Crystal of spiro compound and method for producing the same

Abstract

The present disclosure relates to crystalline forms of spiro compounds of formula (I), their methods of preparation, and their use in the treatment of diseases caused by coronavirus infections and / or small RNA virus infections. TIFF2026502598000031.tif43170

Description

[Technical Field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims priority to and the benefit of Chinese patent application No. 202310064780.5, filed with the State Intellectual Property Office of China on January 16, 2023, the entire disclosure of which is incorporated herein by reference.

[0002] The present disclosure relates to the field of medicinal chemistry, and in particular to crystalline forms of spiro compounds, methods for their preparation, and pharmaceutical compositions and uses containing them. [Background technology]

[0003] Coronaviruses are single-stranded, sense-RNA viruses, some of which can spread widely within populations and cause severe symptoms. Currently, there are seven known coronaviruses capable of infecting humans: HCoV-229E, HCoV-OC43, HCoV-NL63, HCoV-HKU1, SARS-CoV, MERS-CoV, and SARS-CoV-2. Most of the functional proteins of coronaviruses are encoded by the ORF1ab gene, which is first translated into a single polyprotein and then cleaved into multiple active proteins by 3CL proteases and PL proteases. Therefore, inhibiting the activity of 3CL proteases can effectively inhibit viral replication. Different coronavirus 3CL proteases share a high degree of structural homology; therefore, 3CL protease inhibitors typically possess broad-spectrum anticoronavirus activity.

[0004] In addition to coronaviruses, 3CL protease also plays an important role in hydrolyzing polyproteins encoded by small RNA viruses, and 3CL protease inhibitors can effectively inhibit the replication of small RNA viruses. Enterovirus 71 is a small RNA virus that is one of the common viruses that causes hand, foot, and mouth disease and can also cause various diseases such as meningitis, brainstem encephalitis, and myocarditis. In recent years, enterovirus 71 has caused multiple outbreaks in infants and young children, and clinically, there is still a lack of effective therapeutic agents.

[0005] Therefore, there remains a clinical need for the development of pharmaceutical active substances useful for inhibiting RNA / small RNA viruses, including coronaviruses, enterovirus 71, and the like. Summary of the Invention

[0006] In one aspect, the disclosure provides a crystal of the compound of formula (I). [ka]

[0007] In another aspect, the present disclosure provides crystalline Form A of the compound of formula (I): The X-ray powder diffraction pattern of the A-type crystals, expressed in terms of diffraction angle 2θ, has diffraction peaks at 10.88±0.20°, 15.09±0.20°, 17.67±0.20°, 18.28±0.20° and 20.64±0.20°.

[0008] In a further aspect, the present disclosure provides a method for preparing crystalline Form A of the compound of formula (I), comprising: The present invention provides a method for producing a compound of formula (I) by crystallizing the compound of formula (I) with solvent (i) and solvent (ii), followed by separating the resulting solid, wherein the solvent (i) is at least one selected from isopropyl acetate, ethyl acetate, and isopropyl alcohol, and the solvent (ii) is at least one selected from n-hexane and n-heptane.

[0009] In another aspect, the disclosure provides a solvate of a compound of formula (I): The solvate is selected from a methyl tert-butyl ether solvate and a 2-methyltetrahydrofuran solvate.

[0010] In yet another aspect, the present disclosure provides crystalline Form B of a methyl tert-butyl ether solvate of the compound of Formula (I), The X-ray powder diffraction pattern of the B-type crystals, expressed in terms of diffraction angle 2θ, has diffraction peaks at 6.26±0.20°, 17.66±0.20°, and 20.24±0.20°.

[0011] In a further aspect, the present disclosure provides a method for preparing Type B crystals, comprising mixing the compound of formula (I) with methyl tert-butyl ether, stirring, and then separating the solid.

[0012] In another aspect, the present disclosure provides crystalline Form C of the 2-methyltetrahydrofuran solvate of the compound of Formula (I): The X-ray powder diffraction pattern of the C-type crystals, expressed in terms of diffraction angle 2θ, has diffraction peaks at 6.21±0.20°, 17.48±0.20°, and 20.86±0.20°.

[0013] In yet another aspect, the present disclosure provides a method for producing Form C crystals, the method comprising mixing the compound of Formula (I) with 2-methyltetrahydrofuran, stirring, and separating the solid after crystallization.

[0014] In a further aspect, the present disclosure provides a drug combination comprising crystalline Form A, crystalline Form B, crystalline Form C, or a combination thereof, of the compound of Formula (I) described herein and another antiviral agent.

[0015] In another aspect, the present disclosure provides a pharmaceutical composition comprising Form A crystals, Form B crystals, Form C crystals, or a combination thereof, of the compound of Formula (I) described herein, and pharmaceutically acceptable adjuvants.

[0016] In yet another aspect, the present disclosure provides use of Form A crystals, Form B crystals, Form C crystals, or a combination thereof of the compound of formula (I) described herein, a drug combination described herein, or a pharmaceutical composition described herein, in the manufacture of a medicament for preventing or treating an associated disease caused by coronavirus and / or small RNA virus infection. [Brief explanation of the drawings]

[0017] [Figure 1] 1 is an XRPD pattern of crystalline form C of the 2-methyltetrahydrofuran solvate of the compound of formula (I). [Figure 2] 1 is a DSC pattern of type C crystals of the 2-methyltetrahydrofuran solvate of the compound of formula (I). [Figure 3] 1 is an XRPD pattern of type B crystals of the methyl tert-butyl ether solvate of the compound of formula (I). [Figure 4] 1 is a DSC pattern of type B crystals of the methyl tert-butyl ether solvate of the compound of formula (I). [Figure 5] 1 is an XRPD pattern of crystalline form A of the compound of formula (I). [Figure 6] 1 is a DSC pattern of type A crystals of the compound of formula (I). [Figure 7] 1 is a TGA pattern of type A crystals of the compound of formula (I). [Figure 8] 1A and 1B are graphs showing the inhibitory effect of the compound of formula (I) on the virus titer in mouse lungs 2 days (FIG. A) and 4 days (FIG. B) after infection in Test Example 4. FIG. [Figure 9] FIG. 1 is a graph showing changes in mouse body weight in Test Example 4. [Figure 10] 1 is a graph showing the inhibitory effect of the compound of formula (I) on the viral titer in mouse brain 4 days after infection in Test Example 4. FIG. DETAILED DESCRIPTION OF THE INVENTION

[0018] The compound of formula (I) is a novel 3CL protease inhibitor having broad-spectrum anti-coronavirus activity, and the compound of formula (I) has the following structure: [ka]

[0019] The present disclosure provides a crystal of the compound of formula (I).

[0020] The present disclosure provides a type A crystal of the compound of formula (I), and the X-ray powder diffraction pattern of the type A crystal, expressed in terms of diffraction angle 2θ, has diffraction peaks at 10.88±0.20°, 15.09±0.20°, 17.67±0.20°, 18.28±0.20°, and 20.64±0.20°.

[0021] In some embodiments, the X-ray powder diffraction pattern of the A-type crystal of the compound of formula (I), expressed in terms of diffraction angles 2θ, has diffraction peaks at 10.88±0.20°, 15.09±0.20°, 16.61±0.20°, 17.67±0.20°, 18.28±0.20°, 18.50±0.20°, 20.09±0.20°, and 20.64±0.20°.

[0022] In some embodiments, the X-ray powder diffraction pattern of the A-type crystal of the compound of formula (I), expressed in terms of diffraction angles 2θ, has diffraction peaks at 10.27±0.20°, 10.88±0.20°, 11.86±0.20°, 15.09±0.20°, 16.61±0.20°, 17.67±0.20°, 18.28±0.20°, 18.50±0.20°, 20.09±0.20°, and 20.64±0.20°.

[0023] In some embodiments, the X-ray powder diffraction pattern of the A-type crystal of the compound of formula (I), represented by diffraction angles 2θ, is 9.29±0.20°, 10.27±0.20°, 10.88±0.20°, 10.97±0.20°, 11.86±0.20°, 14.27±0.20°, 14.92±0.20°, 15.09±0.20°. It has diffraction peaks at 15.61±0.20°, 15.78±0.20°, 16.61±0.20°, 17.67±0.20°, 18.28±0.20°, 18.50±0.20°, 20.09±0.20°, 20.64±0.20°, 22.18±0.20°, 23.80±0.20° and 25.63±0.20°.

[0024] In some embodiments, the X-ray powder diffraction pattern of the A-type crystal of the compound of formula (I) above, expressed in terms of diffraction angle 2θ, has the diffraction peaks shown in Table 3.

[0025] In some embodiments, the Form A crystal of the compound of formula (I) has an X-ray powder diffraction pattern, expressed in terms of diffraction angles 2θ, substantially as shown in FIG.

[0026] In some embodiments, the Type A crystals of the compound of formula (I) have a DSC pattern with a peak at 213.78°C ± 5.0°C.

[0027] In some embodiments, the DSC pattern of the Form A crystals of the compound of formula (I) is substantially as shown in FIG.

[0028] The present disclosure provides a method for preparing crystalline Form A of the compound of formula (I), comprising: The present invention further provides a method, which comprises mixing the compound of formula (I) with solvent (i) and solvent (ii), crystallizing, and separating the solid, wherein the solvent (i) is at least one selected from isopropyl acetate, ethyl acetate, and isopropyl alcohol, and preferably isopropyl acetate, and the solvent (ii) is at least one selected from n-hexane and n-heptane, and preferably n-heptane.

[0029] The method of mixing the compound of formula (I) with solvent (i) and solvent (ii) includes at least a specific method of first mixing the compound of formula (I) with solvent (i) and then further mixing with solvent (ii), or mixing the compound of formula (I) with a mixed solvent of solvent (i) and solvent (ii).

[0030] In some embodiments, in the method for producing the A-type crystal, the volume (mL) of the solvent (i) is 1 to 100 times, preferably 2 to 50 times the mass (g) of the compound.

[0031] In some embodiments, in the method for producing the Type A crystal, the volume (mL) of the solvent (ii) is 2 to 100 times, preferably 5 to 50 times, the mass (g) of the compound.

[0032] In some embodiments, the crystallization in the method for producing Type A crystals is carried out at a temperature of 10 to 80°C, preferably 15 to 45°C, and more preferably 20 to 30°C.

[0033] In some embodiments, in the method for producing Type A crystals, the separating step comprises filtering, collecting the solid, and drying.

[0034] The present disclosure further provides a solvate of a compound of formula (I) selected from a methyl tert-butyl ether solvate and a 2-methyltetrahydrofuran solvate.

[0035] In some embodiments, the molar ratio of the compound of formula (I) to the solvent in the solvate of the compound of formula (I) is about 0.5 to 2.

[0036] In some embodiments, in the solvate of the compound of formula (I), the molar ratio of the compound of formula (I) to the solvent is about 0.8 to 1.2.

[0037] In some embodiments, in the solvate of the compound of formula (I), the molar ratio of the compound of formula (I) to the solvent is about 1.0.

[0038] The present disclosure further provides a type B crystal of the methyl tert-butyl ether solvate of the compound of formula (I), wherein the X-ray powder diffraction pattern of the type B crystal, expressed in terms of diffraction angles 2θ, has diffraction peaks at 6.26±0.20°, 17.66±0.20°, and 20.24±0.20°.

[0039] In some embodiments, the X-ray powder diffraction pattern of the B-type crystals, expressed in terms of diffraction angles 2θ, has diffraction peaks at 6.26±0.20°, 7.27±0.20°, 11.47±0.20°, 13.06±0.20°, 15.57±0.20°, 17.66±0.20°, 20.24±0.20°, and 22.72±0.20°.

[0040] In some embodiments, the X-ray powder diffraction pattern of the B-type crystals, expressed in terms of diffraction angles 2θ, has diffraction peaks at 6.26±0.20°, 7.27±0.20°, 8.84±0.20°, 10.18±0.20°, 10.30±0.20°, 11.47±0.20°, 13.06±0.20°, 14.44±0.20°, 15.57±0.20°, 17.66±0.20°, 20.24±0.20°, 20.51±0.20°, and 22.72±0.20°.

[0041] In some embodiments, the X-ray powder diffraction pattern of the B-type crystals, expressed in terms of diffraction angle 2θ, has the diffraction peaks shown in Table 2.

[0042] In some embodiments, the Type B crystals have an X-ray powder diffraction pattern, expressed in terms of diffraction angles 2θ, substantially as shown in FIG.

[0043] In some embodiments, the Type B crystals have a DSC pattern with peaks at 93.36°C±5.0°C, 170.25°C±5.0°C, and 214.18°C±5.0°C.

[0044] In some embodiments, the DSC pattern of the B-type crystals is substantially as shown in FIG.

[0045] The present disclosure provides a method for producing B-type crystals, comprising: There is further provided a process comprising combining a compound of formula (I) with methyl tert-butyl ether and stirring, followed by separating the solids.

[0046] In some embodiments, in the method for producing the B-type crystals, the volume (mL) of the methyl tert-butyl ether is 2 to 100 times, preferably 5 to 50 times, and more preferably 10 to 30 times the mass (g) of the compound of formula (I).

[0047] In some embodiments, in the method for producing the B-type crystals, the stirring step is carried out at 15 to 55°C, preferably 45 to 55°C, and more preferably 50 to 55°C.

[0048] In some embodiments, in the method for producing the type B crystals, the compound of formula (I) is a 2-methyltetrahydrofuran solvate of the compound of formula (I).

[0049] The present disclosure further provides a C-type crystal of the 2-methyltetrahydrofuran solvate of compound of formula (I), wherein the X-ray powder diffraction pattern of the C-type crystal, expressed in terms of diffraction angles 2θ, has diffraction peaks at 6.21±0.20°, 17.48±0.20°, and 20.86±0.20°.

[0050] In some embodiments, the X-ray powder diffraction pattern of the C-type crystals, expressed in terms of diffraction angle 2θ, has diffraction peaks at 6.21±0.20°, 7.04±0.20°, 11.71±0.20°, 17.48±0.20°, 20.59±0.20°, and 20.86±0.20°.

[0051] In some embodiments, the X-ray powder diffraction pattern of the C-type crystals, expressed in terms of diffraction angles 2θ, has diffraction peaks at 6.21±0.20°, 7.04±0.20°, 11.71±0.20°, 14.74±0.20°, 15.77±0.20°, 17.48±0.20°, 20.59±0.20°, and 20.86±0.20°.

[0052] In some embodiments, the X-ray powder diffraction pattern of the C-type crystals, expressed in terms of diffraction angle 2θ, has the diffraction peaks shown in Table 1.

[0053] In some embodiments, the Form C crystal has an X-ray powder diffraction pattern, expressed in terms of diffraction angles 2θ, substantially as shown in FIG.

[0054] In some embodiments, the C-type crystals have a DSC pattern with peaks at 90.39°C±5.0°C, 167.59°C±5.0°C, and 214.22°C±5.0°C.

[0055] In some embodiments, the DSC pattern of the C-type crystals is substantially as shown in FIG.

[0056] The present disclosure provides a method for producing a C-type crystal, comprising: There is further provided a process comprising mixing a compound of formula (I) with 2-methyltetrahydrofuran, stirring, crystallizing, and then isolating the solid.

[0057] In some embodiments, in the method for producing the C-type crystal, the volume (mL) of the 2-methyltetrahydrofuran is 2 to 50 times, preferably 3 to 20 times, and more preferably 5 to 15 times the mass (g) of the compound of formula (I).

[0058] In some embodiments, in the method for producing the C-type crystals, the stirring step is carried out at a temperature of 15 to 80°C, preferably 40 to 80°C, for example, 50 to 60°C or 70 to 80°C.

[0059] In some embodiments, in the method for producing C-type crystals, the crystallization is temperature-reducing crystallization.

[0060] In some embodiments, in the method for producing a C-type crystal, the temperature of the reaction system is lowered to 0 to 35°C, preferably 5 to 30°C, and more preferably 10 to 25°C in the temperature lowering step.

[0061] In another embodiment, the present disclosure further provides a drug combination comprising the above-described crystalline form A, crystalline form B, crystalline form C, or a combination thereof of the compound of formula (I), and another antiviral agent.

[0062] In another aspect, the present disclosure provides a pharmaceutical composition comprising the Type A crystals, Type B crystals, Type C crystals or a combination thereof of the compound of formula (I) above, and pharmaceutically acceptable auxiliary materials, optionally further comprising another antiviral agent.

[0063] In some embodiments, the other antiviral agent is ritonavir.

[0064] In another aspect, the present disclosure provides use of Type A crystals, Type B crystals, Type C crystals or a combination thereof of the compound of formula (I) above, or the drug combination, or the pharmaceutical composition above, in the manufacture of a medicament for preventing or treating related diseases caused by coronavirus and / or small RNA virus infection.

[0065] In another aspect, the present disclosure provides use of the Type A crystals, Type B crystals, Type C crystals or a combination thereof of the compound of formula (I), or the drug combination or the pharmaceutical composition, in the prevention or treatment of related diseases caused by coronavirus and / or small RNA virus infection.

[0066] In another aspect, the present disclosure provides crystalline form A, crystalline form B, crystalline form C, or a combination thereof of the compound of formula (I), or the drug combination, or the pharmaceutical composition, for preventing or treating related diseases caused by coronavirus and / or small RNA virus infection.

[0067] In another aspect, the present disclosure provides a method for treating a coronavirus and / or related disease caused by a small RNA virus infection, comprising administering to a subject (e.g., a mammal, preferably a human) in need of such treatment a therapeutically effective amount of the Form A crystals, Form B crystals, Form C crystals, or a combination thereof of the compound of formula (I) above, or the drug combination or the pharmaceutical composition.

[0068] Associated diseases caused by coronavirus and / or small RNA virus infections described in this disclosure include, but are not limited to, respiratory infections, pneumonia, or complications thereof.

[0069] The coronavirus described herein is selected from SARS-CoV, MERS-CoV, H229E-CoV, HKU1-CoV, NL63-CoV, OC43-CoV, or SARS-CoV-2. The small RNA virus described herein is selected from Enterovirus 71.

[0070] The type A crystals, type B crystals, or type C crystals of the compound of formula (I) described in the present disclosure have at least one advantage in terms of pharmacological efficacy, physicochemical properties, etc., and are suitable for preparation into a desired pharmaceutical composition.

[0071] Definitions and explanations of terms Unless otherwise specified, the terms used in this disclosure have the following meanings, and the definitions of groups and terms described in this disclosure, including their illustrative definitions, exemplary definitions, preferred definitions, definitions described in the tables, definitions of specific compounds in the examples, etc., can be arbitrarily combined or combined with each other. Unless specifically defined, a particular term should not be considered indefinite or unclear, but should be understood according to its general meaning in the art. When a trade name appears in this specification, it is intended to refer to the corresponding trade name or its active ingredient.

[0072] The term "solvate" refers to a complex or aggregate formed by one or more molecules of a solute and one or more molecules of a solvent. A solvate typically has an essentially fixed molar ratio of solute to solvent. The term also includes clathrate compounds, including clathrate compounds with water. Representative solvents include, for example, water, methanol, ethanol, isopropyl alcohol, acetic acid, 2-methyltetrahydrofuran, methyl tert-butyl ether, and the like. When the solvent is water, the solvate formed is a hydrate.

[0073] The term "about" is used in this disclosure to mean approximately, on the order of, roughly, or around. When the term "about" is used in conjunction with a numerical range, it modifies that range by expanding the limits of the numerical range set forth. Unless otherwise specified, the term "about" is used herein to modify the upper and lower limits of a set forth value by a numerical value of 10% deviation.

[0074] Unless otherwise specified, the terms "comprise", "containing" or "comprise" and their English variants, such as comprises or comprising, are to be understood in an open and non-exclusive sense, i.e., "including but not limited to".

[0075] "Alternative embodiments" or "embodiments" as referred to in this disclosure refer to at least one embodiment that includes the relevant specific referenced elements, structures, or features described in that embodiment. Thus, the appearances of the phrase "alternative embodiments" or "embodiments" in different places in this disclosure do not necessarily all refer to the same embodiment. Furthermore, specific elements, structures, or features may be combined in any suitable manner in one or more embodiments.

[0076] Room temperature as referred to in this disclosure refers to 20±5.0°C.

[0077] A range "m to n" as used in this disclosure represents a shorthand notation for combining any real numbers from m to n, where both m and n are real numbers.

[0078] The "X-ray powder diffraction pattern" described in this disclosure is obtained by measurement using CuKα radiation.

[0079] The term "X-ray powder diffraction pattern or XRPD pattern" as used herein refers to the Bragg equation, 2d Sinθ=nλ (where d is the crystallographic spacing, θ is the diffraction angle, λ is the wavelength of the incident X-ray, and the diffraction order n is any positive integer, generally the first-order diffraction peak is taken, where n=1), where the Bragg equation can be satisfied when X-rays are incident on a crystal or on a certain atomic plane having a d-spacing in some crystal sample at a sweep angle θ (the complementary angle of the incident angle, also called the Bragg angle), thereby measuring this set of X-ray powder diffraction patterns.

[0080] For the same crystalline form of the same compound, the peak positions in the XRPD pattern are similar throughout, and the relative intensity error may be relatively large. In addition, when identifying an adulterant, it is necessary to note that it is not necessary to rely on all the diffraction peaks observed in a high-purity sample, since some diffraction lines may be missing due to factors such as a decrease in content, and even one diffraction peak may be characteristic of a given crystal.

[0081] "2θ or 2θ angle" as used in this disclosure refers to the diffraction angle, where θ is the Bragg angle, and is measured in ° or degrees.

[0082] Those skilled in the art will understand that due to factors such as crystal defects and measurement errors, the molar ratios of the compounds disclosed herein and the acid / base molecules and solvent molecules in solvates will always have some error, and generally, ±10% is within a reasonable error range. Although there will be some error variation depending on the context of where it is used, the error variation is ±10% or less, preferably ±5% or less.

[0083] The term "therapeutically effective amount" refers to a dose of a compound of the present disclosure that (i) treats a particular disease, condition, or disorder, (ii) reduces, ameliorates, or eliminates one or more symptoms of a particular disease, condition, or disorder, or (iii) delays the onset of one or more symptoms of a particular disease, condition, or disorder described herein. The amount of a compound of the present disclosure that constitutes a "therapeutically effective amount" varies depending on the compound, the disease state and its severity, the mode of administration, and the age of the mammal being treated, but can be determined routinely by one of ordinary skill in the art according to their own knowledge and the present disclosure.

[0084] The term "treatment" means administering a compound or formulation described herein to ameliorate or eliminate a disease or one or more symptoms associated with said disease; and (i) inhibiting the disease or disease state, i.e., inhibiting its progression; (ii) alleviating the disease or disease state, i.e., causing the disease or disease state to regress; Includes.

[0085] The term "prevention" refers to the administration of a compound or formulation described herein to prevent a disease or one or more symptoms associated with said disease, and This includes preventing the appearance of a disease or disease state in a subject (e.g., a mammal), particularly where such subject is susceptible to the disease state but has not been diagnosed as suffering from the disease state.

[0086] As used herein, the term "subject" or "patient" includes mammals and non-mammals. Examples of mammals include, but are not limited to, any member of the class Mammalia, including humans, non-human primates (e.g., chimpanzees and other apes and monkeys), livestock such as cows, horses, sheep, goats, and pigs, pet animals such as rabbits, dogs, and cats, and laboratory animals including rodents such as rats, mice, and guinea pigs. Examples of non-human mammals include, but are not limited to, birds and fish.

[0087] The term "pharmaceutically acceptable" refers to compounds, materials, compositions and / or dosage forms that are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response or other problem or complication, commensurate with a reasonable benefit / risk ratio.

[0088] The term "pharmaceutically acceptable auxiliary material" refers to an auxiliary material that does not have a significant irritating effect on the organism and does not impair the biological activity and performance of the active compound. Suitable auxiliary materials are well known to those skilled in the art, such as carbohydrates, waxes, water-soluble and / or water-swellable polymers, hydrophilic or hydrophobic materials, gelatin, oils, solvents, water, etc.

[0089] The term "drug combination" refers to a combination of two or more active ingredients. In some embodiments of the present disclosure, the active ingredients in the drug combination may be administered simultaneously, and in some embodiments of the present disclosure, the active ingredients in the drug combination may be administered separately or sequentially.

[0090] The pharmaceutical compositions of the present disclosure can be prepared by combining the compounds of the present disclosure or their medicinal salts or solvates with suitable pharmaceutically acceptable auxiliary materials, and can be prepared into solid, semi-solid, liquid or gaseous preparations such as tablets, pills, capsules, powders, granules, ointments, emulsions, suspensions, suppositories, injections, inhalants, gels, microspheres and aerosols.

[0091] Typical routes of administration of the compounds of the present disclosure or their pharmaceutically acceptable salts or solvates, or pharmaceutical compositions comprising said compounds or their pharmaceutically acceptable salts or solvates, include, but are not limited to, oral, rectal, topical, inhalation, parenteral, sublingual, intravaginal, intranasal, intraocular, intraperitoneal, intramuscular, subcutaneous, and intravenous administration.

[0092] The pharmaceutical compositions of the present disclosure can be manufactured by conventional mixing, dissolving, granulating, emulsifying, lyophilizing or other methods well known in the art.

[0093] In some embodiments, the pharmaceutical composition is in oral dosage form.For oral administration, the pharmaceutical composition can be prepared by mixing the active compound with pharmaceutically acceptable auxiliary materials well known in the art.With these auxiliary materials, the compound of the present disclosure or its medicinal salt or its solvate can be prepared into tablets, pills, lozenges, dragees, capsules, liquids, gels, slurries, suspensions, etc. for oral administration to patients.

[0094] The solid oral compositions can be prepared by conventional blending, filling or tabletting methods, for example by mixing the active compound with a solid auxiliary material, optionally comminute the resulting mixture, optionally adding other suitable auxiliary materials as needed, and then processing the mixture into granules to give tablets or dragee cores.

[0095] The pharmaceutical compositions are also applicable for parenteral administration, such as sterile solutions, suspensions, or lyophilized products in appropriate unit dosage forms.

[0096] The therapeutically effective amount of the crystalline form of the compound of formula (I) or a solvate thereof contained in the pharmaceutical composition of the present disclosure is selected from 0.001 mg / kg body weight to 1000 mg / kg body weight, for example, 0.01 mg / kg body weight to 500 mg / kg body weight, either in the form of a single or separate dose.

[0097] Those skilled in the art will recognize that measurement data of XRPD peak positions and / or intensities for a given crystalline form of the same compound will vary within an error range. The 2θ values ​​in this disclosure encompass an appropriate error range, and the error range is typically expressed as "±." For example, in this disclosure, a 2θ value expressed as a specific angle value of ±0.20° indicates that the error variation range of the specific angle value is ±0.20°, i.e., a 2θ of 5.92±0.20° indicates that 2θ is within the range of 6.12 to 5.72°. Depending on sample preparation techniques, calibration techniques applied to the instrument, human operating variations, etc., those skilled in the art will recognize that an appropriate error range for XRPD diffraction angles may be ±0.20°, ±0.15°, ±0.10°, ±0.05°, or less, and that a certain degree of variability in peak intensities is acceptable. When used to describe an XRPD pattern, the terms "substantially the same" or "substantially shown in" refer to a pattern that contains diffraction peaks for which at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, or at least 99% of the diffraction angles are within a standard deviation of ±0.2° 2θ.

[0098] Those skilled in the art will recognize that measurement data for DSC patterns of a given crystalline form of the same compound will vary within acceptable error limits. The peak value (expressed in degrees Celsius) of a single peak allows for an appropriate error range. Typically, the error range is expressed as "±." For the same crystalline form of the same compound, the thermal transition temperature and melting point error in consecutive analyses is typically within ±5.0°C. For example, a peak value of "170.25±5.0" indicates a range of 165.25 to 175.25°C. Those skilled in the art will recognize that, depending on sample preparation techniques, calibration techniques applied to the instrument, human operating variance, etc., an appropriate error range for the peak value of a single peak may be ±5.0, ±4.0, ±3.0, ±2.0, or even less.

[0099] The salt forms and / or crystalline forms of the present disclosure may also be isotopically labeled. The present disclosure further includes isotopically labeled compounds of the present disclosure that are the same as those described herein, except that one or more atoms are replaced with an atom having an atomic mass or mass number different from the atomic mass or mass number normally found in nature. Illustrative examples of isotopes that can be bound to compounds of the present disclosure include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, sulfur, fluorine, iodine, and chlorine, for example, 2 H, 3 H, 11 C. 13 C. 14 C. 13 N, 15 N, 15 O. 17 O. 18 O. 31 P, 32 P, 35 S, 18 F, 123 I, 125 I and 36 Cl, etc.

[0100] Some isotopically labeled compounds of the present disclosure (e.g., 3 H and 14 C) may be used in compound and / or substrate tissue distribution analysis. 3 H) and carbon-14 (i.e. 14 C) Isotopes are particularly preferred for their ease of preparation and detectability. 15 O. 13 N, 11 C and 18 Positron-emitting isotopes, such as F, may be used in positron emission tomography (PET) studies to measure substrate occupancy. Isotopically labeled compounds of the present disclosure can generally be prepared in the following schemes and / or procedures similar to those disclosed in the Examples by substituting isotopically labeled reagents for non-isotopically labeled reagents.

[0101] Note that relatively heavy isotopes (e.g., deuterium (i.e. 2Substitution with deuterium may provide several therapeutic advantages (e.g., increased in vivo half-life or reduced dosage requirements) resulting from greater metabolic stability and may therefore be preferable in some cases, where deuterium substitution may be partial or complete, with partial deuterium substitution referring to the replacement of at least one hydrogen with deuterium.

[0102] The compounds of the present disclosure or their pharmaceutically acceptable salts or solvates thereof can be prepared by a variety of synthetic methods well known to those skilled in the art, including the specific embodiments listed below, embodiments formed by combining them with other chemical synthetic methods, and equivalent substitution forms well known to those skilled in the art, and preferred embodiments include, but are not limited to, the examples of the present disclosure.

[0103] The chemical reactions of the specific embodiments of the present disclosure are completed in a suitable solvent, which must be suitable for the chemical transformations of the present disclosure and the necessary reagents and materials. To obtain the compounds of the present disclosure or their pharmaceutically acceptable salts or solvates, those skilled in the art may need to modify or select synthetic steps or reaction processes based on existing embodiments.

[0104] Test conditions for the equipment used in the experiments of this disclosure: 1. X-ray powder diffraction Model number: Bruker D8 Focus X-ray light source:Cu Kα Kα1(Å): 1.54060, Kα2(Å): 1.54439, Kα2 / Kα1 intensity ratio: 0.50 Wavelength λ (Å): 1.54060 Slit (°): 2.5 Scanning method: θ / 2θ, Scanning range: 3 to 40° (2θ angle) Residence time (sec): 0.12 Scan step (°2θ): 0.01 Scanning speed: 5° / min Voltage: 40kV Current: 40mA 2. Differential scanning calorimeter Device model number:Discvery DSC2500 Purge gas: Nitrogen gas Sample disc: Aluminum disc, non-sealing lid holder Method: Linear heating Heating rate: 10℃ / min Temperature range: 30℃~300℃ or 30℃~400℃, 3, Thermogravimetric analyzer Device model number:Discvery TA 55 Purge gas: Nitrogen gas Sample disc: platinum, aperture Method: Linear heating Heating rate: 10℃ / min Temperature range: 30℃~300℃ 4. Dynamic moisture absorption device Device model number: DVS Intrinsic PLUS DVS parameters: Temperature: 25℃, Equilibrium: dm / dt=0.002% / min RH(%) test step: 10% RH(%) test step range: 0%~90%~0%.

[0105] The compounds of the present disclosure can be prepared by a variety of synthetic methods well known to those skilled in the art, including the specific embodiments listed below, embodiments formed by combining them with other chemical synthetic methods, and equivalent substitution forms well known to those skilled in the art, and preferred embodiments include, but are not limited to, the examples of the present disclosure.

[0106] The chemical reactions of specific embodiments of the present disclosure are completed in a suitable solvent, which must be suitable for the chemical transformations of the present disclosure and the necessary reagents and materials. To obtain compounds of the present disclosure, one skilled in the art may need to modify or select synthetic steps or reaction processes based on existing embodiments.

[0107] Specific Embodiments The present invention will be described in detail below with reference to examples, but these examples are not intended to limit the present disclosure in any way. Although the present specification has already described the present disclosure in detail and disclosed specific examples, it will be apparent to those skilled in the art that various modifications and improvements can be made to the specific embodiments of the present disclosure without departing from the spirit and scope of the present disclosure. All reagents used in the present disclosure are commercially available and can be used without further purification.

[0108] Unless otherwise specified, the ratio of the mixed solvent is a volumetric mixing ratio. Unless otherwise specified, % refers to wt%.

[0109] Compounds are named artificially or with ChemDraw® software; commercially available compounds use names from the manufacturer's catalog.

[0110] The structures of the compounds were determined by nuclear magnetic resonance (NMR) and / or mass spectrometry (MS). NMR shifts are in units of 10 -6 The solvents used for NMR measurements were deuterated dimethyl sulfoxide, deuterated chloroform, deuterated methanol, etc., and the internal standard was tetramethylsilane (TMS).

[0111] Example 1: Preparation of compounds of formula (I) 1.1. Preparation of Compound 1-1: [ka] Step 1: The starting material SMA (2.74 g, 11.85 mmol), 35 mL of dichloromethane, and 35 mL of DMF were added to a reaction flask and cooled to 0 °C. The starting materials SMB (3.56 g, 11.86 mmol), benzotriazol-1-yloxytris(dimethylamino)phosphonium hexafluorophosphate (BOP, 6.29 g, 14.22 mmol), and N-methylmorpholine (NMM, 3.91 mL, 35.56 mmol) were added sequentially. The temperature was raised to room temperature and the reaction was allowed to proceed for 10 h. After completion of the reaction, an appropriate amount of dichloromethane was added, and the organic phase was washed sequentially with 1 N aqueous hydrochloric acid and saturated brine. After washing, the organic phase was dried over anhydrous sodium sulfate, concentrated to dryness, and purified by column chromatography to obtain 3.71 g of INT-1. ESI-MS: m / z 433.2 [M+H] + ; 1 H NMR (400 MHz, DMSO-d):δ H : 6.75 (d, J = 9.2 Hz, 1H), 4.38 (t, J = 8.2 Hz, 1H), 4.25 (d, J = 10.9 Hz, 1H), 4.11 (d, J = 9.3 Hz, 1H), 3.93 (t, J = 9.3 Hz, 1H), 3.62 (s, 3H), 3.40-3.31 (m, 4H), 2.70 (dd, J = 13.1, 7.9 Hz, 1H), 2.37 (dd, J = 13.2, 8.4 Hz, 1H), 1.37 (s, 9H), 0.94 (s, 9H).

[0112] Step 2: INT-1 (3.71 g, 8.58 mmol), 37 mL of THF, 37 mL of purified water, and lithium hydroxide monohydrate (0.72 g, 17.16 mmol) were added to a reaction flask and reacted at room temperature for 2 h. After the reaction was completed, the pH was adjusted to 4 with concentrated hydrochloric acid and filtered to obtain 3.4 g of compound 1-1. ESI-MS: 419.2 m / z [M+H] + ; 1 H NMR (400 MHz, DMSO-d):δ H:12.68 (s, 1H), 6.71 (d, J = 9.4 Hz, 1H), 4.38-4.19 (m, 2H), 4.11 (d, J = 9.4 Hz, 1H), 3.88 (d, J = 10.9 Hz, 1H), 3.41-3.29(m, 4H), 2.69 (dd, J = 13.1, 7.9 Hz, 1H), 2.34 (dd, J = 13.2, 8.9 Hz, 1H), 1.38 (s, 9H), 0.94 (s, 9H).

[0113] 1.2. Preparation of Compound 1-2: [ka] Ammonia gas-methanol solution (700 mL, 7 mol / L) and the starting material SMD (100 g, 0.349 mol) were added to the reaction flask, stirred to dissolve, and then incubated at 25±5°C for 36 hours. After the reaction was complete, the reaction mixture was concentrated to approximately 250 mL. 300 mL of isopropyl alcohol was added, and the mixture was concentrated under reduced pressure (repeated three times) until the remaining reaction mixture was approximately 250 mL. The mixture was then purged with nitrogen gas and cooled to 10±5°C. 500 mL of hydrogen chloride-isopropyl alcohol solution (4 mol / L) was added to the reaction vessel. After the addition, the mixture was heated to 25±5°C, maintained at 25±5°C, and incubated for 9 hours. After the reaction was completed, the reaction solution was concentrated under reduced pressure until the volume of the remaining reaction solution was approximately 250 mL, 300 mL of isopropyl alcohol was added, and the concentration under reduced pressure was continued until the volume of the remaining reaction solution was approximately 250 mL (repeated twice), 100 mL of isopropyl alcohol was added, and the mixture was stirred for 30±5 minutes, filtered, and the filter cake was rinsed with 50 mL of isopropyl alcohol to obtain a wet product, which was dried under vacuum at 45±5°C to obtain 66.7 g of compound 1-2. 1 H NMR (400 MHz, DMSO-d):δ H:8.45 (d, J = 5.1 Hz, 3H), 8.25-8.04 (m, 1H), 7.95 (s, 1H), 7.67-7.49 (m, 1H), 3.85-3.80 (m, 1H), 3.19-3.13 (m, 2H), 2.59-2.51 (m, 1H), 2.32-2.27 (m, 1H), 2.05-1.98 (m, 1H), 1.82-1.66 (m, 2H);ESI-MS: 172.1 m / z [M+H] + .

[0114] 1.3. Preparation of compounds of formula (I): [ka] Step 1: Compound 1-1 (419 mg, 1 mmol) was placed in a two-neck flask and, under nitrogen gas protection, 5 mL of dichloromethane was added, followed by 2-(7-azabenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (400 mg, 1.1 mmol). The reaction mixture was stirred at room temperature for 1 h. Compound 1-2 (1 mmol) was dissolved in 1 mL of dichloromethane and added to the above system. N,N-diisopropylethylamine (2 mmol) was then added in an ice-water bath. The ice-water bath was removed, and the system was stirred at room temperature overnight. After workup, 50 mL of dichloromethane was added, followed by washing three times with 1 M aqueous hydrochloric acid, three times with saturated aqueous sodium bicarbonate, and the organic phase with saturated brine. The mixture was dried over anhydrous sodium sulfate, filtered, and rotary evaporated to give compound 1-3 (469 mg). ESI-MS: m / z 572.3 [M+H] + .

[0115] Step 2: Compound 1-3 (572 mg, 1 mmol) was dissolved in 3 mL of 4 M hydrogen chloride / 1,4-dioxane solution and stirred at ambient temperature. After thin-layer chromatography showed that the reaction was essentially complete, the solvent was thoroughly spun off. The resulting crude product was dissolved in 2 mL of dichloromethane and protected with nitrogen gas. Triethylamine (3 mmol) was added, and the system was placed in an ice-water bath. Trifluoroacetic anhydride (1.2 mmol) was added dropwise. After thin-layer chromatography showed that the reaction was essentially complete, 50 mL of dichloromethane was added, followed by washing three times with 1 M aqueous hydrochloric acid, three times with saturated aqueous sodium bicarbonate, and the organic phase with saturated brine. The organic phase was then dried over anhydrous sodium sulfate and purified by column chromatography to obtain compound 1-4 (265 mg). ESI-MS: m / z 568.3 [M+H] + .

[0116] Step 3: Compound 1-4 (113 mg, 0.2 mmol) and Burgess reagent (1.5 eq) were added to a two-neck flask, which was filled and vented with nitrogen three times. Dichloromethane dried over molecular sieves was added and the mixture was stirred at room temperature overnight. Thin layer chromatography showed that the reaction was essentially complete. After workup, column chromatography afforded compound (I) (41 mg), which was confirmed to be amorphous by XRPD. 1H NMR (400 MHz, DMSO-d6) δ 9.46 (d, J = 8.7 Hz, 1H), 9.05 (d, J = 8.6 Hz, 1H), 7.67 (s, 1H), 4.97 (ddd, J = 11.0, 8.5, 5.0 Hz, 1H), 4.53 (d, J = 8.7 Hz, 1H), 4.34 (dd, J = 9.9, 7.1 Hz, 1H), 4.26-4.14 (m, 1H), 3.92 (d, J = 10.9 Hz, 1H), 3.50-3.34 (m, 4H), 3.22-3.11 (m, 1H), 3.06 (td, J = 9.3, 7.1 Hz, 1H), 2.68-2.58 (m, 1H), 2.50-2.43 (m, 1H), 2.31 (dd, J = 13.0, 10.0 Hz, 1H), 2.23-2.07 (m, 2H), 1.71 (tdd, J = 14.9, 10.3, 7.4 Hz, 2H), 0.99 (s, 9H). ESI-MS: 550.3 m / z [M+H] + .

[0117] Example 2: Preparation of Type C Crystals of 2-methyltetrahydrofuran Solvate of Compound of Formula (I) Compound 1-4 (10.00 g) was dissolved in 100 mL of dichloromethane, 8.39 g of Burgess reagent was added, and the mixture was incubated at 25-26 °C for 1 hour. After the reaction was completed, the mixture was quenched with 50 mL of saturated aqueous sodium bicarbonate solution. The organic phase was washed with 50 mL of 1 M aqueous hydrochloric acid solution and 50 mL of saturated aqueous sodium chloride solution, and then concentrated under reduced pressure to obtain the compound of formula (I). The obtained compound of formula (I) was added to 100 mL of 2-methyltetrahydrofuran, heated to 70-80 °C, dissolved into a clear solution, gradually cooled to allow crystallization, and filtered to obtain 7.92 g of a solid. The obtained solid was identified as type C crystal of the 2-methyltetrahydrofuran solvate of the compound of formula (I) by methods such as nuclear magnetism, TGA, and X-ray powder diffraction pattern. The XRPD pattern of type C crystal is shown in FIG. 1, its DSC pattern is shown in FIG. 2, and its XRPD diffraction peak positions are shown in Table 1 below.

[0118] TIFF2026502598000007.tif94170

[0119] 1 H NMR(400 MHz, DMSO-d6) δ 9.47 (d, J = 7.6 Hz, 1H), 9.06 (d, J = 8.6 Hz, 1H), 7.68 (s, 1H), 4.99-4.93(m, 1H), 4.52 (d, J = 7.0 Hz, 1H), 4.32 (dd, J = 9.9Hz, 7.1 Hz, 1H), 4.19 (d, J = 11.0 Hz, 1H), 3.91 (d, J = 10.9 Hz, 1H), 3.79-3.86(m,1H),3.78-3.71(m,1H),3.58-3.51(m,1H),3.47-3.35 (m, 4H), 3.14 (t, J = 9.3 Hz, 1H), 3.08-3.02 (m, 1H), 2.61 (dd, J = 13.0Hz, 7.1 Hz, 1H), 2.46-2.39 (m, 1H), 2.35-2.25 (m, 1H), 2.18-2.07 (m, 2H), 2.00-1.75(m,3H),1.75-1.67 (m, 2H),1.35-1.27 (m, 1H), 1.12(d, J=6.1Hz,3H), 0.97 (s, 9H).

[0120] Example 3: Preparation of Type B Crystals of Methyl tert-butyl Ether Solvate of Compound of Formula (I) 1.6 kg of the 2-methyltetrahydrofuran solvate of the compound of formula (I) was dissolved in 4.5 L of dichloromethane and concentrated under reduced pressure until dry. The resulting solid was pulverized and added in portions to 45 L of methyl tert-butyl ether. After the addition was complete, the temperature was raised to 50-55°C and slurried for 1 hour. The temperature was then gradually lowered to 20°C, filtered, and dried to obtain 1.41 kg of a solid. The resulting solid was identified as type B crystals of the methyl tert-butyl ether solvate of the compound of formula (I) by nuclear magnetism, TGA, and X-ray powder diffraction pattern. The XRPD pattern of type B crystals is shown in Figure 3, its DSC pattern is shown in Figure 4, and its XRPD diffraction peak positions are shown in Table 2 below.

[0121] TIFF2026502598000008.tif110170

[0122] 1H NMR (400 MHz, DMSO-d6): δ 9.48 (d, J = 8.7 Hz, 1H), 9.06 (d, J = 8.6 Hz, 1H), 7.68 (s, 1H), 5.00-4.94 (m, 1H), 4.53 (d, J = 8.7 Hz, 1H), 4.33 (dd, J = 9.8 Hz, 7.1 Hz, 1H), 4.19 (d, J = 10.7 Hz, 1H), 3.92 (d, J = 10.9 Hz, 1H), 3.44-3.37 (m, 4H), 3.17-3.02 (m, 2H), 3.08 (s, 3H), 2.61 (dd, J = 12.8 Hz, 7.4 Hz, 1H), 2.46-2.43 (m, 1H), 2.30 (dd, J = 12.9 Hz, 10.1 Hz, 1H), 2.18-2.08 (m, 2H), 1.75-1.65 (m, 2H), 1.11 (s, 9H), 0.98 (s, 9H).

[0123] Example 4: Preparation of Form A Crystals of the Compound of Formula (I) Method 1: 270 g of the 2-methyltetrahydrofuran solvate of the compound of formula (I) was added to 540 mL of a mixed solvent of isopropyl acetate and 5.4 L of n-heptane, heated to 55-65°C, slurried and crystallized for 12 hours, gradually cooled to 20-30°C, stirred at the same temperature for 1 hour, filtered, and dried to obtain 215 g of a solid. The resulting solid was identified as type A crystal of the compound of formula (I) by nuclear magnetic resonance and X-ray powder diffraction patterns. The XRPD pattern of type A crystal is shown in Figure 5, its DSC pattern in Figure 6, and its TGA pattern in Figure 7, and its XRPD diffraction peak positions are shown in Table 3 below.

[0124] TIFF2026502598000009.tif187170

[0125] 1 H NMR (400 MHz, DMSO-d6):δ 9.47 (d, J = 8.4 Hz, 1H), 9.06 (d, J = 8.6 Hz, 1H), 7.68 (s, 1H), 5.00-4.94 (m, 1H), 4.53 (d, J = 8.5 Hz, 1H), 4.33 (dd, J = 9.8 Hz, 7.1 Hz, 1H), 4.19 (d, J = 10.8 Hz, 1H), 3.92 (d, J = 11.0 Hz, 1H), 3.44-3.32 (m, 4H), 3.17-3.02 (m, 2H), 2.61 (dd, J = 12.7 Hz, 5.6 Hz, 1H), 2.50-2.43 (m, 1H), 2.30 (dd, J = 12.8 Hz, 10.0 Hz, 1H), 2.19-2.08 (m, 2H), 1.75-1.68 (m, 2H), 0.98 (s, 9H).

[0126] Method 2: 10 mg of the methyl tert-butyl ether solvate of the compound of formula (I) was dissolved in 50 μL of ethyl acetate, and 50 μL of n-heptane was added thereto. The solution became cloudy, and after the solvent was evaporated, the resulting solid was identified as type A crystal of the compound of formula (I) based on the X-ray powder diffraction pattern.

[0127] Method 3: 15 mg of the methyl tert-butyl ether solvate of the compound of formula (I) was dissolved in 150 μL of isopropyl alcohol, 400 μL of n-heptane was added, the solution became cloudy, stirring was continued for 2 hours, 40 μL of n-heptane was added, the solution became cloudy, and after evaporating the solvent, the obtained solid was identified as type A crystal of the compound of formula (I) based on the X-ray powder diffraction pattern.

[0128] Biological activity and related property test examples Test Example 1-1: Inhibitory activity test of the compound of formula (I) against SARS-CoV-2 3CLpro Using fluorescence resonance energy transfer (FER) to detect SARS-CoV-2 3CL proThe inhibitory activity of the compound of formula (I) against enzyme activity was evaluated. The total volume of the enzyme reaction system was 120 μL, the final concentration of protease was 30 nM, and the final concentration of substrate was 20 μM. The reaction buffer contained 50 mM Tris pH 7.3 and 1 mM EDTA. SARS-CoV-2 3CL was placed in a 96-well plate. pro Protease and different concentrations of compounds were added, incubated at 30°C for 10 min, and then substrate was added and the plate was quickly read in a microplate reader. The excitation and emission light were 320 nM and 405 nM, respectively. The test time was 3.5 min, and fluorescence readings were taken every 35 s. The final results were fitted to the reaction rate using the readings from the first 2 min and compared with the control (DMSO) to calculate the inhibition rate. Fitting was performed using the software GraphPad Prism 8, and the IC 50 The values ​​and inhibition rate curves were obtained.

[0129] Experimental results show that the compound of formula (I) inhibits SARS-CoV-2 3CL pro IC for 50 The value was <0.1 μM, indicating a strong inhibitory effect.

[0130] Test Example 1-2: Inhibitory activity test of the compound of formula (I) against SARS-CoV-2 Omicron strain mutant 3CL protease Experimental principle: The inhibitory effect of the compound of formula (I) of the present disclosure on the activity of the mutant 3CL protease of the Omicron strain (P132H) was studied using a method in which fluorescence resonance energy transfer (FRET) occurs by reacting an enzyme with a substrate.

[0131] The experimental materials are shown in the table below.

[0132] TIFF2026502598000010.tif51170

[0133] Laboratory equipment and devices: TIFF2026502598000011.tif33170

[0134] Testing Procedure: A reaction buffer containing 20 mM Tris-HCl, 1 mM EDTA, 0.01% BSA, 1 mM DTT, and 100 mM NaCl was prepared. Using the Echo pipette system, test compounds were diluted to different concentrations in dimethyl sulfoxide (DMSO) and transferred to a 384-well plate. Mutant 3CL protease was diluted in the reaction buffer and added to the 384-well plate at 10 μL / well. The mixture was centrifuged at 1000 rpm for 1 min and then incubated at room temperature for 30 min. Next, 10 μL / well of substrate was added and the mixture was centrifuged at 1000 rpm for 30 s to initiate the enzyme reaction. The final enzyme concentration in the reaction system was 50 nM, the final substrate concentration was 20 μM, and the compound concentrations ranged from 10,000 nM to 0.51 nM. Next, the Flexstation 3 microplate reader was set to Kinetic Reduction Vmax mode, and the fluorescence intensity at 490 nm was read every 75 seconds for a total of 35 consecutive readings. The reaction velocity (V) was obtained, and the inhibition rate was calculated. The XLfit software was used to perform a four-parameter fitting to determine the half-maximal inhibitory concentration (IC). 50 The inhibition rate was calculated as follows: Inhibition rate=(V max -V Compound ) / (V max -V min )×100% where V max is the reaction rate value for wells containing only enzyme and substrate, and V min is the reaction rate value for wells containing substrate only, and V Compound was the reaction rate value for the wells containing the test compound, enzyme and substrate.

[0135] Experimental results: The compound of formula (I) still retained significant inhibitory activity against the 3CL protease harboring the P132H mutation in the SARS-CoV-2 Omicron strain.

[0136] TIFF2026502598000012.tif36170

[0137] Test Example 2: Inhibitory activity test of compounds of formula (I) against coronavirus 3CL proteases of different origins Experimental Objective: To study the inhibitory effect of the compound of formula (I) on the activity of 3CL proteases from six other coronaviruses that can infect humans, namely SARS-CoV, MERS-CoV, H229E-CoV, HKU1-CoV, NL63-CoV and OC43-CoV.

[0138] Test materials: 3CL protease: Recombinant full-length coronavirus 3CL proteases were produced in-house according to the coronavirus genome sequences. The GenBank numbers of the SARS-CoV, MERS-CoV, H229E-CoV, HKU1-CoV, NL63-CoV, and OC43-CoV genomes used were AAP13442.1, MT387202.1, AF304460.1, AY597011.2, AY567487.2, and AY903459.1, respectively. The DNA sequences required for protein expression of the six coronavirus 3CL proteases were purchased from Nanjing Jinsirui Biotechnology Co., Ltd.

[0139] 3CL protease substrate was purchased from Nanjing Jinsirui Biotechnology Co., Ltd.

[0140] Chymotrypsin substrate was purchased from Jier Biochemical Co., Ltd.

[0141] Other reagents are listed in the table below.

[0142] TIFF2026502598000013.tif27170

[0143] Testing Procedure: A reaction buffer (containing 50 mM Tris and 1 mM EDTA) was prepared. Test compounds were dissolved in DMSO to a 100 mM stock solution and further diluted two-fold with the reaction buffer to a total of 11 concentrations. 3CL protease and different concentrations of compounds were added to a 96-well plate and incubated at room temperature for 10 minutes. Substrate was then added and the plate was quickly read in a microplate reader. The total volume of the enzyme reaction system was 120 μL. The final concentrations of SARS-CoV, MERS-CoV, H229E-CoV, HKU1-CoV, NL63-CoV, and OC43-CoV proteases were 30 nM, 80 nM, 30 nM, 20 nM, 30 nM, and 10 nM, respectively, and the final substrate concentration was 10 μM. The excitation and emission wavelengths during reading were 340 nm and 490 nm, respectively. The test lasted for 10 minutes, with fluorescence readings taken every minute. The final results were calculated by fitting the reaction rate using the readings from the first 5 minutes to calculate the inhibition rate, which was calculated using the formula: inhibition rate = 1 - (reaction rate of the test group / reaction rate of the control group).

[0144] Experimental Results: As shown in Table 5, the compound of formula (I) exhibited relatively good inhibitory effects against 3CL proteases derived from six other coronaviruses, suggesting that the compound of formula (I) may have broad-spectrum anti-coronavirus activity.

[0145] TIFF2026502598000014.tif66170

[0146] Test Example 3: Inhibitory effect of the compound of formula (I) on SARS-CoV-2 Vero E6 original strain (WIV04), Delta strain (B.1.617.2), and Omicron strain (B.1.1.529) at the cellular level Experimental Objective: This study investigated the inhibitory effect of the compound of formula (I) on the replication of SARS-CoV-2 original strain (WIV04 strain), Delta strain (B.1.617.2), and Omicron strain (B.1.1.529) in Vero E6 cells by detecting viral copy numbers in the culture supernatant using real-time fluorescent quantitative PCR. Because Vero E6 cells highly express the efflux transporter protein P-gp, the compounds were co-incubated with 0.5 μM of the P-gp inhibitor CP-100356.

[0147] Test materials: Vero E6 was purchased from ATCC (product number CRL-1586), and the SARS-CoV-2 original strain (SARS-CoV-2-WIV04 strain), Delta strain (B.1.617.2), and Omicron strain (B.1.1.529) viruses were derived from the Microbial (Virulent) Seed Deposit Center of the Wuhan Institute of Virology, Chinese Academy of Sciences.

[0148] Other reagents are listed in the table below.

[0149] TIFF2026502598000015.tif81170

[0150] Experimental equipment: Biosafety cabinet (AC2-3S1, ESCO, Singapore) Carbon dioxide incubator (Thermo Scientific HERAcell 150i, Thermo Scientific, USA) Pure water equipment (Yuyuan SYS ultrapure water machine, Chengdu) StepOne Plus Real-time PCR system (4376600, ABI, USA) TC20™ Automated Cell Counter (1450102, BIO-RAD, USA) T100™ Thermal Cycler (1861096, BIO-RAD, USA) Centrifuge (Micro21 / 21R Thermo Fisher, Thermo Scientific, USA)

[0151] Testing Procedure: Vero E6 cells were digested with trypsin, placed in 90% DMEM, 10% fetal bovine serum, and seeded at 50,000 cells / well in a 48-well plate for overnight incubation. Test compounds were dissolved in DMSO to a 40 mM stock solution, which was then diluted with 0.5 μM Pgp inhibitor-containing medium to obtain the required concentration. The final test compound concentrations ranged from 1 μM to 0.004 μM. The cell supernatant was removed, and diluted compounds (containing 0.5 μM Pgp inhibitors) were added to each well and incubated for 1 hour. Different strains of SARS-CoV-2 were added at a multiplicity of infection (MOI) of 0.01 or 0.001 in a Biosafety Level 3 (BSL-3) laboratory. After 1 hour of incubation, the supernatant was removed and washed with PBS. 200 μL of diluted compounds (containing 0.5 μM Pgp inhibitors) were added per well. The supernatant was collected at 24 or 72 hours post-infection. Supernatant viral RNA was extracted, and the viral copy number in the supernatant was detected using real-time fluorescent quantitative PCR. The compound inhibition rate was calculated based on the viral copy number, and the compound's IC was calculated using GraphPad Prism 8. 50 was calculated.

[0152] For the cytotoxicity test, Vero E6 cells were digested and placed in medium (90% DMEM, 10% fetal bovine serum), seeded at 20,000 cells / well in a 96-well plate, and cultured overnight. The test compound was dissolved in DMSO to prepare a 40 mM stock solution, which was then gradient-diluted with medium or medium containing 0.5 μM Pgp inhibitor to obtain the required concentration for the test. The final concentration range of the test compound in the experiment was 500 μM to 1.95 μM. The cell supernatant was removed from the 96-well plate, and 100 μL / well of medium containing the test compound (single agent or 0.5 μM Pgp inhibitor) was added. After 24 hours of incubation, the cell activity was detected using a CCK8 detection kit, and the inhibition rate and median cytotoxic concentration (CCT) were measured. 50 ) was calculated.

[0153] Test results: As shown in Table 6, after combined use with the P-gp inhibitor CP-100356, the compound of formula (I) can dose-dependently inhibit the replication of Delta strain in Vero E6 cells, with an IC 50 The IC value was 0.040 μM. In the original strain, the compound of formula (I) in combination with a P-gp inhibitor also exerted a relatively strong inhibitory effect, with an IC value of 0.040 μM. 50 Furthermore, the combination of the compound of formula (I) with a P-gp inhibitor was able to significantly inhibit the replication of Omicron strains in Vero E6 cells, with an IC 50 The compound of formula (I) alone and in combination with a P-gp inhibitor did not show any significant cytotoxicity against the proliferation of Vero E6 cells, and CC 50 >500 μM.

[0154] TIFF2026502598000016.tif90170

[0155] Test Example 4: In vivo antiviral activity of the compound of formula (I) against SARS-CoV-2 Delta strain in hACE2-K18 transgenic mice Experimental Objective: This study evaluated the antiviral activity of compound of formula (I) against SARS-CoV-2 Delta strain in K18 transgenic mice stably expressing human angiotensin-converting enzyme 2 (ACE2) (K18-hACE2).

[0156] Test materials: Seven- to eight-week-old K18-hACE2 transgenic mice were purchased from Jiangsu Jixi Yaokang Biotechnology Co., Ltd. SARS-CoV-2 Delta strain virus was derived from the Microbial (Virulent) Species Depository Center of the Wuhan Institute of Virology, Chinese Academy of Sciences.

[0157] Ritonavir was purchased from Shanghai Disinfectant Chemical Pharmaceutical Co., Ltd.

[0158] Vero E6 cells were purchased from ATCC (product number CRL-1586).

[0159] Other reagents are listed in the table below.

[0160] TIFF2026502598000017.tif76170

[0161] Experimental equipment: Biosafety cabinet (AC2-3S1, ESCO, Singapore) Carbon dioxide incubator (Thermo Scientific HERAcell 150i, Thermo Scientific, USA) Pure water equipment (Yuyuan SYS ultrapure water machine, Chengdu) StepOne Plus Real-time PCR system (4376600, ABI, USA) TC20™ Automated Cell Counter (1450102, BIO-RAD, USA) T100™ Thermal Cycler (1861096, BIO-RAD, USA) Centrifuge (Micro21 / 21R Thermo Fisher, Thermo Scientific, USA) Tissue polishing machine (JXFSTPRP-CL, Shanghai Jingxin, China)

[0162] Experimental Procedure: K18-hACE2 transgenic mice were infected intranasally with the SARS-CoV-2 delta strain (day 0). Two hours after infection, mice were intragastrically administered vehicle, 50 mg / kg, or 200 mg / kg of the compound of formula (I) (combined with 50 mg / kg of the cytochrome P450 inhibitor ritonavir) twice daily for 2 days (once on day 0, twice on day 1, and once on day 2) or 4 days (once on day 0, twice on days 1, 2, and 3). Mouse weight changes were recorded, and lung and brain tissues were collected at the end of the study. The left lung was fixed with formaldehyde, embedded, sectioned, and stained with H&E for histopathological examination. The right lung and brain tissues were each divided into two portions. One portion was polished, homogenized, and RNA was extracted and reverse-transcribed for viral copy number detection by real-time fluorescent quantitative PCR. The other portion was polished, homogenized, and viral titer detection by plaque assay. Plaque assay method: Vero E6 cells were seeded at 12,000 cells / well in a 24-well plate and cultured overnight. The stock tissue homogenate was diluted 10-fold with DMEM medium for use. The cell supernatant was removed, and the diluted tissue homogenate was added and incubated for 1 hour. The supernatant was then removed, and medium containing 1% sodium methylcellulose and 2% FBS was added and cultured for 4 days. The medium was then removed, and the cells were fixed with paraformaldehyde and stained with 1% (w / v) crystal violet. The number of plaques in each well was counted.

[0163] Test results: As shown in Table 7, two days after infection, compared with the model group (mean viral copy number was 9.19±0.30 log10 copies / g), when combined with ritonavir, both 50 mg / kg and 200 mg / kg of compound of formula (I) significantly reduced the viral load in the lungs, with the mean copy numbers being 7.66±0.27 log10 copies / g and 6.79±0.30 log10 copies / g, respectively, with the 200 mg / kg dose reducing the viral copy number by 2.4 log10 copies / g. Four days after infection, the compound of formula (I) showed a sustained inhibitory effect on the viral copy number.

[0164] Regarding viral titer, as shown in Figure 8, a significant inhibitory effect of the compound of Formula (I) was observed. Two days after infection, a dose of 200 mg / kg completely inhibited viral replication, and no titer was measured. At 50 mg / kg, the viral titer was reduced by more than 3 log10 PFU / g compared to the model group. Four days after infection, the compound of Formula (I) demonstrated a sustained inhibitory effect on viral titer. Regarding body weight, as shown in Figure 9, four days after infection, mice in the model group lost approximately 10% of their body weight, while the group treated with the compound of Formula (I) showed no significant weight loss, indicating that sustained administration of the compound of Formula (I) did not result in any significant toxicity. Viral load in the mouse brain was further detected, and no significant infection was observed in either group two days after infection. Four days after infection, compared with the model group, compound of formula (I) at both 50 mg / kg and 200 mg / kg doses significantly reduced the viral copy number in the mouse brain. At 200 mg / kg, the viral copy number in the brain was comparable to that of the uninfected control group. Four days after infection, viral titers were also measured in the brain. As shown in Figure 10, compared with the model group, no viral titers were measured at either dose of compound of formula (I), demonstrating the potent inhibitory effect of compound of formula (I). Furthermore, histopathological analysis of the lungs showed that compound of formula (I) at a dose of 200 mg / kg significantly improved lung damage, including reducing the degree of alveolar atrophy or dilation and the degree of alveolar membrane thickening, compared with the model group.

[0165] TIFF2026502598000018.tif66170

[0166] Test Example 5: Selectivity of compounds of formula (I) against kinases Experimental Objective: To detect the inhibitory activity of the compound of formula (I) against 413 kinases and to study the selectivity of the compound of formula (I) against kinases using the KinaseProfile experimental platform.

[0167] Test materials: The Full Human Panel [10 uM ATP] KinaseProfiler is a test product provided by Eurofins, product number 50-005KP10, which contains 413 kinases.

[0168] Testing Procedure: Compounds were tested against each selected kinase using the Eurofins standard KinaseProfiler assay, following the relevant standard operating procedures. Protein kinases were detected by radioactivity, while lipid kinases were detected by HTRF. The ATP concentration during the experiments was 10 μM. Detailed information about each kinase is available on the Eurofins website: https: / / www.eurofinsdiscoveryservices.com / catalogmanagement / viewItem / Full-Human-Panel-10-uM-ATP-KinaseProfiler / 50-005KP10.

[0169] Experimental results: The compound of formula (I) showed less than 30% inhibition against 413 kinases at a concentration of 10 μM, indicating no obvious inhibitory effect, suggesting that the compound of formula (I) has excellent selectivity.

[0170] Test Example 6: Selectivity of compounds of formula (I) for safety targets Experimental Objective: To detect the effects of compound of formula (I) on 47 safety-related targets using Safetyscan experimental platform.

[0171] Test materials: The Safety47 Panel Dose Response SAFETYscan was a test product provided by Eurofins with product number 87-1003DR. This product included 78 tests related to 47 safety targets.

[0172] Testing Procedure: For the 78 tests related to the 47 safety targets, the experimental methods used included cAMP experiments, calcium flux experiments, hormone nuclear receptor experiments, kinase binding experiments, enzyme activity experiments, neurotransmitter transporter experiments, ion channel experiments, and transporter experiments. The specific methods for each experiment were available on the eurofins website, which can be found at: https: / / www.eurofinsdiscoveryservices.com / catalogmanagement / viewItem / Safety47-Panel-Dose-Response-SAFETYscan-DiscoverX / 87-1003DR.

[0173] Experimental results: The compound of formula (I) had no significant inhibitory or activating effect on any of the 47 safety-related targets at a concentration of 100 μM (EC 50 and β-glucan were all greater than 100 μM), suggesting that the compound of formula (I) has excellent selectivity.

[0174] Test Example 7: Human plasma protein binding test of the compound of formula (I) Experimental materials Human plasma was purchased from BioIVT, anticoagulated with EDTA K2, and stored at −80° C. 96-well balance dialysis plates were purchased from HTDialysis LLC. Balance dialysis membranes were purchased from Gales Ferry.

[0175] Experimental procedure A basic solution of 14.2 g / L disodium hydrogen phosphate and 8.77 g / L sodium chloride was prepared using ultrapure water, and the basic solution could be stored for 7 days at 4°C. An acidic solution of 12.0 g / L sodium dihydrogen phosphate and 8.77 g / L sodium chloride was prepared using ultrapure water, and the acidic solution could be stored for 7 days at 4°C. The basic solution was titrated with the acidic solution to a pH of 7.4, and the buffer solution could be stored for 7 days at 4°C. On the day of the experiment, the pH of the buffer solution was measured, and if it was outside the range of 7.4 ± 0.1, the pH was adjusted.

[0176] To separate the membrane into two pieces, the dialysis membrane was immersed in ultrapure water for 60 minutes, then in 20% ethanol for 20 minutes, and finally in dialysis buffer for 20 minutes.

[0177] Frozen plasma was rapidly thawed at room temperature.

[0178] The plasma was centrifuged at 3220 g for 10 minutes at 4°C to remove clots, and the supernatant was collected in a new centrifuge tube. The pH of the plasma was measured and recorded.

[0179] A 10 mM DMSO stock solution of the test substance was prepared. 2 μL of the stock solution (10 mM) was diluted with 98 μL of DMSO to obtain a working solution (200 μM). 3 μL of the working solution was taken and 597 μL of human plasma was added to give a final concentration of 1 μM (0.5% DMSO). The mixture was vortexed thoroughly and uniformly.

[0180] A 120 μL sample of drug-containing plasma was added to one side of the dialysis membrane, and an equal volume of dialysate (phosphate buffer) was added to the other side. Experiments were performed in duplicate. The dialysis plate was sealed and placed in an incubator and incubated at 37°C, 5% CO2, and a rotation speed of approximately 100 rpm for 6 hours. After incubation, the sealing membrane was removed, and 50 μL of each well was aspirated from the buffer and plasma side and transferred to a different well of a new plate.

[0181] 50 μL of blank plasma was added to the phosphate buffer sample, and an equal volume of blank phosphate buffer was added to the plasma sample. 300 μL of room-temperature quencher (internal standard acetonitrile (IS), containing 500 nM labetalol, 100 nM alprazolam, and 2 μM ketoprofen)) was added to precipitate proteins. The mixture was vortexed for 5 minutes and centrifuged at 3220 g for 30 minutes at 4°C. 100 μL of the supernatant was transferred to a new plate. Depending on the LC / MS response signal and peak shape of the analyte, the supernatant was diluted with 100 μL or 200 μL of water. After uniform mixing, the sample was analyzed by LC / MS.

[0182] All calculations were performed using Microsoft Excel. The peak areas of the test compounds on the buffer and plasma sides were determined. The calculation formulas for the plasma protein binding rates of the test compounds and control drugs were as follows: release rate = (ratio of sample peak area to internal standard peak area on the buffer side / ratio of sample peak area to internal standard peak area on the plasma side) × 100%, binding rate = 1 − release rate, recovery rate = (ratio of sample peak area to internal standard peak area on the buffer side + ratio of sample peak area to internal standard peak area on the plasma side) / (ratio of sample peak area to internal standard peak area on the initial plasma sample) × 100%. The ratio of sample peak area to internal standard peak area on the buffer side represents the free concentration of the compound, the ratio of sample peak area to internal standard peak area on the plasma side represents the sum of the free and bound concentrations of the compound, and the ratio of sample peak area to internal standard peak area on the initial plasma sample represents the total concentration of the compound at the start of sample incubation.

[0183] Test results: The results are shown in Table 8. When 1 μM of the compound of formula (I) was incubated at 37° C. for 6 hours, the average release rate was 46.63%, the binding rate was 53.37%, and the recovery rate was 88.02%.

[0184] TIFF2026502598000019.tif22170

[0185] Test Example 8: Tissue distribution test by single intragastric administration of the compound of formula (I) Test materials: Balb / c mice (purchased from Shanghai Minchang Biotechnology Co., Ltd.) were used, half male and half female, a total of 60 mice, weighing 18-25 g.

[0186] Testing Procedure: The compound of formula (I) was administered intragastrically to Balb / c mice in a single dose of 100 mg / kg in a volume of 10 mL / kg.

[0187] Before administration and at 5 min, 0.25, 1.0, 2.0, 3.0, 5.0, 7.0, and 10 h post-administration (six mice, half male and half female per time point), 0.2 mL of blood was collected from the retrobulbar venous plexus in EDTA-K2 tubes and centrifuged at 11,000 rpm for 5 min to separate the plasma, which was then frozen in a -70°C refrigerator. Immediately after the whole blood collection at 0.25, 1.0, 3.0, and 7.0 h, lung tissues were dissected and washed with cold saline to remove any remaining blood and contents from the tissue surface. After drying, the tissues were labeled and stored at -70°C for further analysis. The content of compound of formula (I) in plasma and lung tissues was measured by LC / MS-MS, and the lung-blood ratio was calculated.

[0188] Test results: After a single intragastric administration of the compound of formula (I) to Balb / c mice, the ratio of lung tissue exposure to plasma exposure was 0.62, with the compound of formula (I) showing higher lung tissue exposure.

[0189] Test Example 9: Safety pharmacology test of the effect of intragastric administration of the compound of formula (I) on the cardiovascular system of cynomolgus monkeys The effects of compounds of formula (I) on the cardiovascular system were investigated together in a two-week repeated dose toxicity study in cynomolgus monkeys.

[0190] Test materials: Thirty-two cynomolgus monkeys, half male and half female, were aged 2.5 to 5 years at the time of administration.

[0191] Animal origin: Yunnan Yingmao Biological Technology Co., Ltd., Guangxi Yusen Primate Experimental Animal Culture Development Co., Ltd., Zhongke Lingrui (Zhanjiang) Biotechnology Co., Ltd.

[0192] Systolic blood pressure (SBP), diastolic blood pressure (DBP), and mean arterial pressure (MBP) were measured in all conscious animals using a smart noninvasive sphygmomanometer BP-98E with a Provantis / v10.2.3.1 electronic data acquisition system (PV-02).

[0193] Experimental Procedure: Thirty-two cynomolgus monkeys (five animals / sex / group in Groups 1 and 4, and three animals / sex / group in Groups 2 and 3, for a total of four groups) were randomly assigned to receive either the compound of Formula (I) (40, 160, or 600 mg / kg / day) or a control formulation (98.9% vehicle formulation + 1.1% MTBE (methyl tert-butyl ether), 0 mg / kg / day) by nasal gavage twice daily for a total of 14 days, followed by a 14-day recovery period. All animals were enrolled in this study to evaluate the effects of treatment on ECG parameters (including heart rate, PR interval, QRS duration, QT interval, and QTcF) and blood pressure during the pre-treatment, treatment, and recovery periods.

[0194] Test results: In this test condition, cynomolgus monkeys were administered the compound of formula (I) (40, 160 and 600 mg / kg / day) twice daily by nasogastric gavage for 14 days, and no test-related cardiovascular changes were observed, no test-related arrhythmias were observed, and no test-related changes in ECG parameters or blood pressure were observed throughout the test process.

[0195] Test Example 10: Chemical Stability Consideration Test Test method: The chemical stability of type A crystals of the compound of formula (I) and type B crystals of the methyl tert-butyl ether solvate of the compound of formula (I) under conditions such as high temperature, light irradiation, acceleration, and high humidity was examined. Both unpackaged and packaged (packaged using two layers of polyethylene and one layer of medicinal composite film) under light irradiation conditions were simultaneously examined, and the results are shown in Table 9.

[0196] Test results: When Type B crystals were left under each of the conditions of the influencing factors for 12 days, and Type A crystals were left under each of the conditions of the influencing factors for 12 days and 30 days, there was no significant change in the content, and the chemical stability of both Type A crystals and Type B crystals was stable.

[0197] TIFF2026502598000020.tif85170

[0198] Test Example 11: Crystalline form stability study Test Method: Form A crystals of the compound of formula (I) were packaged according to the following protocol, and the samples were left standing. After 30 days, the samples were taken out and subjected to XRPD measurement.

[0199] TIFF2026502598000021.tif42170

[0200] The test results showed that when the A-type crystal of the compound of formula (I) was left for 30 days under the conditions of each influencing factor, the characteristic diffraction peaks of the crystalline form were consistent, and there was no significant difference in the crystallinity, indicating that the crystalline form was stable.

[0201] Test Example 12: Hygroscopicity test Test Method: (1) Experimental equipment: Dynamic moisture adsorption device DVS Intrinsic PLUS, (2) Experimental conditions: Each sample (appropriate amount) was taken and placed on a DVS sample disk for testing.

[0202] (3) DVS parameters: Temperature: 25℃, Equilibrium: dm / dt≦0.002% / min RH(%) test step: 10% RH(%) test step range: 0%~90%~0%.

[0203] Hygroscopicity evaluation standard (based on the description of hygroscopicity characteristics and the definition of weight increase due to moisture absorption in "9103 Guide Principles for Drug Hygroscopicity" of Part 4 of the 2020 edition of the Chinese Pharmacopoeia): TIFF2026502598000022.tif35170

[0204] *Weight gain due to moisture absorption at 25±1°C and 80±2% RH.

[0205] Test results: The weight gain of the A-type crystals of the compound of formula (I) due to moisture absorption was only 0.5% and 0.6% under conditions of 80% RH and 90% RH, respectively.

Claims

1. A crystal of the compound of formula (I). 【Chemistry 1】

2. A type A crystal of the compound of formula (I), 【Chemistry 2】 The X-ray powder diffraction pattern of the A-type crystals, expressed in terms of diffraction angle 2θ, has diffraction peaks at 10.88±0.20°, 15.09±0.20°, 17.67±0.20°, 18.28±0.20°, and 20.64±0.20°; or The X-ray powder diffraction pattern of the A-type crystals, expressed in terms of diffraction angle 2θ, has diffraction peaks at 10.88±0.20°, 15.09±0.20°, 16.61±0.20°, 17.67±0.20°, 18.28±0.20°, 18.50±0.20°, 20.09±0.20°, and 20.64±0.20°; or The X-ray powder diffraction pattern of the A-type crystals, expressed in terms of diffraction angle 2θ, has diffraction peaks at 10.27±0.20°, 10.88±0.20°, 11.86±0.20°, 15.09±0.20°, 16.61±0.20°, 17.67±0.20°, 18.28±0.20°, 18.50±0.20°, 20.09±0.20°, and 20.64±0.20°; or The X-ray powder diffraction pattern of the A-type crystal, represented by the diffraction angle 2θ, is 9.29±0.20°, 10.27±0.20°, 10.88±0.20°, 10.97±0.20°, 11.86±0.20°, 14.27±0.20°, 14.92±0.20°, 15.09±0.20°, 15.61±0.20°. having diffraction peaks at 15.78±0.20°, 16.61±0.20°, 17.67±0.20°, 18.28±0.20°, 18.50±0.20°, 20.09±0.20°, 20.64±0.20°, 22.18±0.20°, 23.80±0.20° and 25.63±0.20°; or The X-ray powder diffraction pattern of the A-type crystal, expressed in terms of diffraction angle 2θ, has the diffraction peaks shown in Table 3, 【change】 or The A-type crystal has an X-ray powder diffraction pattern, represented by diffraction angles 2θ, substantially as shown in FIG. Form A crystal of the compound of formula (I).

3. having a DSC pattern with a peak at 213.78°C ± 5.0°C; or The DSC pattern of the A-type crystals is substantially as shown in FIG. A type A crystal of the compound of formula (I) according to claim 2.

4. A method for producing type A crystals of the compound of formula (I) according to claim 2 or 3, comprising: the method includes mixing the compound of formula (I) with solvents (i) and (ii), crystallizing the compound, and separating the solid, wherein the solvent (i) is at least one selected from isopropyl acetate, ethyl acetate, and isopropyl alcohol, and is preferably isopropyl acetate, and the solvent (ii) is at least one selected from n-hexane and n-heptane, and is preferably n-heptane; method.

5. A solvate of a compound of formula (I), 【Transformation 3】 The solvate is selected from methyl tert-butyl ether solvate and 2-methyltetrahydrofuran solvate. Solvates of compounds of formula (I).

6. 6. The solvate of the compound of formula (I) according to claim 5, wherein the molar ratio of the compound of formula (I) to the solvent is about 0.5 to 2, preferably about 0.8 to 1.2, more preferably about 1.

0.

7. A type B crystal of a methyl tert-butyl ether solvate of the compound of formula (I), 【Chemistry 4】 The X-ray powder diffraction pattern of the B-type crystals, expressed in terms of diffraction angle 2θ, has diffraction peaks at 6.26±0.20°, 17.66±0.20°, and 20.24±0.20°; or The X-ray powder diffraction pattern of the B-type crystals, expressed in terms of diffraction angle 2θ, has diffraction peaks at 6.26±0.20°, 7.27±0.20°, 11.47±0.20°, 13.06±0.20°, 15.57±0.20°, 17.66±0.20°, 20.24±0.20°, and 22.72±0.20°; or The X-ray powder diffraction pattern of the B-type crystals, expressed in terms of diffraction angle 2θ, has diffraction peaks at 6.26±0.20°, 7.27±0.20°, 8.84±0.20°, 10.18±0.20°, 10.30±0.20°, 11.47±0.20°, 13.06±0.20°, 14.44±0.20°, 15.57±0.20°, 17.66±0.20°, 20.24±0.20°, 20.51±0.20°, and 22.72±0.20°; or The X-ray powder diffraction pattern of the B-type crystals, expressed in terms of diffraction angle 2θ, has the diffraction peaks shown in Table 2, 【change】 or The B-type crystal has an X-ray powder diffraction pattern, represented by diffraction angles 2θ, substantially as shown in FIG. B type crystal.

8. a DSC pattern exhibiting peaks at 93.36°C ± 5.0°C, 170.25°C ± 5.0°C, and 214.18°C ± 5.0°C; or The DSC pattern of the B-type crystals is substantially as shown in FIG. The B-type crystal according to claim 7.

9. A method for producing the B-type crystals according to claim 7 or 8, comprising mixing the compound of formula (I) with methyl tert-butyl ether, stirring, and then separating the solid.

10. A C-type crystal of the 2-methyltetrahydrofuran solvate of the compound of formula (I), 【Transformation 5】 The X-ray powder diffraction pattern of the C-type crystals, expressed in terms of diffraction angle 2θ, has diffraction peaks at 6.21±0.20°, 17.48±0.20°, and 20.86±0.20°; or The X-ray powder diffraction pattern of the C-type crystals, expressed in terms of diffraction angle 2θ, has diffraction peaks at 6.21±0.20°, 7.04±0.20°, 11.71±0.20°, 17.48±0.20°, 20.59±0.20°, and 20.86±0.20°; or The X-ray powder diffraction pattern of the C-type crystals, expressed in terms of diffraction angle 2θ, has diffraction peaks at 6.21±0.20°, 7.04±0.20°, 11.71±0.20°, 14.74±0.20°, 15.77±0.20°, 17.48±0.20°, 20.59±0.20°, and 20.86±0.20°; or The X-ray powder diffraction pattern of the C-type crystal, expressed in terms of diffraction angle 2θ, has the diffraction peaks shown in Table 1, 【change】 or The C-type crystal has an X-ray powder diffraction pattern, represented by diffraction angles 2θ, substantially as shown in FIG. C type crystal.

11. a DSC pattern exhibiting peaks at 90.39°C ± 5.0°C, 167.59°C ± 5.0°C, and 214.22°C ± 5.0°C; or The DSC pattern of the C-type crystals is substantially as shown in FIG. The C-type crystal according to claim 10.

12. 12. A method for producing the C-type crystals according to claim 10 or 11, comprising mixing the compound of formula (I) with 2-methyltetrahydrofuran, stirring, crystallizing, and then separating the solid.

13. A drug combination comprising type A crystals of the compound of formula (I) according to claim 2 or 3, type B crystals according to claim 7 or 8, type C crystals according to claim 10 or 11, or a combination thereof, and another antiviral drug, preferably wherein the other antiviral drug is ritonavir.

14. A pharmaceutical composition comprising type A crystals of the compound of formula (I) according to claim 2 or 3, type B crystals according to claim 7 or 8, type C crystals according to claim 10 or 11, or a combination thereof, and pharmaceutically acceptable auxiliary materials.

15. Use of type A crystals of compound of formula (I) according to claim 2 or 3, type B crystals according to claim 7 or 8, type C crystals according to claim 10 or 11 or a combination thereof, or a drug combination according to claim 13, or a pharmaceutical composition according to claim 14, in the manufacture of a drug for preventing or treating related diseases caused by coronavirus and / or small RNA virus infection.

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