project]

[0006] The present invention provides a type A crystal of a compound represented by formula (I), characterized in that its powder X-ray diffraction spectrum has characteristic diffraction peaks at the following 2θ angles: 10.655±0.200°, 11.988±0.200°, 16.055±0.200°, 18.356±0.200°, and 20.083±0.200°. [ka]

[0007] In some embodiments of the present invention, a type A crystal of the compound represented by formula (I) contains at least 6, 7, or 8 diffraction peaks selected from 6.917±0.200°, 10.655±0.200°, 11.988±0.200°, 14.481±0.200°, 16.055±0.200°, 17.653±0.200°, 18.356±0.200°, and 20.083±0.200° in a powder X-ray diffraction spectrum expressed as a 2θ angle.

[0008] In some embodiments of the present invention, the A-type crystal of the compound represented by formula (I) has characteristic diffraction peaks in its powder X-ray diffraction spectrum at the following 2θ angles: 6.917±0.200°, 10.655±0.200°, 11.988±0.200°, 14.481±0.200°, 16.055±0.200°, 17.653±0.200°, 18.356±0.200°, and 20.083±0.200°.

[0009] In some embodiments of the present invention, the A-type crystal of the compound represented by formula (I) has powder X-ray diffraction spectra expressed in 2θ angles, with values ​​of 6.917±0.200°, 10.655±0.200°, 11.988±0.200°, 13.837±0.200°, 14.481±0.200°, 16.055±0.200°, 17.653±0.200°, and 18. It includes at least 12, 13, 14, 15, or 16 diffraction peaks selected from 0.356±0.200°, 20.083±0.200°, 20.839±0.200°, 21.388±0.200°, 22.379±0.200°, 24.577±0.200°, 25.104±0.200°, 26.402±0.200°, and 31.540±0.200°.

[0010] In some embodiments of the present invention, the powder X-ray diffraction spectrum of the A-type crystal of the compound represented by formula (I) is as follows: 2θ angles: 6.917±0.200°, 10.655±0.200°, 11.988±0.200°, 13.837±0.200°, 14.481±0.200°, 16.055±0.200°, 17.653±0 Characteristic diffraction peaks are observed at 0.200°, 18.356±0.200°, 20.083±0.200°, 20.839±0.200°, 21.388±0.200°, 22.379±0.200°, 24.577±0.200°, 25.104±0.200°, 26.402±0.200°, and 31.540±0.200°.

[0011] In some embodiments of the present invention, the powder X-ray diffraction spectrum of the A-type crystal of the compound represented by formula (I) is as shown in Figure 1.

[0012] In some embodiments of the present invention, the peak position, interplane distance, and relative intensity of the diffraction peak in the powder X-ray diffraction (XRPD) spectrum of a type A crystal of the compound represented by formula (I) are as shown in Table 1. [Table 1]

[0013] In some embodiments of the present invention, the A-type crystal of the compound represented by the above formula (I) has a peak value of an endothermic peak in its differential scanning calorimetry curve (DSC) at 173.5 °C ± 3 °C.

[0014] In some embodiments of the present invention, the A-type crystal of the compound represented by the above formula (I) has a differential scanning calorimetry curve as shown in FIG. 2.

[0015] In some embodiments of the present invention, the A-type crystal of the compound represented by the above formula (I) has a 0.00% weight loss at 150.0 °C ± 3 °C in its thermogravimetric analysis curve (TGA).

[0016] In some embodiments of the present invention, the A-type crystal of the compound represented by the above formula (I) has a thermogravimetric analysis curve as shown in FIG. 3.

[0017] The present invention further provides a method for producing the A-type crystal of the compound represented by the above formula (I), which includes a step of stirring the compound represented by the formula (I) in water and further includes a step of separation.

[0018] In some embodiments of the present invention, the A-type crystal of the compound represented by the formula (I) of the present invention (1) A method of adding water to the compound represented by the formula (I) and stirring (2) A method of filtering and concentrating under reduced pressure to remove moisture is produced by.

[0019] In some embodiments of the present invention, the stirring in the above step (1) is carried out under heating conditions, and preferably, the stirring temperature is 50 °C.

[0020] The present invention further provides a B-type crystal of the compound represented by formula (I), characterized in that its powder X-ray diffraction spectrum has characteristic diffraction peaks at the following 2θ angles: 9.202±0.200°, 13.394±0.200°, 14.212±0.200°, 15.516±0.200°, and 19.917±0.200°. [ka]

[0021] In some embodiments of the present invention, a type B crystal of the compound represented by formula (I) contains at least 6, 7, or 8 diffraction peaks selected from 9.202±0.200°, 10.968±0.200°, 13.394±0.200°, 14.212±0.200°, 15.516±0.200°, 16.483±0.200°, 18.306±0.200°, and 19.917±0.200° in a powder X-ray diffraction spectrum expressed as a 2θ angle.

[0022] In some embodiments of the present invention, the powder X-ray diffraction spectrum of the compound type B crystal represented by formula (I) has characteristic diffraction peaks at the following 2θ angles: 9.202±0.200°, 10.968±0.200°, 13.394±0.200°, 14.212±0.200°, 15.516±0.200°, 16.483±0.200°, 18.306±0.200°, and 19.917±0.200°.

[0023] In some embodiments of the present invention, the B-type crystal of the compound represented by formula (I) has powder X-ray diffraction spectra expressed in 2θ angles, with values ​​of 5.459±0.200°, 7.096±0.200°, 9.202±0.200°, 10.968±0.200°, 13.394±0.200°, 14.212±0.200°, 15.516±0.200°, and 16. It includes at least 12, 13, 14, 15, or 16 diffraction peaks selected from 483±0.200°, 18.306±0.200°, 19.488±0.200°, 19.917±0.200°, 20.681±0.200°, 21.526±0.200°, 22.409±0.200°, 24.622±0.200°, and 25.661±0.200°.

[0024] In some embodiments of the present invention, the powder X-ray diffraction spectrum of the B-type crystal of the compound represented by formula (I) is as follows: 2θ angles: 5.459±0.200°, 7.096±0.200°, 9.202±0.200°, 10.968±0.200°, 13.394±0.200°, 14.212±0.200°, 15.516±0. Characteristic diffraction peaks are observed at 200°, 16.483±0.200°, 18.306±0.200°, 19.488±0.200°, 19.917±0.200°, 20.681±0.200°, 21.526±0.200°, 22.409±0.200°, 24.622±0.200°, and 25.661±0.200°.

[0025] In some embodiments of the present invention, the type B crystal of the compound represented by formula (I) has a powder X-ray diffraction (XRPD) spectrum as shown in Figure 4.

[0026] In some embodiments of the present invention, the peak position, interplane distance, and relative intensity of the diffraction peak in the powder X-ray diffraction spectrum of the B-type crystal of the compound represented by formula (I) are as shown in Table 2. [Table 2]

[0027] In some embodiments of the present invention, the type B crystal of the compound represented by formula (I) has a differential scanning calorimetry curve (DSC) with a peak value of an endothermic peak at 128.4°C ± 3°C.

[0028] In some embodiments of the present invention, the B-type crystal of the compound represented by formula (I) has a differential scanning calorimetry curve as shown in Figure 5.

[0029] In some embodiments of the present invention, the B-type crystal of the compound represented by formula (I) decreases in weight by 9.77% when its thermogravimetric analysis curve (TGA) is at 150.0°C ± 3°C.

[0030] In some embodiments of the present invention, the B-type crystal of the compound represented by formula (I) has a thermogravimetric analysis curve as shown in Figure 6.

[0031] In some embodiments of the present invention, the B-type crystal of the compound represented by formula (I) may exist in the form of a solvate crystal.

[0032] The present invention further provides a method for producing type B crystals of the compound represented by formula (I), comprising the steps of stirring the compound represented by formula (I) in an alcohol-based / n-heptane mixed solvent at room temperature, and further comprising the step of separation.

[0033] In some embodiments of the present invention, the B-type crystal of the compound represented by formula (I) of the present invention is (1) A method in which an alcohol-based / n-heptane mixed solvent is added to the compound represented by formula (I) and stirred at room temperature. (2) Method of volatilization at low temperature and concentration under reduced pressure It is manufactured by [company name].

[0034] In some embodiments of the present invention, the alcohol-based / n-heptane mixed solvent in step (1) above is preferably ethanol / n-heptane.

[0035] In some embodiments of the present invention, the solvent ratio of ethanol / n-heptane is preferably 1 / 4 (by volume).

[0036] In some embodiments of the present invention, the low temperature in step (2) above is preferably 5°C.

[0037] The present invention further provides a C-type crystal of a compound represented by formula (I), characterized in that its powder X-ray diffraction spectrum has characteristic diffraction peaks at the following 2θ angles: 6.334±0.200°, 8.399±0.200°, 9.435±0.200°, 12.060±0.200°, and 18.126±0.200°. [ka]

[0038] In some embodiments of the present invention, a C-type crystal of the compound represented by formula (I) contains at least 6, 7, or 8 diffraction peaks selected from 4.883±0.200°, 6.334±0.200°, 8.399±0.200°, 8.928±0.200°, 9.435±0.200°, 11.168±0.200°, 12.060±0.200°, and 18.126±0.200° in a powder X-ray diffraction spectrum expressed as a 2θ angle.

[0039] In some embodiments of the present invention, the powder X-ray diffraction spectrum of the compound C-type crystal represented by formula (I) has characteristic diffraction peaks at the following 2θ angles: 4.883±0.200°, 6.334±0.200°, 8.399±0.200°, 8.928±0.200°, 9.435±0.200°, 11.168±0.200°, 12.060±0.200°, and 18.126±0.200°.

[0040] In some embodiments of the present invention, the C-type crystal of the compound represented by formula (I) has powder X-ray diffraction spectra expressed in 2θ angles, with values ​​of 4.883±0.200°, 6.334±0.200°, 8.399±0.200°, 8.928±0.200°, 9.435±0.200°, 11.168±0.200°, 12.060±0.200°, and 12.9 It includes at least 12, 13, 14, 15, or 16 diffraction peaks selected from 88±0.200°, 14.821±0.200°, 16.182±0.200°, 18.126±0.200°, 18.545±0.200°, 19.474±0.200°, 20.199±0.200°, 24.294±0.200°, and 25.697±0.200°.

[0041] In some embodiments of the present invention, the powder X-ray diffraction spectrum of the C-type crystal of the compound represented by formula (I) is as follows: 2θ angles: 4.883±0.200°, 6.334±0.200°, 8.399±0.200°, 8.928±0.200°, 9.435±0.200°, 11.168±0.200°, 12.060±0.2 Characteristic diffraction peaks are observed at 00°, 12.988±0.200°, 14.821±0.200°, 16.182±0.200°, 18.126±0.200°, 18.545±0.200°, 19.474±0.200°, 20.199±0.200°, 24.294±0.200°, and 25.697±0.200°.

[0042] In some embodiments of the present invention, the C-type crystal of the compound represented by formula (I) is characterized in that its powder X-ray diffraction spectrum is as shown in Figure 7.

[0043] In some embodiments of the present invention, the peak position, interplane distance, and relative intensity of the diffraction peak in the powder X-ray diffraction spectrum of the C-type crystal of the compound represented by formula (I) are as shown in Table 3. [Table 3]

[0044] In some embodiments of the present invention, the differential scanning calorimetry curve of a C-type crystal of the compound represented by formula (I) has an endothermic peak value at 113.5°C ± 3°C.

[0045] In some embodiments of the present invention, the DSC spectrum of the C-type crystal of the compound represented by formula (I) is as shown in Figure 8.

[0046] In some embodiments of the present invention, the thermogravimetric analysis curve of the C-type crystal of the compound represented by formula (I) shows a weight decrease of 3.52% at 110.0°C ± 3°C and a weight decrease of 10.58% at 150.0°C ± 3°C.

[0047] In some embodiments of the present invention, the TGA spectrum of the C-type crystal of the compound represented by formula (I) is as shown in Figure 9.

[0048] In some embodiments of the present invention, the C-type crystal of the compound represented by formula (I) may exist in the form of a solvate crystal.

[0049] The present invention further provides a method for producing C-type crystals of a compound represented by formula (I), comprising the steps of stirring the compound represented by formula (I) in a mixed solvent of ethyl acetate / n-heptane at room temperature, and further comprising the step of separation.

[0050] In some embodiments of the present invention, the C-type crystal of the compound represented by formula (I) of the present invention is (1) A method in which a compound represented by formula (I) is added to a mixed solvent of ethyl acetate / n-heptane and stirred at room temperature. (2) Method of volatilization at low temperature and concentration under reduced pressure It is manufactured by [company name].

[0051] In some embodiments of the present invention, the solvent ratio of ethyl acetate / n-heptane in step (1) above is preferably 1 / 4 (by volume).

[0052] In some embodiments of the present invention, the low temperature in step (2) above is preferably -20°C.

[0053] The present invention further provides a D-type crystal of the compound represented by formula (I), characterized in that its powder X-ray diffraction spectrum has characteristic diffraction peaks at the following 2θ angles: 9.136±0.200°, 9.908±0.200°, 17.787±0.200°, 18.329±0.200°, and 24.298±0.200°. [ka]

[0054] In some embodiments of the present invention, the powder X-ray diffraction spectrum of the compound D-type crystal represented by formula (I) has characteristic diffraction peaks at the following 2θ angles: 9.136±0.200°, 9.908±0.200°, 17.787±0.200°, 18.329±0.200°, and 24.298±0.200°.

[0055] In some embodiments of the present invention, the XRPD spectrum of the D-type crystal of the compound represented by formula (I) is as shown in Figure 10.

[0056] In some embodiments of the present invention, the peak position, interplane distance, and relative intensity of the diffraction peak in the powder X-ray diffraction spectrum of a D-type crystal of the compound represented by formula (I) are as shown in Table 4. [Table 4]

[0057] In some embodiments of the present invention, the differential scanning calorimetry curve of a D-type crystal of the compound represented by formula (I) has an endothermic peak value at 113.3°C ± 3°C.

[0058] In some embodiments of the present invention, the DSC spectrum of the D-type crystal of the compound represented by formula (I) is as shown in Figure 11.

[0059] In some embodiments of the present invention, the thermogravimetric analysis curve of the D-type crystal of the compound represented by formula (I) shows a weight decrease of 6.59% at 90.0°C ± 3°C and a weight decrease of 15.60% at 150.0°C ± 3°C.

[0060] In some embodiments of the present invention, the TGA spectrum of the D-type crystal of the compound represented by formula (I) is as shown in Figure 12.

[0061] In some embodiments of the present invention, the D-type crystal of the compound represented by formula (I) may exist in the form of a solvate crystal.

[0062] The present invention further provides a method for producing D-type crystals of the compound represented by formula (I), comprising the steps of stirring the compound represented by formula (I) in a mixed solvent of dichloromethane / n-heptane at room temperature, and further comprising the step of separation.

[0063] In some embodiments of the present invention, the D-type crystal of the compound represented by formula (I) of the present invention is (1) A method in which a compound represented by formula (I) is added to a dichloromethane / n-heptane mixed solvent and stirred at room temperature. (2) Method of volatilization at low temperature and concentration under reduced pressure It is manufactured by [company name].

[0064] In some embodiments of the present invention, the solvent ratio of dichloromethane / n-heptane in step (1) above is preferably 1 / 4 (by volume).

[0065] In some embodiments of the present invention, the low temperature in step (2) above is preferably -20°C.

[0066] The present invention further provides an E-type crystal of a compound represented by formula (I), characterized in that its powder X-ray diffraction spectrum has characteristic diffraction peaks at the following 2θ angles: 5.987±0.200°, 9.928±0.200°, 11.998±0.200°, 13.499±0.200°, and 18.044±0.200°. [ka]

[0067] In some embodiments of the present invention, the powder X-ray diffraction spectrum of the compound E-type crystal represented by formula (I) has characteristic diffraction peaks at the following 2θ angles: 5.987±0.200°, 9.928±0.200°, 11.998±0.200°, 13.499±0.200°, 18.044±0.200°, 19.492±0.200°, and 25.173±0.200°.

[0068] In some embodiments of the present invention, the XRPD spectrum of the E-type crystal of the compound represented by formula (I) is as shown in Figure 13.

[0069] In some embodiments of the present invention, the peak position, interplane distance, and relative intensity of the diffraction peak in the powder X-ray diffraction spectrum of the E-type crystal of the compound represented by formula (I) are as shown in Table 5. [Table 5]

[0070] In some embodiments of the present invention, the differential scanning calorimetry curve of the E-type crystal of the compound represented by formula (I) above has an endothermic peak value at 113.7°C ± 3°C.

[0071] In some embodiments of the present invention, the DSC spectrum of the E-type crystal of the compound represented by formula (I) is as shown in Figure 14.

[0072] In some embodiments of the present invention, the thermogravimetric analysis curve of the E-type crystal of the compound represented by formula (I) shows a weight reduction of 13.97% at 150.0°C ± 3°C.

[0073] In some embodiments of the present invention, the TGA spectrum of the E-type crystal of the compound represented by formula (I) is as shown in Figure 15.

[0074] In some embodiments of the present invention, the TGA spectrum of the E-type crystal of the compound represented by formula (I) is as shown in Figure 15.

[0075] The present invention further provides a method for producing E-type crystals of the compound represented by formula (I), comprising the steps of stirring the compound represented by formula (I) in toluene at room temperature, and further comprising the step of separation.

[0076] In some embodiments of the present invention, the E-type crystal of the compound represented by formula (I) of the present invention is (1) A method of adding toluene to the compound represented by formula (I) and stirring at room temperature, (2) Method of volatilization and concentration under reduced pressure It is manufactured by [company name].

[0077] In some embodiments of the present invention, the volatilization temperature in step (2) above is preferably room temperature.

[0078] The present invention further provides a crystalline composition comprising type A, type B, type C, type D, or type E crystals of a compound represented by formula (I), wherein the crystals constitute more than 50%, preferably more than 80%, more preferably more than 90%, and most preferably more than 95% of the weight of the crystalline composition.

[0079] The present invention further provides a pharmaceutical composition comprising a therapeutically effective amount of type A, type B, type C, type D, or type E crystal of a compound represented by formula (I), or the above-mentioned crystalline composition. The pharmaceutical composition of the present invention may or may not contain pharmaceutically acceptable excipients. Furthermore, the pharmaceutical composition of the present invention may further contain one or more other therapeutic agents.

[0080] The present invention further provides a method for treating coronavirus infection, comprising administering a therapeutically effective amount of type A, type B, type C, type D, or type E crystals of the compound represented by formula (I) of the present invention, or the crystalline composition thereof, or the pharmaceutical composition thereof, to an individual in need. The present invention further provides the use of type A crystals, type B crystals, type C crystals, type D crystals, or type E crystals of the compound represented by formula (I), or the crystalline composition thereof, or the pharmaceutical composition thereof, in the manufacture of a pharmaceutical for the treatment of diseases related to coronavirus infection.

[0081] In some embodiments of the present invention, the disease associated with the coronavirus infection is SARS-CoV-2 virus infection.

[0082] [Technical effects] The crystals of the compound of the present invention possess good PK properties and therapeutic effects against the SARS-CoV-2 virus. The crystals are stable, have good hygroscopicity, and are resistant to light and heat.

[0083] [Definitions and explanations] Unless otherwise specified, the following terms and collocations used herein have the following meanings. Unless otherwise specifically defined, any particular collocation or term should be understood as having its ordinary definition, not as uncertain or ambiguous. When a trade name appears herein, it refers to the corresponding product or its active ingredient.

[0084] It is well known in the field of crystallography that, for any given crystal form, the relative intensity of diffraction peaks may change due to preferential orientation caused by factors such as crystal morphology. When preferential orientation is present, the peak intensity changes, but the position of the diffraction peaks in the crystal cannot be changed. Furthermore, it is well known in the field of crystallography that even for any given crystal, there may be slight errors in the position of the peaks. For example, changes in temperature during sample analysis, movement of the sample, or calibration of the instrument may cause the position of the peaks to shift, and the measurement error of the 2θ value may be approximately ±0.2 degrees. Therefore, it is well known to those skilled in the art that this error must be taken into consideration when determining the structure of each crystal.

[0085] DSC measures the transition temperature at which heat is absorbed or released due to a change in crystal structure or crystal melting. For the same crystal of the same compound, the error between the thermal transition temperature and melting point in sequential analysis is usually within approximately 5°C or 3°C, and if a compound has a specific DSC peak or melting point, this means that the DSC peak or melting point is within ±5°C or ±3°C. DSC provides an auxiliary method for distinguishing different crystals. Different crystals can be identified based on their different transition temperature characteristics. It should be noted that in the case of mixtures, their DSC peak or melting point can vary over a wider range. Also, since the melting process of a substance involves decomposition, the melting temperature is related to the heating rate.

[0086] For the same crystal, the occurrence of the TGA weight loss temperature may differ depending on the measuring instrument, measurement method / conditions, etc. There may be an error in the weight loss temperature for any particular crystal, and the error may be approximately ±5°C or approximately ±3°C.

[0087] It should be noted that in the production of drug crystals, during the process of contact between drug molecules and solvent molecules, it is difficult to avoid the formation of eutectic formation between solvent molecules and compound molecules due to external conditions and internal factors, and their persistence in the solid material. Therefore, solvates are formed, specifically including stoichiometric solvates and non-stoichiometric solvates. All of the aforementioned solvates are included within the scope of the present invention.

[0088] The aforementioned "pharmaceutically acceptable excipients" refer to inert substances administered simultaneously with the active ingredient to facilitate the administration of the active ingredient, and include, but are not limited to, any fluidizing agents, sweeteners, diluents, preservatives, dyes / colorants, flavor enhancers, surfactants, wetting agents, dispersants, disintegrants, suspending agents, stabilizers, isotonic agents, solvents, or emulsifiers that are permitted for use in humans or animals (e.g., livestock) as approved by the National Food and Drug Administration.

[0089] The term "crystalline composition" refers to a mixture of crystals of the compound represented by formula (I) of the present invention and other crystals or amorphous materials or other impurities of the compound. For example, a crystalline composition of type A crystals of the compound represented by formula (I) includes not only type A crystals of the compound represented by formula (I), but also other crystals or amorphous materials of the compound represented by formula (I), or other impurities.

[0090] The term "pharmaceutical composition" refers to a mixture of one or more compounds of the present invention or salts thereof with pharmaceutically acceptable excipients. The purpose of the pharmaceutical composition is to facilitate the administration of the compounds of the present invention to living organisms.

[0091] The therapeutic dose of the compound of the present invention may be determined, for example, based on the specific therapeutic use, the method of administration of the compound, the patient's health condition, and the judgment of the prescribing physician. The ratio or concentration of the compound of the present invention in the pharmaceutical composition does not have to be fixed and depends on various factors, including the dosage, chemical properties (e.g., hydrophobicity), and route of administration.

[0092] The term “treatment” means administering a compound or formulation described in the present invention to improve or eliminate a disease or one or more symptoms associated with said disease, and includes: (i) To inhibit a disease or disease state, that is, to prevent its onset. (ii) Alleviation of the disease or disease state, i.e., resolution of the disease or disease state.

[0093] The term “therapeutic dose” means the amount of the compound of the present invention that (i) treats a particular disease, condition, or disorder; (ii) alleviates, improves, or eliminates one or more symptoms of a particular disease, condition, or disorder; or (iii) prevents or delays the onset of one or more symptoms of a particular disease, condition, or disorder as described herein. The amount of the compound of the present invention constituting the “therapeutic dose” varies depending on the compound, the disease condition and its severity, the method of administration, and the age of the mammal being treated, but can be routinely determined by those skilled in the art in light of their knowledge and the contents of this disclosure.

[0094] Unless otherwise specifically required in this invention, the word “comprise,” and its English variations such as “comprises” and “comprising,” shall be interpreted throughout this specification and the subsequent claims in an open-ended and comprehensive sense, i.e., “comprise but not limited to.”

[0095] Throughout this specification, any reference to “one embodiment,” “an embodiment,” “another embodiment,” or “in some embodiments” means that at least one embodiment includes certain reference elements, structures, or features described in that embodiment. Therefore, the phrases “one embodiment,” “an embodiment,” “another embodiment,” or “in some embodiments” appearing in various places throughout this specification do not necessarily all refer to the same embodiment. Furthermore, certain elements, structures, or features may be combined in any suitable manner in one or more embodiments.

[0096] In the specification and appended claims of this invention, the singular noun "one" (corresponding to the English "a," "an," and "the") should be understood to include multiple subjects unless explicitly stated otherwise in the context. Therefore, for example, a reference to a reaction including a "catalyst" includes one catalyst, or two or more catalysts. Furthermore, unless explicitly indicated otherwise in the context, the term "or" should also be understood to generally include "and / or."

[0097] The intermediate compounds of the present invention can be produced by various synthetic methods familiar to those skilled in the art, including, but not limited to, the specific embodiments listed below, embodiments combined with other chemical synthesis methods, and equivalent alternative methods familiar to those skilled in the art. Preferred embodiments include, but are not limited to, the examples of the present invention.

[0098] The chemical reactions of specific embodiments of the present invention are completed in a suitable solvent, which should be suitable for the chemical changes of the present invention and the reagents and materials required therefor. To obtain the compounds of the present invention, it may be necessary for those skilled in the art to modify or select synthesis steps or reaction schemes based on existing embodiments.

[0099] The present invention will be specifically described below with reference to examples, but these examples do not limit the present invention in any way.

[0100] All solvents used in this invention are commercially available and may be used as is without further purification.

[0101] The compounds are named according to the usual naming conventions in this field, or ChemDraw (R) Compounds are named using software, and commercially available compounds are named according to the supplier's catalog.

[0102] Instruments and analytical methods

[0103] 1.1 The present invention relates to the powder X-ray diffraction (X-ray powder diffractometer, XRPD) method. Instrument model: PANalytacal X-ray diffractometer Test method: Approximately 1-2 mg of sample is used for XRPD detection. The detailed XRPD parameters are shown below. X-ray tube: Cu, kα, Kα1(Å): 1.540598, Kα2(Å): 1.544426, Kα2 / Kα1 intensity ratio: 0.50 Tube voltage: 45kV, Tube current: 40mA Divergence slit: 1 / 8° Scanning mode: Continuous Scanning range (°2Theta): 3~40 Scan time per step (s): 46.7 Scanning step size (°2Theta): 0.0263 Test time: ~5 minutes

[0104] 1.2 Differential Thermal Analysis (Differential Scanning Calorimeter, DSC) Method of the Present Invention Instrument model: TA 2500 Differential Scanning Calorimeter Measurement method: Take a sample (~1 mg) in a DSC aluminum crucible and perform the measurement. Heat the sample from 25°C (room temperature) to the installation temperature at a heating rate of 10°C / min under conditions of 50 mL / min N2.

[0105] 1.3 The Thermogravimetric Analysis (TGA) Method of the Present Invention Instrument model: TA 5500 thermogravimetric analyzer Measurement method: Take a sample (~1 mg) in a TGA platinum crucible and perform the measurement. Heat the sample from room temperature to the installation temperature at a heating rate of 10°C / min under conditions of 25 mL / min N2. Or: Thermogravimetric analyzer TGA550 Measurement method: Take a sample (5-10 mg) onto the built-in aluminum plate of a TGA platinum crucible and perform the measurement. Heat the sample from room temperature to 300°C at a heating rate of 10°C / min under conditions of 60 mL / min N2.

[0106] 1.4 Dynamic Vapor Soaping (DVS) Method of the Present Invention Equipment Model: SMS DVS Advantage Dynamic Steam Solubleizer Measurement conditions: Samples (10-30 mg) were placed in a DVS sample pan and the test was performed. The detailed DVS parameters are shown below. Temperature: 25℃ Balance: dm / dt = 0.002% / min (minimum: 10 minutes, maximum: 180 minutes) RH (%) range: 70%RH~95%RH~0%RH~95%RH RH (%) gradient: 10% (90%RH~0%RH~90%RH), 5% (95%RH~90%RH and 90%RH~95%RH). [Brief explanation of the drawing]

[0107] [Figure 1] This is the XRPD spectrum of the Cu-Kα line of a type A crystal of the compound represented by formula (I). [Figure 2] This is the DSC spectrum of the A-type crystal of the compound represented by formula (I). [Figure 3] This is the TGA spectrum of a type A crystal of the compound represented by formula (I). [Figure 4] This is the XRPD spectrum of the Cu-Kα line of a type B crystal of the compound represented by formula (I). [Figure 5] This is the DSC spectrum of a type B crystal of the compound represented by formula (I). [Figure 6] This is the TGA spectrum of a B-type crystal of the compound represented by formula (I). [Figure 7] This is the XRPD spectrum of the Cu-Kα line of a C-type crystal of the compound represented by formula (I). [Figure 8] This is the DSC spectrum of the C-type crystal of the compound represented by formula (I). [Figure 9] This is the TGA spectrum of a C-type crystal of the compound represented by formula (I). [Figure 10] This is the XRPD spectrum of the Cu-Kα line of a D-type crystal of the compound represented by formula (I). [Figure 11] This is the DSC spectrum of a D-type crystal of the compound represented by formula (I). [Figure 12] This is the TGA spectrum of a D-type crystal of the compound represented by formula (I). [Figure 13] This is the XRPD spectrum of the Cu-Kα line of the E-type crystal of the compound represented by formula (I). [Figure 14] This is the DSC spectrum of the E-type crystal of the compound represented by formula (I). [Figure 15] This is the TGA spectrum of the E-type crystal of the compound represented by formula (I). [Figure 16] This is the DVS spectrum of the A-type crystal of the compound represented by formula (I). [Modes for carrying out the invention]

[0108] The present invention will be described in detail below with reference to examples, but this does not mean that there are any unfavorable limitations to the present invention. The present invention has been described in detail herein, and specific embodiments thereof have also been disclosed, and it will be apparent to those skilled in the art that various changes and modifications can be made to specific embodiments of the present invention without departing from the spirit and scope of the invention.

[0109] Example 1: Preparation of Compound 1 [ka]

[0110] Step 1: Synthesis of Compounds 1-2 Compound 1-1 (5 g, 54.32 mmol) was dissolved in methanol (50 mL) and refluxed at 70°C for 48 hours. The reaction system was concentrated under reduced pressure to obtain the crude target product. The crude product was of high purity and was used directly in the next step to obtain compound 1-2. 1 H NMR (400 MHz, CDCl3) δ = 4.81 (s, 1H), 3.77 (s, 3H), 3.43 (s, 3H).

[0111] Step 2: Synthesis of Compounds 1-3 Compounds 1-2 were dissolved in toluene (3 mL), cooled to 0°C, and compound (R)-(+)phenethylamine (1.5 g, 12.38 mmol, 1.60 mL) was slowly added dropwise. The mixture was stirred at 20°C for 1 hour. Ethyl acetate (60 mL) and saturated saline (30 mL) were added to the reaction system for extraction. The organic phase was separated, dried over anhydrous sodium sulfate, and spin-dried to obtain the crude product. The crude product was purified by silica gel column chromatography (petroleum ether:ethyl acetate = 1:0-5:1) to obtain target compounds 1-3. 1 H NMR (400 MHz, CDCl3) δ = 7.95~7.56 (m, 1H), 7.31~7.17 (m, 5H), 4.71~4.40 (m, 1H), 3.95~3.71 (m, 3H), 1.67~1.51 (m, 3H).

[0112] Step 3: Synthesis of Compounds 1-4 Compounds 1-3 (0.5 g, 2.61 mmol) were dissolved in 2,2,2-trifluoroethanol (5 mL), trifluoroacetic acid (313.04 mg, 2.75 mmol, 203.28 μL) was added, and the mixture was cooled to -10°C and stirred for 1 hour. The temperature was then controlled to -10°C, and cyclopentadiene (207.40 mg, 3.14 mmol) was slowly added dropwise, with stirring continued for 0.5 hours. The reaction system was concentrated under reduced pressure, methyl tert-butyl ether (60 mL) and saturated sodium bicarbonate solution (30 mL x 2) were added, and the mixture was stirred for 10 minutes. The organic phase was extracted, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. Compounds 1-4 were purified by silica gel column chromatography (petroleum ether:ethyl acetate = 1:0~5:1), and their stereochemistry was confirmed by two-dimensional NMR. 1H NMR (400 MHz, CDCl3) δ = 7.34~7.18 (m, 5H), 6.59~6.41 (m, 1H), 6.31 (dd, J = 1.6, 5.6 Hz, 1H), 4.35 (br d, J = 1.3 Hz, 1H), 3.39 (s, 3H), 3.18~3.03 (m, 1H), 2.95 (br s, 1H), 2.33~2.22 (m, 1H), 2.14 (br d, J = 8.4 Hz, 1H), 1.54~1.41 (m, 4H), MS m / z(ESI):[M+H] + = 258.2.

[0113] Step 4: Synthesis of Compounds 1-5 Compound 1-4 (100.00 mg, 388.61 μmol) was dissolved in tetrahydrofuran (1.25 mL), cooled to -70°C, and boranetetrahydrofuran complex (1 M, 427.47 μL) was slowly added dropwise. The mixture was slowly heated to 20°C and stirred for 1 hour. The mixture was cooled to 0°C, and a 10% sodium hydroxide aqueous solution (0.55 mL) and a 30% hydrogen peroxide solution (220.28 mg, 1.94 mmol, 186.68 μL) were added. The mixture was slowly heated to 20°C and stirred for 1 hour. A saturated sodium thiosulfate aqueous solution (10 mL) was added to the reaction system and stirred for 10 minutes to quench the reaction. Then, saturated saline solution (20 mL) and ethyl acetate (60 mL x 2) were added for extraction, and the organic phase was separated. A small amount of the sample solution was taken, the pH was adjusted to less than 8 with 3% citric acid, and after a potassium iodide starch test showed a negative result, it was dried over anhydrous sodium sulfate and concentrated under reduced pressure at 30°C. Compounds 1-5 were obtained by purification using silica gel column chromatography (petroleum ether:ethyl acetate = 1:0~5:1). 11H NMR (400 MHz, CDCl3) δ = 7.30~7.13 (m, 5H), 3.93 (br d, J = 6.5 Hz, 1H), 3.78 (br s, 1H), 3.70~3.54 (m, 1H), 3.39~3.32 (m, 1H), 3.31~3.24 (m, 3H), 2.49~2.40 (m, 1H), 2.26 (s, 1H), 2.09~2.00 (m, 1H), 1.72 (br d, J = 10.1 Hz, 1H), 1.46 (br d, J = 6.5 Hz, 1H), 1.41~1.33 (m, 3H), MS m / z(ESI):[M+H] + =276.1。

[0114] Step 5: Synthesis of hydrochloride salt of Compound 1-6 Compound 1-5 (3 g, 10.90 mmol) was dissolved in ethanol (80 mL), and hydrochloric acid (1.19 g, 32.69 mmol) and wet palladium carbon (15 g, 10.68 mmol) were added. The reaction system was stirred at 20 °C for 16 hours. The reaction solution was filtered through diatomaceous earth and then directly spin-dried to obtain the crude product, hydrochloride salt of Compound 1-6. 1 1H NMR (400 MHz, DMSO-d6) δ = 10.32~9.78 (m, 1H), 8.94~8.43 (m, 1H), 5.51~5.12 (m, 1H), 4.05~3.98 (m, 1H), 3.96~3.86 (m, 2H), 3.83~3.71 (m, 3H), 2.70~2.60 (m, 1H), 2.36~2.20 (m, 1H), 1.92~1.81 (m, 1H), 1.51~1.31 (m, 2H), MS m / z(ESI):[M+H] + =172.0。

[0115] Step 6: Synthesis of Compound 1-8 It should be noted that there are some possible typos in the original text (such as "1H" in the NMR data which might be a mistake). This translation is based on the provided text as accurately as possible.Compound 1-7 (1.87 g, 10.90 mmol) was dissolved in N,N-dimethylformamide (20 mL), and O-(7-azabenzotriazol-1-yl)-N,N,N,N-tetramethyluronium hexafluorophosphate (4.78 g, 12.58 mmol) and diisopropylethylamine (4.34 g, 33.55 mmol) were added. The mixture was stirred for 30 minutes, and then the hydrochloride salt of compound 1-6 (190 mg, 1.12 mmol) was added. The reaction system was stirred at 20°C for 16 hours. Water (15 mL) was added to the reaction mixture, and the mixture was extracted twice with ethyl acetate (60 mL). The organic phases were combined, the organic phases were washed twice with 5% citric acid (30 mL), washed four times with saline solution (20 mL), dried over anhydrous sodium sulfate, filtered, and spin-dried. Compounds 1-8 were obtained by purification using column chromatography (petroleum ether:ethyl acetate = 3:1). 1 H NMR (400 MHz, CDCl3) δ = 5.28~5.16 (m, 1H), 4.50 (br s, 1H), 4.28 (d, J = 9.8 Hz, 1H), 3.92 (s, 1H), 3.74 (s, 3H), 2.81 (s, 1H), 2.67 (s, 1H), 2.17 (br dd, J = 6.1, 12.7 Hz, 1H), 1.99~1.93 (m, 1H), 1.90~1.84 (m, 1H), 1.59 (br d, J = 13.3 Hz, 2H), 1.43 (s, 9H), 1.04 (s, 9H), MS m / z(ESI):[M+H] + = 385.2.

[0116] Step 7: Synthesis of Compounds 1-9 Compound 1-8 (500 mg, 1.30 mmol) was dissolved in acetonitrile (7.5 mL), and 2-iodoxybenzoic acid (976.31 mg, 3.49 mmol) was added. The mixture was stirred at 60°C for 16 hours. The reaction solution was filtered directly through diatomaceous earth and spin-dried. Compound 1-9 was obtained without purification. MS m / z (ESI): [M-55] + = 327.1.

[0117] Step 8: Synthesis of Compounds 1-10 Compound 1-9 (0.7 g, 1.83 mmol) was dissolved in tetrahydrofuran (14 mL), and TEBBE (μ-chloro-μ-methylene[bis(cyclopentadienyl)titanium]dimethylaluminum) reagent (0.5 M, 14.64 mL) was added at 0°C. The mixture was stirred for the primary time at 0°C, and then the temperature was raised to 15°C and stirring continued for 3 hours. The reaction mixture was slowly poured into saturated sodium bicarbonate solution (50 mL), filtered through diatomaceous earth, extracted with ethyl acetate (30 mL x 3), and washed with saturated brine (30 mL x 2). Compound 1-10 was obtained by purification by column chromatography (petroleum ether:ethyl acetate = 5:1). 1 H NMR (400 MHz, DMSO-d6) δ = 6.63~6.54 (m, 1H), 5.21~5.15 (m, 1H), 4.88~4.82 (m, 1H), 4.76~4.68 (m, 1H), 4.25~4.19 (m, 1H), 3.90~3.85 (m, 1H), 3.66~3.61 (m, 3H), 3.19~3.11 (m, 1H), 2.42~2.28 (m, 2H), 1.98~1.92 (m, 1H), 1.61~1.53 (m, 1H), 1.38 (s, 9H), 1.00~0.93 (m, 9H), MS m / z(ESI):[M+H] + = 381.1.

[0118] Step 9: Synthesis of Compounds 1-11 Protected with nitrogen gas, diethylzinc (1M, 13.14 mL) was slowly added to 1,2-dichloroethane (80 mL) at 0°C. The mixture was stirred for 0.25 hours, and diiodomethane (7.04 g, 26.28 mmol, 2.12 mL) was slowly added to the reaction mixture at 0°C, and the mixture was stirred for 0.25 hours. Trifluoroacetic acid (149.84 mg, 1.31 mmol, 97.30 μL) was slowly added to the reaction system, and stirring was continued for 0.5 hours. Compound 1-10 (0.5 g, 1.31 mmol) in 1,2-dichloroethane (5 mL) was added to the reaction system, the temperature was raised to 20°C, and stirring was continued for 12 hours. The reaction system was quenched with saturated sodium bicarbonate solution (200 mL), extracted with dichloromethane (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by preparative HPLC (column type: Phenomenex luna C18 80×0mm×3μm; mobile phase: [H2O(HCl)-acetonitrile]; acetonitrile %: 1%~30%, 7 min) to obtain compounds 1-11. MS m / z(ESI): [M+H] + = 295.2.

[0119] Step 10: Synthesis of Compounds 1-12 Compound 1-11 (0.1 g, 339.69 μmol) was dissolved in 1,4-dioxane (3 mL), then potassium carbonate (187.79 mg, 1.36 mmol) and a solution of di-tert-butyl dicarbonate (111.20 mg, 509.53 μmol, 117.06 μL) in water (1 mL) were added, and the reaction system was stirred at 15 °C for 12 hours. The reaction mixture was poured into water (30 mL), extracted with ethyl acetate (20 mL x 3), dried over anhydrous sodium sulfate, filtered, and concentrated. Compound 1-12 was obtained by purification by column chromatography (petroleum ether:ethyl acetate = 5:1). 1H NMR (400 MHz, DMSO-d6) δ = 12.52~12.16 (m, 1H), 6.56~6.31 (m, 1H), 4.66~4.58 (m, 1H), 4.24~4.18 (m, 1H), 4.02 (s, 1H), 3.36~3.28 (m, 1H), 1.97~1.91 (m, 2H), 1.81~1.73 (m, 2H), 1.66~1.59 (m, 1H), 1.36 (s, 9H), 0.99~0.93 (m, 9H), 0.80~0.70 (m, 1H), 0.64~0.53 (m, 1H), 0.49~0.33 (m, 2H) MS m / z(ESI):[M+H] + =395.2.

[0120] Step 11: Synthesis of Compounds 1-13 Compound 1-12 (88.13 mg, 223.40 μmol) was dissolved in tetrahydrofuran (2 mL) and methanol (0.6 mL), and lithium hydroxide monohydrate (28.12 mg, 670.21 μmol) dissolved in water (0.6 mL) was added. The reaction system was stirred at 15°C for 2 hours. The pH of the reaction system was adjusted to approximately 5 with 3% citric acid, extracted with ethyl acetate (20 mL x 3), dried over anhydrous sodium sulfate, filtered, and concentrated to obtain compound 1-13. MS m / z(ESI):[M+H] + = 381.3.

[0121] Step 12: Synthesis of Compounds 1-15 Compound 1-13 (0.056 g, 148.79 μmol) was dissolved in N,N-dimethylformamide (2 mL), and O-(7-azabenzotriazol-1-yl)-N,N,N,N-tetramethyluronium hexafluorophosphate (84.86 mg, 223.18 μmol) was added to the reaction system, and the reaction system was stirred at 15°C for 0.5 hours. Next, diisopropylethylamine (76.92 mg, 595.16 μmol, 103.67 μL) was added to the reaction mixture, and further, the hydrochloride salt of compound 1-14 (43.26 mg, 208.31 μmol) was dissolved in N,N-dimethylformamide (0.5 mL) solution and added to the reaction system, and the reaction system was stirred at 15°C for 12 hours. The reaction system was diluted with water (20 mL), extracted with ethyl acetate (20 mL x 3), the organic phase was washed with 3% citric acid (20 mL), washed with saturated sodium chloride (20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated to obtain compounds 1-15. 1 H NMR (400 MHz, DMSO-d6) δ = 8.22~8.08 (m, 1H), 7.56 (s, 1H), 7.32~7.16 (m, 1H), 7.01 (s, 1H), 6.51 (br d, J = 9.4 Hz, 1H), 4.60~4.49 (m, 1H), 4.28~4.17 (m, 2H), 4.14 (s, 1H), 3.18~2.98 (m, 2H), 2.47~2.35 (m, 1H), 2.17~2.09 (m, 2H), 1.96~1.84 (m, 2H), 1.76 (br d, J = 11.0 Hz, 1H), 1.71~1.42 (m, 4H), 1.39~1.34 (m, 9H), 0.95 (s, 8H), 0.84~0.77 (m, 1H), 0.73~0.65 (m, 1H), 0.39 (br s, 2H), MS m / z(ESI):[M+H] + = 534.4.

[0122] Step 13: Synthesis of Compounds 1-16 Compound 1-15 (0.02 g, 37.48 μmol) was dissolved in dichloromethane (2 mL), and trifluoroacetic acid (141.02 mg, 1.24 mmol, 91.57 μL) was added to the reaction system. The mixture was stirred at 15°C for 1 hour. The reaction system was directly quenched with sodium bicarbonate solution (10 mL), extracted with dichloromethane (5 mL x 5), the organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain compound 1-16. 1 H NMR (400 MHz, DMSO-d6) δ = 8.37~8.26 (m, 1H), 8.14 (br d, J = 3.6 Hz, 3H), 7.59~7.53 (m, 1H), 7.40~7.32 (m, 1H), 7.05~6.92 (m, 1H), 4.33~4.21 (m, 2H), 3.93 (br d, J = 5.0 Hz, 1H), 3.17~3.08 (m, 1H), 3.06~2.97 (m, 1H), 2.44~2.32 (m, 1H), 2.18~2.08 (m, 2H), 1.96~1.86 (m, 2H), 1.82~1.57 (m, 4H), 1.51~1.40 (m, 1H), 1.04 (s, 9H), 0.95~0.85 (m, 1H), 0.76~0.67 (m, 1H), 0.54~0.34 (m, 2H), MS m / z(ESI):[M+H] + = 434.2.

[0123] Step 14: Synthesis of Compound 1 Compound 1-16 (0.03 g, 69.20 μmol) was dissolved in dichloromethane (1 mL), and trifluoroacetic anhydride (58.13 mg, 276.79 μmol, 38.50 μL) was added to the reaction system. The mixture was stirred at 15°C for 1 hour. The reaction system was directly quenched with sodium bicarbonate solution (10 mL), extracted with dichloromethane (5 mL x 5), the organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was separated by preparative HPLC (column type: Waters Xbridge BEH C18 100 × 30 mm × 10 μm; mobile phase: [H2O(NH4HCO3)-acetonitrile]; acetonitrile %: 10%~50%, 8 min) to obtain compound 1, which was the compound represented by formula (I). 1 H NMR (400 MHz, DMSO-d6) δ = 9.31 (br d, J = 8.0 Hz, 1H), 8.87 (d, J = 8.6 Hz, 1H), 7.66 (s, 1H), 5.01~4.87 (m, 1H), 4.72~4.59 (m, 2H), 4.07 (s, 1H), 3.17~3.10 (m, 1H), 3.09~2.98 (m, 1H), 2.45~2.33 (m, 1H), 2.19~2.05 (m, 3H), 1.85~1.64 (m, 5H), 1.56 (br d, J = 12.0Hz, 1H), 1.20~1.13 (m, 1H), 1.01 (s, 8H), 0.82~0.67 (m, 2H), 0.54~0.46 (m, 1H), 0.45~0.34 (m, 1H), MS m / z(ESI):[M+H] + = 512.2.

[0124] Example 2: Preparation of type A crystals of the compound represented by formula (I) The compound represented by formula (I) (9.4 g, 18.38 mmol) was dissolved in H2O (235 mL) and stirred at 50°C for 48 hours. A white solid was obtained by filtration, and water was removed by concentrated under reduced pressure to obtain type A crystals, which were detected by XRPD.

[0125] Example 3: Preparation of type B crystals of the compound represented by formula (I) A compound represented by formula (I) (15 mg, 29.32 μmol) was added to a glass vial, and 0.5 mL of ethanol / n-heptane (1:4) solvent was added. The sample was left at room temperature and magnetically stirred (1000 rpm) for about 5 days. Then, it was cooled to a low temperature (5°C) while stirring to allow volatilization, and the solid was collected by centrifugation. The solid was concentrated under reduced pressure to obtain type B crystals, which were detected by XRPD.

[0126] Example 4: Preparation of C-type crystals of the compound represented by formula (I) A compound represented by formula (I) (15 mg, 29.32 μmol) was added to a glass vial, and 0.5 mL of ethyl acetate / n-heptane (1:4) solvent was added. The sample was left at room temperature and magnetically stirred (1000 rpm) for about 5 days. Then, it was cooled to a low temperature (-20°C) while stirring to allow volatilization, and the solid was collected by centrifugation. The solid was concentrated under reduced pressure to obtain C-type crystals, which were detected by XRPD.

[0127] Example 5: Preparation of D-type crystals of the compound represented by formula (I) A compound represented by formula (I) (15 mg, 29.32 μmol) was added to a glass vial, and 0.5 mL of dichloromethane / n-heptane (1:4) solvent was added. The sample was left at room temperature and magnetically stirred (1000 rpm) for about 5 days. Then, it was cooled to a low temperature (-20°C) while stirring to allow volatilization, and the solid was collected by centrifugation. The solid was concentrated under reduced pressure to obtain D-type crystals, which were detected by XRPD.

[0128] Example 6: Preparation of E-type crystals of the compound represented by formula (I) A compound represented by formula (I) (15 mg, 29.32 μmol) was added to a glass vial, 0.5 mL of toluene solvent was added, and the mixture was left at room temperature until the sample dissolved into a clear state. Magnetic stirring (1000 rpm) was performed for approximately 5 days, and then the mixture was cooled to a low temperature (-20°C) while stirring, but no precipitate was observed. The mixture was then heated to room temperature to allow volatilization, yielding a white solid. This solid was concentrated under reduced pressure to obtain E-type crystals, which were detected by XRPD.

[0129] Example 7: Study of the hygroscopicity of type A crystals of the compound represented by formula (I) Experimental materials: SMS DVS Advantage Dynamic Steam Soluble Unit Experimental method: 10-30 mg of type A crystals of the compound represented by formula (I) were placed in a DVS sample pan and measured. Experimental results: The DVS spectrum of the A-type crystal of the compound represented by formula (I) was ΔW = 0.114%, as shown in Figure 16. Experimental conclusion: The A-type crystals of the compound represented by formula (I) showed a moisture absorption weight increase of 0.114% at 25°C and 80%RH, indicating little to no hygroscopicity.

[0130] Example 8: Stability study of type A crystals of the compound represented by formula (I) The results of stability tests of type A crystals under different conditions are shown in Table 6. [Table 6] Experimental conclusion: The A-type crystals of the compound of the present invention exhibit good stability under conditions of light irradiation, high temperature, and high humidity.

[0131] Biological test data:

[0132] Experimental Example 1: Evaluation of the in vitro anti-COVID-19 Mpro protease activity of the test compound. 1. Experimental materials: 1.1 Reagents and consumables are as shown in Table 7. [Table 7]

[0133] 1.2 The equipment and brands are as shown in Table 8. [Table 8]

[0134] 2. Experimental method: The compound was dissolved in DMSO and gradient diluted 3-fold at 10 concentration points according to the test concentration using Echo655. Each concentration was added to a 384-well plate with two replication wells. The Mpro protein and substrate were diluted with test buffer (100 mM NaCl, 20 mM Tris-HCl, 1 mM EDTA). The Mpro protein was added to the 384-well test plate and incubated with the compound at room temperature for 30 minutes. Then the substrate was added, resulting in a test concentration of 25 nM for the Mpro protein and 25 μM for the substrate. The mixture was incubated in a 30°C incubator for 60 minutes. Next, the fluorescence signal values ​​at Ex / Em = 340 nm / 490 nm were detected using a microplate reader. Simultaneously, a background well containing the substrate and compound but without the Mpro protein was detected as a control.

[0135] 3. Data Analysis: 1) The inhibition rate was calculated using the following formula: Inhibition rate % = [(Compound - BG 化合物 )-(ZPE-BG ZPE )] / [(HPE-BG HPE )-(ZPE-BG ZPE )] × 100% # HPE: 100% inhibitory control, containing 25 nM Mpro protein + 25 μM substrate + 1 μM GC376 ZPE: No inhibition control, 25 nM Mpro protein + 25 μM substrate, no compound included. Compound: Test compound well contains 25 nM Mpro protein + 25 μM substrate + compound. BG: Background control well, containing 25 μM of substrate + compound, but without Mpro protein. 2) Using GraphPad Prism software, perform log(agonist) vs. response--Variable slope nonlinear fitting analysis on the inhibition rate data (inhibition rate %) of the compound, and determine the IC of the compound. 50 The value was obtained. [Table 9] Conclusion: The compounds of the present invention have good in vitro anti-COVID-19 Mpro protease activity.

[0136] Example 2: Evaluation of the in vitro anticoronavirus activity of a compound using a cytopathic model. 1. The experimental materials are as shown in Tables 10 and 11. [Table 10] [Table 11]

[0137] 1.1 Cells and Viruses MRC5 cells and coronavirus HCoV OC43 were purchased from ATCC. MRC5 cells were cultured in MEM(Sigma) medium supplemented with 10% fetal bovine serum (Excell), 1% biantibody (Hyclone), 1% L-glutamine (Gibco), and 1% non-essential amino acids (Gibco). MEM(Sigma) medium supplemented with 5% fetal bovine serum (Excell), 1% biantibody (Hyclone), 1% L-glutamine (Gibco), and 1% non-essential amino acids (Gibco) was used as the experimental medium.

[0138] 2. Experimental Method [Table 12] Cells were seeded at a predetermined density (Table 12) in a 96-well microplate and cultured overnight in an incubator at 5% CO2 and 37°C. The following day, diluted compounds (8 concentration points, 2 replication wells) were added at 50 μL per well. Next, diluted viruses were added at 100 TCID per well. 5050 μL was added. Cell controls (cells, no compound treatment or viral infection), viral controls (virus-infected cells, no compound treatment), and culture medium controls (culture medium only) were established. The final volume of the experimental culture medium was 200 μL, and the final DMSO concentration in the medium was 0.5% in each case. Cells were cultured for 5 days in an incubator at 5% CO2 and 33°C. Cell viability was detected using the CellTiter Glo (Promega) cell activity detection kit. The conditions for the cytotoxicity experiment were the same as for the antiviral experiment, but there was no viral infection.

[0139] 3. Data Analysis The antiviral activity and cytotoxicity of the compounds were expressed as the percentage inhibition of virus-induced cytopathic effects and the percentage cell viability at different concentrations of the compound. The calculation formula is as follows: Inhibition rate (%) = (Reading in test well - Average value of virus control) / (Average value of cell control - Average value of virus control) × 100 Cell viability (%) = (Reading in test well - Average value of culture medium control) / (Average value of cell control - Average value of culture medium control) × 100 Using GraphPad Prism, a nonlinear fitting analysis was performed on the inhibition rate and cell viability of the compound, and the semi-effective concentration (EC) of the compound was determined. 50 ) and semi-cytotoxic concentration (CC 50 The value was calculated. [Table 13] Conclusion: The compounds of the present invention exhibit good in vitro anticoronavirus activity at the cellular level and are non-cytotoxic.

[0140] Experimental Example 3: Pharmacokinetic Study in Rats In this study, male and female SD rats were selected as test animals. Compound 1 type A crystals were administered as a single intravenous injection at a dose of 2 mg / kg and as a single intragastric administration at a dose of 30 mg / kg. Plasma drug concentrations of the test compound were then quantitatively measured at different time points using LC / MS / MS to evaluate the pharmacokinetic properties of the test drug in rats. The animals were divided into groups of 3 animals / sex, and each group was administered type A crystals of compound 1. Plasma samples were then collected at 0.083 (intravenous injection group only), 0.25, 0.5, 1, 2, 4, 6, 8, and 24 hours after administration, and plasma drug concentrations were measured using LC-MS / MS. The experimental results are shown in Table 14. [Table 14] Experimental conclusion: The compounds of the present invention showed high exposure levels and high bioavailability in plasma.

[0141] Experimental Example 4: Evaluation of the in vivo antiviral effect of a test compound using an infant mouse infection model with coronavirus strain OC43. C57BL / 6J infant mice were infected with a lethal dose of coronavirus via intranasal administration and treated with compound 1 in a solvent (5% DMSO + 40% PEG400 + 55% water) two hours prior to infection. During the experiment, the infant mice's body weight, health status, and survival were monitored daily, and the protective effect of compound 1 at different doses was evaluated. Infant mice in the solvent group continued to lose body weight on day 6 after viral inoculation, and the endpoint survival rate was 0%. When compound 1 (12.5, 25, 50 mpk) was administered for the first time 2 hours before infection, the endpoint survival rates were 87.5%, 100%, and 100%, respectively. The experimental results are shown in Table 15.

[0142] [Table 15] Experimental conclusion: Compound 1 exhibited excellent anti-coronavirus activity in the body and showed a good dose-response relationship.

Claims

1. A crystal, wherein the crystal is a crystal of type A, type B, type C, type D, or type E of a compound represented by formula (I), 【Chemistry 1】 (1) The type A crystal has a powder X-ray diffraction spectrum that includes characteristic diffraction peaks at the following 2θ angles: 10.655±0.200°, 11.988±0.200°, 16.055±0.200°, 18.356±0.200° and 20.083±0.200°; (2) The type B crystal has a powder X-ray diffraction spectrum that includes characteristic diffraction peaks at the following 2θ angles: 9.202±0.200°, 13.394±0.200°, 14.212±0.200°, 15.516±0.200° and 19.917±0.200°; (3) The C-type crystals have powder X-ray diffraction spectra that include characteristic diffraction peaks at the following 2θ angles: 6.334±0.200°, 8.399±0.200°, 9.435±0.200°, 12.060±0.200° and 18.126±0.200°; (4) The type D crystals have powder X-ray diffraction spectra that include characteristic diffraction peaks at the following 2θ angles: 9.136±0.200°, 9.908±0.200°, 17.787±0.200°, 18.329±0.200° and 24.298±0.200°; (5) The E-type crystals have powder X-ray diffraction spectra that include characteristic diffraction peaks at the following 2θ angles: 5.987±0.200°, 9.928±0.200°, 11.998±0.200°, 13.499±0.200° and 18.044±0.200°; crystal.

2. (1) The type A crystal contains one, two, or three additional diffraction peaks selected from 6.917 ± 0.200°, 14.481 ± 0.200° and 17.653 ± 0.200° in its powder X-ray diffraction spectrum expressed at a 2θ angle; (2) The type B crystal contains one, two or three additional diffraction peaks selected from 10.968 ± 0.200°, 16.483 ± 0.200° and 18.306 ± 0.200° in a powder X-ray diffraction spectrum expressed at a 2θ angle; (3) The C-type crystal contains one, two, or three additional diffraction peaks selected from 4.883 ± 0.200°, 8.928 ± 0.200°, and 11.168 ± 0.200° in its powder X-ray diffraction spectrum expressed at a 2θ angle; The crystal according to claim 1.

3. (1) The type A crystal has a powder X-ray diffraction spectrum that includes characteristic diffraction peaks at the following 2θ angles: 6.917±0.200°, 10.655±0.200°, 11.988±0.200°, 14.481±0.200°, 16.055±0.200°, 17.653±0.200°, 18.356±0.200° and 20.083±0.200°; (2) The type B crystal contains characteristic diffraction peaks at the following 2θ angles: 9.202±0.200°, 10.968±0.200°, 13.394±0.200°, 14.212±0.200°, 15.516±0.200°, 16.483±0.200°, 18.306±0.200° and 19.917±0.200°; (3) The C-type crystal contains characteristic diffraction peaks at the following 2θ angles: 4.883±0.200°, 6.334±0.200°, 8.399±0.200°, 8.928±0.200°, 9.435±0.200°, 11.168±0.200°, 12.060±0.200° and 18.126±0.200°; (4) The E-type crystal contains characteristic diffraction peaks at the following 2θ angles: 5.987±0.200°, 9.928±0.200°, 11.998±0.200°, 13.499±0.200°, 18.044±0.200°, 19.492±0.200° and 25.173±0.200°; The crystal according to claim 1.

4. (1) The type A crystal contains one to eleven additional diffraction peaks selected from the following in its powder X-ray diffraction spectrum, expressed at a 2θ angle: 6.917±0.200°, 13.837±0.200°, 14.481±0.200°, 17.653±0.200°, 20.839±0.200°, 21.388±0.200°, 22.379±0.200°, 24.577±0.200°, 25.104±0.200°, 26.402±0.200° and 31.540±0.200°; (2) The type B crystal, in a powder X-ray diffraction spectrum expressed at a 2θ angle, contains one to eleven additional diffraction peaks selected from: 5.459±0.200°, 7.096±0.200°, 10.968±0.200°, 16.483±0.200°, 18.306±0.200°, 19.488±0.200°, 20.681±0.200°, 21.526±0.200°, 22.409±0.200°, 24.622±0.200° and 25.661±0.200°; (3) The C-type crystal contains one to eleven additional diffraction peaks selected from 4.883±0.200°, 8.928±0.200°, 11.168±0.200°, 12.988±0.200°, 14.821±0.200°, 16.182±0.200°, 18.126±0.200°, 18.545±0.200°, 19.474±0.200°, 20.199±0.200°, 24.294±0.200° and 25.697±0.200° in its powder X-ray diffraction spectrum expressed at a 2θ angle; The crystal according to claim 1.

5. (1) The powder X-ray diffraction spectra of the type A crystals are as follows (2θ angles): 6.917±0.200°, 10.655±0.200°, 11.988±0.200°, 13.837±0.200°, 14.481±0.200°, 16.055±0.200°, 17.653±0.200°, 18.356± It includes characteristic diffraction peaks at 0.200°, 20.083±0.200°, 20.839±0.200°, 21.388±0.200°, 22.379±0.200°, 24.577±0.200°, 25.104±0.200°, 26.402±0.200° and 31.540±0.200°; (2) The powder X-ray diffraction spectra of the type B crystals are as follows (2θ angles): 5.459±0.200°, 7.096±0.200°, 9.202±0.200°, 10.968±0.200°, 13.394±0.200°, 14.212±0.200°, 15.516±0.200°, 16.483±0. It includes characteristic diffraction peaks at 200°, 18.306±0.200°, 19.488±0.200°, 19.917±0.200°, 20.681±0.200°, 21.526±0.200°, 22.409±0.200°, 24.622±0.200° and 25.661±0.200°; (3) The C-type crystals have powder X-ray diffraction spectra that include characteristic diffraction peaks at the following 2θ angles: 4.883±0.200°, 6.334±0.200°, 8.399±0.200°, 8.928±0.200°, 9.435±0.200°, 11.168±0.200°, 12.060±0.200°, 12.988±0.200°, 14.821±0.200°, 16.182±0.200°, 18.126±0.200°, 18.545±0.200°, 19.474±0.200°, 20.199±0.200°, 24.294±0.200° and 25.697±0.200°; The crystal according to claim 1.

6. (1) The differential scanning calorimetry curve of the type A crystal has a peak value of endothermic peak at 173.5°C ± 3°C; (2) The differential scanning calorimetry curve of the type B crystal has a peak value of endothermic peak at 128.4°C ± 3°C; (3) The differential scanning calorimetry curve of the C-type crystal has a peak value of endothermic peak at 113.3°C ± 3°C; (4) The differential scanning calorimetry curve of the type D crystal has a peak value of the endothermic peak at 113.3°C ± 3°C; (5) The differential scanning calorimetry curve of the E-type crystal has a peak value of the endothermic peak at 113.7°C ± 3°C; The crystal according to claim 1.

7. (1) The thermogravimetric analysis curve of the type A crystal shows a 0.00% decrease in weight at 150.0°C ± 3°C; (2) The thermogravimetric analysis curve of the B-type crystal showed a weight decrease of 9.77% at 150.0°C ± 3°C; (3) The thermogravimetric analysis curve of the C-type crystal showed a weight decrease of 3.52% at 110.0°C ± 3°C and a weight decrease of 10.58% at 150.0°C ± 3°C; (4) The thermogravimetric analysis curve of the D-type crystal showed a weight decrease of 6.59% at 90.0°C ± 3°C and a weight decrease of 15.60% at 150.0°C ± 3°C; (5) The thermogravimetric analysis curve of the E-type crystal shows a 13.97% decrease in weight at 150.0°C ± 3°C; The crystal according to claim 1.

8. (1) The type B crystals exist in the form of solvate crystals; (2) The C-type crystals exist in the form of solvate crystals; (3) The D-type crystals exist in the form of solvate crystals; (4) The E-type crystals exist in the form of solvate crystals; The crystal according to claim 1.

9. A crystalline composition comprising the crystal described in any one of claims 1 to 8.

10. A pharmaceutical composition comprising the crystal described in any one of claims 1 to 8.

11. A pharmaceutical composition comprising the crystalline composition described in claim 9.

12. A crystal according to any one of claims 1 to 8 for use in the treatment of coronavirus infection.

13. The crystal according to claim 12, wherein the coronavirus infection is SARS-CoV-2 virus infection.

14. The crystalline composition according to claim 9 for use in the treatment of coronavirus infection.

15. The crystalline composition according to claim 14, wherein the coronavirus infection is SARS-CoV-2 virus infection.

16. A pharmaceutical composition according to claim 10 for use in the treatment of coronavirus infection.

17. The pharmaceutical composition according to claim 16, wherein the coronavirus infection is SARS-CoV-2 virus infection.

18. A pharmaceutical composition according to claim 11 for use in the treatment of coronavirus infection.

19. The pharmaceutical composition according to claim 18, wherein the coronavirus infection is SARS-CoV-2 virus infection.

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