Crystal form of compound

Abstract

Provided is crystal form 1 of a compound, wherein the compound has a structural formula as shown in formula I. The crystal form of the compound has a relatively high solubility in a suitable solvent, and a prescription design for liquid preparations with different concentrations can be realized by means of solvent combinations during preparation development. Meanwhile, the crystal form of the compound has no hygroscopicity and also shows good stability in a high-temperature and high-humidity environment, so that the crystal form 1 of the compound is suitable for later preparation development.

Description

Compound crystal form Technical Field

[0001] The present invention relates to compounds, and in particular to stable crystalline forms of specific compounds suitable for development into pharmaceutical formulations. Background Art

[0002] The applicant's patent application PCT / CN2020 / 089692 (publication number WO2020228685A9) and patent application PCT / CN2021 / 129077 disclosed compound No. 16 (hereinafter referred to as Compound I). After further research, it was found that Compound I has excellent anticancer efficacy, toxic side effects, safety and stability, especially in tumor tissues or cells that overexpress aldehyde-keto reductase 1C3 (AKR1C3). It can be targeted for identification and targeted treatment, and can enter the brain or other central nervous systems to exert its efficacy. The Chinese name of Compound I is 1-(3-((2,4'-difluoro-[1,1'-biphenyl]-4-yl)oxy)-4-nitrophenyl)-2,2,2-trifluoroethyl-bis(aziridine-1-yl) phosphate, and its structure is shown below:

[0003] In order to conduct subsequent clinical trials, it is necessary to prepare a suitable dosage form for patient administration: typically oral or injection. Therefore, it is necessary to develop a formulation for Compound I (also referred to as the active pharmaceutical ingredient or API in this application) for the treatment of cancer patients who overexpress aldehyde-keto reductase 1C3 (AKR1C3). According to the above-mentioned patent application, Compound I is a solid at room temperature.

[0004] The applicant disclosed the prescription and preparation method of the injection preparation of compound AST-3424, which has a structure similar to compound I, in patent application PCT / CN2020 / 101870 (publication number WO2021008520A1, corresponding to Chinese patent application CN202080001484.5, publication number CN112469394A). Since compound AST-3424 is a viscous oil at room temperature, it is actually developed as an injection concentrated solution of a propylene glycol-ethanol mixed solvent.

[0005] Since compound I and compound AST-3424 have similar structures and similar mechanisms of action, there is sufficient motivation to draw on the formulation formula of AST-3424 for the development and formulation research of injections. Summary of the Invention

[0006] The inventors further discovered that although the above-mentioned compound I is structurally similar to the compound AST-3424, their solubility in ethanol, propylene glycol, and ethanol-propylene glycol mixed solvents is significantly different:

[0007] The solubility of AST-3424 in ethanol is greater than 1000 mg / ml (almost miscible), the solubility in propylene glycol mixed solvent is greater than 100 mg / ml, and the solubility in ethanol-propylene glycol mixed solvent (volume ratio of 1:1) is greater than 270 mg / mL;

[0008] The solubility of Compound I in ethanol is less than 8 mg / mL, the solubility in propylene glycol is about 2 mg / mL, and the solubility in an ethanol-propylene glycol mixed solvent (volume ratio of 1:1) is about 6 mg / ml.

[0009] In this regard, the applicant's inventor team believes that the difference between the two may be related to the fact that Compound I is a solid: solids have crystalline forms, and different crystalline forms often have different solubility and stability. Therefore, it is necessary to explore the crystalline form of Compound I and confirm the preparation of a stable crystalline form suitable for development into a pharmaceutical preparation.

[0010] The compound crystal form 1 explored and prepared in the present invention has been experimentally verified to be a stable crystal form suitable for development into a pharmaceutical preparation.

[0011] Compound Form 1, the compound structural formula is as shown in Formula I, and the compound Form 1 satisfies one of the following conditions:

[0012] Condition 1: Compound Form 1 was subjected to X-ray powder diffraction analysis using Cu target Kα radiation with a scanning range of 3° to 40°. The resulting spectrum had characteristic peaks at 9.1±0.2, 18.5±0.2, 20.5±0.2, 21.3±0.2, and 25.8±0.2, expressed in 2θ°.

[0013] Condition 2: Compound Form 1 was tested by differential scanning calorimetry. The temperature was increased from 25°C to 180°C at a rate of 10°C / min. The temperature rise curve showed an endothermic melting peak with a peak range of 135.27±0.05°C to 140.19±0.05°C, a corresponding onset point, or melting point, ranging from 131.09±0.05°C to 138.97±0.05°C, and an enthalpy value ΔH ranging from 48.647±0.039 J / g to 83.701±0.067 J / g.

[0014] Condition 3: Solubility test by the following operations:

[0015] 5.0±0.5 mg of Compound Form 1 was accurately weighed into a sample bottle, and then a specific solvent was gradually added at room temperature (25°C) until the solid dissolved or the total volume reached 8 ml. The results are shown in Table 1 below:

[0016] Table 1: Solubility data of compound crystal form 1 in 20 solvents of different polarities

[0017] in,

[0018] Solubility data > 100 mg / ml means that the volume of solvent added is less than 0.05 ml, that is, the substance is completely dissolved until no solid can be observed with the naked eye.

[0019] >50mg / ml means the volume of solvent added is less than or equal to 0.05ml and less than 0.10ml, i.e. the substance is completely dissolved until no solid can be observed with the naked eye.

[0020] <0.6mg / ml means that after the added solvent exceeds the total amount of 8ml in the sample bottle, the solid is still visible to the naked eye and cannot be completely dissolved.

[0021] Others ~33, ~12.5, ~7.5, ~4.5, ~3, ~2.9, and ~2 indicate solubility of 33±3.3 mg / ml, 12.5±1.25 mg / ml, 7.5±30.75 mg / ml, 4.5±0.45 mg / ml, 3±0.3 mg / ml, 2.9±0.29 mg / ml, and 2±0.2 mg / ml, which means that after adding a specific volume of a specific solvent, the solvent is completely dissolved until no solid can be observed with the naked eye.

[0022] As an embodiment of the present application, compound crystal form 1 satisfies the above three conditions at the same time.

[0023] There are many technically available methods for characterization and identification of organic compound crystal forms, as shown in Table 2:

[0024] Table 2: Characterization and identification methods of organic compound crystal forms

[0025] Although the above methods / instruments can all determine certain aspects of the characteristics of the crystalline compound from a certain perspective after detection, X-ray diffraction (especially powder diffraction XRPD) and differential scanning calorimetry (DSC) are relatively speaking. Due to their good reproducibility and ability to provide relatively rich information to form specific spectra, they achieve an effect similar to a "fingerprint" and are gradually considered to be the "gold standard" for characterizing and confirming the crystalline form of a compound, and are unanimously recognized by technicians in related fields.

[0026] For powder diffraction XRPD, the crystal form can be determined by selecting specific characteristic peaks. According to the Bragg equation: for X-ray diffraction, when the optical path difference is equal to an integer multiple of the wavelength, the diffraction line of the crystal plane will be enhanced. The condition satisfied at this time is 2dsinθ=nλ, where d is the interplanar spacing, θ is the angle between the incident ray, the reflected ray and the reflecting crystal plane, λ is the wavelength, and n is the reflection order. The Bragg equation is a necessary condition (but not a sufficient condition) for X-rays to diffract in crystals.

[0027] Because different targets have different characteristic X-ray wavelengths, according to the Bragg equation 2dsinθ = nλ, the diffraction angles of a family of crystal planes with a given spacing d will be different. The diffraction angles of families of crystal planes with different spacing values ​​will exhibit regular changes. Therefore, the positions of the diffraction peaks on the diffraction patterns obtained by X-ray tubes using different target materials will be different.

[0028] Compound I is an organic compound containing C, H, O, N, F, and P elements, but does not contain any metal elements. Its interplanar spacing is generally Therefore, Cu target is selected for testing, which emits three types of X-rays: Kα1 Kα2 Kβ1 For copper targets, the Kα wavelength is the weighted average of Kα1 and Kα2, which is (λ in the Bragg equation).

[0029] The applicant's inventor team and technical experts have determined that the compound crystal form 1 of compound I is tested under the test conditions of Cu target Kα rays and a scanning range 2θ of 3o to 40o. The spectrum obtained by X-ray powder diffraction test has characteristic peaks of 9.1±0.2, 17.2±0.2, 20.5±0.2, and 21.3±0.2 expressed in 2θ°; that is to say, if compound I has the above four characteristic peaks after X-ray powder diffraction test under the above conditions, it can be determined that compound I is compound crystal form 1.

[0030] Due to different conditions during XRPD testing, such as instrument detection accuracy or error, different wear or settings of instrument components (goniometer, divergence slit, receiving slit, anti-scattering slit, Sollar slit), sample preparation process (different sample powder particle sizes), different sample crystallinity and crystal habit, and the sample to be tested may be a mixture (for example, in a preparation, the content of the raw material drug is too little or too low, and if it is too low, some characteristic peaks will be unclear or disappear; if it is too high, other characteristic peaks will appear), other peaks will appear.

[0031] As for XRPD, the XRPD spectrum of compound form 1 under condition 1 also has a characteristic peak of 17.2±0.2 expressed in 2θ°.

[0032] As for XRPD, the XRPD spectrum of compound form 1 under condition 1 also has characteristic peaks at 12.9±0.2 and 19.5±0.2 expressed in 2θ°.

[0033] As for XRPD, the XRPD spectrum of compound form 1 under condition 1 also has characteristic peaks of 15.7±0.2, 16.8±0.2, 18.0±0.2, 26.8±0.2, and 29.8±0.2 expressed in 2θ°.

[0034] As for XRPD, the XRPD spectrum of compound form 1 under condition 1 also has a characteristic peak of 23.7±0.2 expressed in 2θ°.

[0035] For XRPD, the XRPD spectrum of compound form 1 under condition 1 also has characteristic peaks of 12.9±0.2, 15.7±0.2, 16.8±0.2, 18.0±0.2, 19.5±0.2, 23.7±0.2, 24.6±0.2, 24.9±0.2, 26.8±0.2, and 29.8±0.2 expressed in 2θ°.

[0036] “±0.2” is the allowable measurement error range.

[0037] For XRPD, the X-ray powder diffraction test of compound form 1 was performed using a PIXceI 1D The test was conducted using a Panalytical Empyrean X-ray powder diffractometer (Malvern Panalytical Ltd) with a Cu target Kα radiation, a scanning range of 3° to 40° in 2θ, a step size of 0.013° in 2θ°, and an X-ray tube voltage and current of 45 kV and 40 mA, respectively. The spectrum under condition 1 has the characteristic peaks listed in Table 3 below:

[0038] Table 3: Diffraction characteristic peaks of compound form 1 obtained by Panalytical Empyrean test

[0039] For XRPD, the X-ray powder diffraction test of compound form 1 was performed using a PIXceI 1D The test was conducted using a Panalytical Empyrean X-ray powder diffractometer (Malvern Panalytical Ltd) with a Cu target Kα radiation, a scanning range of 3° to 40° in 2θ, a step size of 0.013° in 2θ°, and an X-ray tube voltage and current of 45 kV and 40 mA, respectively. The spectrum under condition 1 has the characteristic peaks and corresponding interplanar distances listed in Table 4 below:

[0040] Table 4: Diffraction characteristic peaks and interplanar distances of compound form 1 obtained by Panalytical Empyrean testing

[0041] The interplanar distances in the table are calculated and corrected based on the Bragg equation.

[0042] For XRPD, the X-ray powder diffraction test of compound form 1 was performed using a PIXceI 1D The test was conducted using a Panalytical Empyrean X-ray powder diffractometer (Malvern Panalytical Ltd) with a Cu target Kα radiation, a scanning range of 3° to 40° in 2θ, a step size of 0.013° in 2θ°, and an X-ray tube voltage and current of 45 kV and 40 mA, respectively. The spectrum under condition 1 has the characteristic peaks and corresponding interplanar distances and relative intensities listed in Table 5 below:

[0043] Table 5: Diffraction characteristic peaks, interplanar distances, and relative intensities of compound form 1 obtained by Panalytical Empyrean testing

[0044] The interplanar distances in the table are calculated and corrected based on the Bragg equation.

[0045] Relative intensity is a calculated value after measurement and has uncertainty. Those skilled in the art understand that the uncertainty of relative intensity is highly dependent on the measurement conditions and has an interval of ±10% of the value. That is, the above-mentioned 78.4% possible interval ranges from 70.56% to 86.24%.

[0046] For XRPD, the X-ray powder diffraction test of compound form 1 was performed using a PIXceI 1D The test was performed using a Panalytical Empyrean X-ray powder diffractometer (Malvern Panalytical Ltd) with a Cu target Kα ray, a scanning range of 3° to 40° in 2θ, a step size of 0.013° in 2θ°, and an X-ray tube voltage and current of 45 kV and 40 mA, respectively. The spectrum under condition 1 is shown in FIG14 .

[0047] The instrument measurement parameters provided by the manufacturer of the X-ray powder diffractometer Panalytical Empyrean (Malvern Panalytical Ltd) are as follows: 2θ linearity equal to or better than ±0.01°. Therefore, the 2θ° measured by the above-mentioned X-ray powder diffractometer Panalytical Empyrean (Malvern Panalytical Ltd) all have an error range of ±0.01°.

[0048] When a substance undergoes changes in physical properties (such as crystallization, melting, or crystal phase transitions), or undergoes chemical changes, these changes are often accompanied by changes in thermodynamic properties such as enthalpy, specific heat, and thermal conductivity. DSC characterizes physical or chemical changes by measuring these changes in thermodynamic properties. Because different crystal forms of the same substance have different enthalpy, specific heat, and thermal conductivity, DSC curves can be used to generate different curves, as well as the onset temperature, endpoint temperature, peak temperature, and enthalpy values ​​of the exothermic / endothermic peaks. By setting the DSC instrument's heating start and end temperatures, as well as the heating rate, a reproducible DSC spectrum can be generated.

[0049] The applicant's inventor team and technical experts determined that the compound form 1 of compound I was tested by differential scanning calorimetry (DSC). The temperature was increased from 25°C to 180°C at a rate of 10°C / min, and the temperature rise curve was recorded, showing an endothermic melting peak with a peak range of 135.27±0.05°C to 140.19±0.05°C, corresponding to the starting point, i.e., the melting point range of 131.09±0.05°C to 138.97±0.05°C, and the enthalpy value ΔH range of 48.647±0.039J / g to 83.701±0.067J. / g; that is, if Compound I is subjected to DSC testing under the above conditions (heating from 25°C to 180°C at 10°C / min) and the temperature rise curve is recorded, if an endothermic melting peak is shown, with a peak value between 135.27±0.05°C and 140.19±0.05°C, a corresponding onset point, i.e., a melting point, between 131.09±0.05°C and 138.97±0.05°C, and an enthalpy value ΔH between 48.647±0.039 J / g and 83.701±0.067 J / g, then Compound I can be identified as Compound Form 1. The number after ± indicates the error range.

[0050] Due to differences in DSC test conditions, such as instrument manufacturers, principles, and sample preparation during testing, the peak value, starting point (melting point), end point, and enthalpy value of the endothermic melting peak in the DSC of the compound crystal form 1 sample may be affected.

[0051] Specifically, the amount of sample and the stacking of samples during sample preparation during testing will cause the shape of the endothermic melting peak to change.

[0052] Small sample size increases the separation ability of adjacent peaks and results in smaller peak shapes. Large sample size increases peak response, broadens the peak shape, and shifts characteristic peaks toward higher temperatures: This is because the larger the sample thickness in the crucible, the slower the heat transfer rate within the sample, forming a larger temperature gradient and increasing the width of the endothermic peak.

[0053] Compound Form 1 is generally a powder sample. When preparing the sample for testing, it is generally flattened into a thin layer at the bottom of the crucible. However, the compactness of the subsequent stacking method and the sample particle size will affect the size of the DSC peak shape: tight stacking is conducive to heat conduction inside the sample, and the measurement results will make the peak shape sharper. The smaller the sample particle size, the larger and sharper the peak. Excessively large particles or irregular particle shapes may cause the melting distance (the difference between the initial point and the end point) to become larger. For this, grinding (the grinding force should not be too strong to destroy the crystal conformation) and tight stacking can avoid the above situation and obtain a DSC spectrum with a small melting distance and moderate peak sharpness.

[0054] In this regard, the difference between the peak value and the onset point, i.e., the melting point, in condition 2 is in the range of 0.86 to 4.18°C, preferably in the range of 0.86 to 1.86°C.

[0055] For DSC, differential scanning calorimetry of compound Form 1 was performed using a differential scanning calorimeter Discovery DSC 250 (TA Instruments, US). The temperature rise curve in condition 2 was selected from the following eight temperature rise curves. These eight temperature rise curves all had endothermic melting peaks. Their peak values, corresponding starting points, i.e., melting points, and corresponding enthalpy values ​​ΔH are listed in Table 8 as follows:

[0056] Table 8: Peak value of endothermic melting peak, corresponding starting point (melting point), and corresponding enthalpy value ΔH of 8 DSC temperature rise curves of compound crystal form 1

[0057] The instrument measurement parameters of the differential scanning calorimeter Discovery DSC 250 (TA Instruments, US) provided by the instrument manufacturer are as follows: temperature accuracy is ±0.05° C., and enthalpy value accuracy is ±0.08%.

[0058] Therefore, for temperature, the above values ​​all have a range of ±0.05°C of the measured value, such as 131.09°C corresponds to a range of 131.04-131.14°C; for enthalpy, they all have a range of ±0.0008 times the measured value, such as 48.647 J / g corresponds to a range of (48.647-0.039)-(48.647+0.039), i.e. 48.608-48.686 J / g.

[0059] The DSC curve of Compound Form 1, i.e., temperature rise curve 7, is shown in the DSC curve in Figure 16. The same Compound Form 1 sample can be subjected to another DSC test to obtain temperature rise curve 8, as shown in the DSC curve in Figure 17.

[0060] Different crystalline forms of a compound have different free energies, resulting in different solubilities in the same solvent. Generally speaking, the greater the free energy, the more unstable the crystalline form and the greater the solubility; conversely, the solubility is lower. The solubility of a substance in a solvent is primarily determined by the properties of the solvent and the solute. Therefore, a certain crystalline form of a compound will have different solubilities in different solvents. By testing the solubility of a series of different solvents (with different polarities and structures), solubility data can be obtained, and the crystalline form can be determined using this series of solubility data.

[0061] The applicant's inventor team and technical experts have determined that the solubility is tested through the following operations:

[0062] 5.0±0.5 mg of Compound Form 1 was accurately weighed into a sample vial. A specific solvent was then gradually added at room temperature (25°C) until the solid dissolved or until a total volume of 8 ml was reached. The results are shown in Table 1: Solubility data of Compound Form 1 in 20 solvents of varying polarity. In other words, the solubility of Compound 1 in the aforementioned 20 solvents was tested according to the following procedure:

[0063] Accurately weigh 5.0±0.5 mg of Compound Form 1 into a sample vial, then gradually add a specific solvent at room temperature (25°C) until the solid dissolves or a total volume of 8 ml is added. Test and record the solubility data. If the data is consistent with Table 1 above, Compound I can be identified as Compound Form 1.

[0064] As a supplementary confirmation, the compound crystal form 1 can also be further confirmed by polarizing microscopy.

[0065] Furthermore, compound Form 1 does not undergo crystal form transformation when stored at 2-8°C, sealed, and in the dark, or at 25°C±2°C, 60%±5% RH, and in the dark for up to 36 months, and the purity and water content do not change substantially.

[0066] According to long-term stability experiments, the compound crystalline form 1 provided by the present invention has excellent stability. After 36 months of storage at 2-8°C, sealed, and in the dark, and at 25°C±2°C, 60%±5% RH, in the dark, XRPD detection of its characteristic peaks of the crystalline form still exists, confirming that no crystalline transformation or amorphization occurs under the above storage conditions, and therefore the crystalline form is stable.

[0067] At the same time, there was no substantial change in HPLC purity and water content. No substantial change refers to data fluctuations or measurement errors caused by sampling and measurement errors during HPLC testing and water content testing:

[0068] Taking HPLC purity testing as an example, in the long-term stability experiment, the HPLC purity of the initial compound Form 1 sample was 99.30%, and the values ​​tested at different times subsequently fluctuated within the range of (99.30±0.02)%, indicating that no substantial change had occurred.

[0069] Taking the water content test as an example, in the long-term stability experiment, the water content of the initial compound crystal form 1 sample was 0.06%, while the values ​​tested at different times were all within %, so there is actually no substantial change.

[0070] That is, according to the long-term stability experiment, the purity of compound crystal form 1 does not change substantially when stored at 2-8°C, sealed, and dark conditions or at 25°C±2°C, 60%±5% RH, and dark conditions for up to 36 months, and the water content does not change substantially. This means that the water content of the initial compound crystal form 1 sample is 0.06%, the HPLC purity is 99.30%, and the water content values ​​detected at different times in the subsequent storage are all within The HPLC purity values ​​fluctuated within the range of (99.30±0.02)%.

[0071] Furthermore, the crystal form 1 of the compound was observed to be rod-shaped under a polarizing microscope, and the specific morphology is shown in FIG15 .

[0072] As a supplementary confirmation, the compound crystal form 1 can be further confirmed by thermogravimetric analysis (TGA).

[0073] Furthermore, thermogravimetric analysis of the compound crystal form 1 showed that it lost weight within the range of 120-150°C, and the weight loss did not exceed 0.8%.

[0074] According to TGA thermogravimetric analysis, compound crystalline form 1 does not contain crystalline water and is an anhydrous crystalline form.

[0075] The TGA curve for Form 1, corresponding to Temperature Ramp Curve 7, is shown in Figure 16 . Its weight loss temperature is 120-150°C, and its weight loss is 0.774%. Another TGA test of the same Form 1 sample yields Temperature Ramp Curve 8, as shown in Figure 17 . Its weight loss temperature is 150°C, and its weight loss is 0.350%.

[0076] As a supplementary confirmation, dynamic water adsorption / desorption (DVS) can be used to further confirm the compound crystal form 1.

[0077] The compound crystal form 1 was subjected to dynamic moisture adsorption and desorption analysis, and its desorption weight loss was 0.13% when tested under 0-90% RH conditions.

[0078] Before determining the crystalline form of compound 1, compound 1 was identified by spectral testing.

[0079] High-resolution mass spectrometry of compound I showed a mass-to-charge ratio of m / z 556.1063 [M+H] + ion peaks and confirmed by any of the following spectral detections:

[0080] Spectral detection: Infrared spectroscopy, using KBr pellet method at 4000-400 cm -1 Infrared absorption spectra were collected within the wavenumber range, with the characteristic absorption wavenumbers, vibration types and structural fragment attributions shown in Table 9 below:

[0081] Table 9: Infrared spectrum measurement results of compound I

[0082] The vibration modes of polyatomic molecules and the various vibration types of groups are represented by the following symbols: ν represents stretching vibration, γ represents out-of-plane bending vibration, and δ represents deformation vibration.

[0083] The specific infrared spectrum of compound I is shown in Figure 1.

[0084] Absorption wave number of infrared absorption spectrum (cm -1 ) measurements are often dependent on a variety of factors, including the laboratory environment and sample preparation. Therefore, relative wavenumber values ​​are subject to normal error. For the same substance, those skilled in the art generally consider infrared (IR) absorption wavenumber measurements to have a measurement range of ±8 for the measured value. This ±8 represents the error.

[0085] Spectral detection 2, nuclear magnetic resonance hydrogen spectrum, detection after dissolution in DMSO-d6,

[0086] The chemical shift, peak shape and proton number are shown in Table 10 below:

[0087] Table 10: H NMR spectrum of Compound I 1 H-NMR measurement results

[0088] Spectral detection three, nuclear magnetic resonance carbon spectrum, detection after dissolution in DMSO-d6,

[0089] The chemical shift and carbon type are shown in Table 11 below:

[0090] Table 11: C NMR spectrum of Compound I 13 C-NMR measurement results

[0091] Since the chemical shift signal will be affected by many factors such as the concentration of the test substance, the test temperature and the test instrument when the test solvent is determined, the relative test shift value will have normal errors. For the same substance in the same solvent, generally speaking, those skilled in the art usually determine that the nuclear magnetic resonance hydrogen spectrum 1 The chemical shift results measured by H-NMR have a measurement range of ±0.2 ppm. 13 C-NMR has a measurement range of ±2 ppm, where ±0.2 ppm and ±2 ppm represent errors.

[0092] HRMS testing has extremely high accuracy and precision. The mass-to-charge ratio is m / z 556.1063 [M+H] + In the measurement result of 556.1063, the first three decimal places are accurate, and only the last digit 3 may have an error. Therefore, the actual measurement range may be 556.1061, 556.1062, 556.1063, 556.1064, 556.1065, 556.1066, 556.1067, 556.1068, and 556.1069.

[0093] As an additional confirmation, you can also 19 F-NMR was used to further confirm compound I. 19 In the F-NMR spectrum, there are three groups of fluorine signals at chemical shifts δ-76.80, δ-114.26, and δ-114.62.

[0094] As an additional confirmation, you can also 31 P-NMR was used to further confirm compound I. 31 In the P-NMR spectrum, there is a group of phosphorus signals at a chemical shift of δ31.84.

[0095] As a supplementary confirmation, compound I can also be further confirmed by UV.

[0096] UV analysis was performed using an acetonitrile sample solution (5.21 μg / mL). The UV absorption detection results and analysis of Compound I are shown in Table 12 below:

[0097] Table 12: UV absorption of compound I in acetonitrile solution

[0098] The UV spectrum of specific compound I is shown in Figure 2.

[0099] Spectral analysis

[0100] Mass spectrometry

[0101] The high-resolution mass spectrum shows a mass-to-charge ratio of m / z 556.1063 [M+H] + The ion peak of compound I has a deviation of less than 5ppm from the theoretical value (compound I molecular formula C 24 H 20 F5N3O5P, relative molecular mass theoretical value 556.1061), suggesting that the sample molecular formula is C 24 H 20 F5N3O5P, consistent with the structure of compound I.

[0102] Infrared IR

[0103] 1) 3090 and 3013 cm -1 The stretching vibration absorption peaks of =CH bond are at 1609, 1599 and 1491 cm -1 The stretching vibration absorption peaks of C=C double bond are at 1163, 1069 and 897 cm -1 The absorption peaks of the in-plane bending vibration of =CH are at 849 and 818 cm -1 The absorption peak at =CH is the out-of-plane bending vibration absorption peak, which is consistent with the absorption peak of the benzene ring fragment contained in the structure;

[0104] 2) 3071 and 2938 cm -1 The stretching vibration absorption peaks of C-H bonds are at 1437, 1431 and 1389 cm -1 The absorption peak at is the bending vibration peak of the C-H bond, which is consistent with the absorption peaks of the CH and CH2 fragments contained in the structure;

[0105] 3) 1530cm -1 The asymmetric contraction vibration absorption peak of nitro group is 1352cm -1 The absorption peak at is the symmetrical stretching vibration of the nitro group, which is consistent with the absorption peak of the nitro fragment contained in the structure;

[0106] 4) 1288 and 1146 cm -1 The stretching vibration absorption peak of the CF bond is at , which is consistent with the CF bond structure in the structure;

[0107] 5)1279cm -1 The stretching vibration absorption peak of COC is at , which is consistent with the presence of aryl ether structure in the structure;

[0108] 6) 1229 and 1188 cm -1 The stretching vibration absorption peaks of P=O are at 962, 947 and 937 cm -1 The absorption peak of POC stretching vibration is consistent with the absorption peak of the phosphate group fragment contained in the structure;

[0109] 7)723cm -1The absorption peak at is the swing vibration of CH, which is consistent with the CH2 structure in the structure; the infrared spectrum characteristics are consistent with the structure of compound I.

[0110] Ultraviolet UV

[0111] The absorption peaks of ultraviolet absorption λmax 247 and 196 nm are the n-π* transition absorption peaks of the conjugated system of the compound and the π-π* transition absorption peaks of the substituted benzene ring, which are consistent with the structure of compound I.

[0112] Nuclear Magnetic Resonance (NMR)

[0113] 1 H-NMR

[0114] The structural formula and atomic number of compound I and the division of CH and HH related fragments

[0115] Table 13: Compound I 1 H-NMR measurement results and H atom assignment

[0116] 1 The H-NMR spectrum showed a total of 19 hydrogen signals, including 8 methylene hydrogens and 11 methine hydrogens. A group of aromatic hydrogen signals at chemical shifts of δH 8.27 (d, J = 8.8 Hz, 1H), δH 7.68 (d, J = 8.4 Hz, 1H), and δH 7.58 (br s, 1H) are characteristic signals of 1,2,4-trisubstituted benzene rings. Based on HMBC correlation, they were assigned to H-5, H-4, and H-2, respectively. Aromatic hydrogen signals at chemical shifts of δH 7.63 (overlap, 1H), δH 7.20 (dd, J = 11.6, 2.4 Hz, 1H), and δH 7.04 (dd, J = 8.4, 2.4 Hz, 1H) are characteristic signals of another group of 1,2,4-trisubstituted benzene rings. Based on HMBC correlation, they were assigned to H-24, H-21, and H-25, respectively. A group of aromatic hydrogen signals at H7.61 (overlap, 2H) and δH7.35-7.31 (m, 2H) are characteristic signals of 1,4-disubstituted benzene rings. Combined with HMBC correlation, they are assigned to H-28, 32 and H-29, 31 respectively; the hydrogen signal with a chemical shift at δH6.39-6.32 (m, 1H) is HMBC-related with C-2, C-3, C-4 and C-10 and is assigned to H-9; the hydrogen signal with a chemical shift at δH2.20-1.98 (m, 8H) is assigned to H-16, 17, 18, 19 according to its chemical shift. The hydrogen spectrum data is consistent with the structure of compound I. The specific hydrogen signal assignments are shown in Table 13. 1 The H-NMR spectrum is shown in Figure 3.

[0117] 13 C-NMR and DEPT 135

[0118] Table 14: Compound I 13 C-NMR measurement results and carbon atom assignment

[0119] 13 The C-NMR spectrum gave a total of 24 carbon signals, which, combined with DEPT 135, showed that there were 4 methylene carbons, 11 methine carbons and 9 quaternary carbons. Through HSQC data, all the above hydrogen-linked carbon signals were assigned, and the remaining quaternary carbons were assigned by chemical shift, coupling constant and HMBC: in the HMBC spectrum, δC 163.54, 161.10 (d, J = 244 Hz) and δC 131.15, 131.13 (d, J = 2 Hz) were related to H-28, 32 and H-29, 31, and combined with the coupling constant, they were assigned to C-30 and C-27, respectively; δC 161.10, 158.64 (d, J = 246 Hz), δC 156.56, 156.45 (d, J = 11 Hz) and δC 124.15, 124.02 (d, J = 13 Hz) are related to H-21, H-24 and H-25, and are attributed to C-22, C-20 and C-23, respectively, in combination with the coupling constant; δC148.91, δC 142.31 and δC 139.51 are related to H-2, H-4 and H-5, and are attributed to C-1, C-6 and C-3, respectively, in combination with the chemical shift; δC 127.52, 127.44, 124.71, 124.63, 121.92, 121.83 (td, J = 281, 8 Hz) are related to H-9, and are attributed to C-10 in combination with the chemical shift and coupling constant; the carbon spectrum data are consistent with the structure of compound I. The specific hydrogen signal assignments are shown in Table 14. 13 The C-NMR spectrum is shown in Figure 4, and the specific DEPT135 spectrum is shown in Figure 5.

[0120] 19 F-NMR spectrum

[0121] exist 19 In the F-NMR spectrum, three groups of fluorine signals were given at chemical shifts δ-76.80, δ-114.26, and δ-114.62, which are consistent with the structure of compound I, as shown in Figure 6.

[0122] 31 P-NMR spectrum

[0123] exist 31 In the P-NMR spectrum, the chemical shift at δ31.84 gave a group of phosphorus signals, which were consistent with the structure of compound I, as shown in FIG7 .

[0124] HSQC and HMBC spectra

[0125] The NMR HSQC and HMBC assignment table is shown in Table 15 below.

[0126] Table 15: HSQC and HMBC spectrum measurement results

[0127] All hydrogen-linked carbon atoms were assigned by HSQC. In the HMBC spectrum, H-28 and 32 correlated with C-27, C-29, 31, and C-30, while H-29 and 31 correlated with C-27, C-28, 32, and C-30, consistent with the presence of fragment A in the structure. H-24 correlated with C-20, C-21, C-22, C-23, and C-25, while H-25 correlated with C-20, C-21, C-22, and C-23. H-21 correlated with C-20, C-22, C-23, C-24, and C-25, consistent with the presence of fragment B in the structure. H-9 correlated with C-10, consistent with the presence of fragment C in the structure. H-28 and 32 correlated with C-23, and H-24 correlated with C-27, indicating that C-27 of fragment A is linked to C-23 of fragment B. H-9 is correlated with C-3, and H-2 and H-4 are correlated with C-9, indicating that C-3 of fragment B is connected to C-9 of fragment C. HSQC and HMBC data are consistent with the structure of compound I, as shown in Figures 8 and 9.

[0128] COSY spectrum

[0129] The NMR COSY value assignment table is shown in Table 16 below.

[0130] Table 16: HH COSY spectrum measurement results of Compound I

[0131] In the COSY spectrum, H-28 correlates with H-29, and H-31 correlates with H-32, further confirming the presence of fragment A in the structure; H-24 correlates with H-25, further confirming the presence of fragment B; H-16 correlates with H-17, and H-18 correlates with H-19, further confirming the presence of fragment C. The COSY spectral data are consistent with the structure of compound I, as shown in Figure 10.

[0132] That is, through the above spectral analysis, it can be known that for a certain unknown organic compound after purification, if the purity of the sample is greater than or equal to 95% or higher using the area normalization method in HPLC detection:

[0133] i. After the above-mentioned high-resolution mass spectrometry and infrared spectrum detection, if it is determined to be consistent, it can be confirmed that the substance is compound I;

[0134] ii. After the above-mentioned high-resolution mass spectrometry and nuclear magnetic resonance hydrogen spectrum detection, if the determination is consistent, it can be confirmed that the substance is compound I;

[0135] iii. After the above-mentioned high-resolution mass spectrometry and carbon nuclear magnetic resonance spectroscopy, if the determination is consistent, it can be confirmed that the substance is compound I;

[0136] Of course, further testing can be done on this basis 19 F-NMR, 31 The results were further confirmed by P-NMR and UV absorption spectroscopy.

[0137] In a spectral identification test, it is usually stipulated that the spectrum of the sample is consistent with the spectrum of the standard (or reference). The consistency in this application means that it is within the error range of the above-mentioned spectral detection results.

[0138] A method for preparing compound Form 1 is provided, which is prepared by one of the following methods:

[0139] Method 1, suspension stirring method: Stir a suspension of Compound I product and the first solvent or mixed solvent at 20-55°C, collect the filter cake by filtration and dry it to obtain Compound Form 1;

[0140] Method 2, antisolvent precipitation method: add a poor solvent to a good solvent solution of Compound I product until a solid precipitate is formed, stir the suspension, filter and collect the filter cake, and dry to obtain Compound Form 1;

[0141] Method 3, cooling crystallization: Cool the good solvent solution of Compound I product, filter and collect the filter cake after solid precipitation, and dry to obtain Compound Form 1;

[0142] Method 4, evaporative crystallization: The solvent of the good solvent solution of Compound I is evaporated, the solid is precipitated, and the filter cake is collected by filtration and dried to obtain Compound Form 1;

[0143] The solvent or mixed solvent in method 1 is selected from a single solvent or mixed solvent of methanol, ethanol, methyl tert-butyl ether, isopropanol, 1,2-propylene glycol, glycerol, n-heptane, and water.

[0144] In methods 2, 3, and 4, the good solvent is independently selected from a single solvent or a mixed solvent of butanone, acetone, N,N-dimethylformamide, N,N-dimethylacetamide, ethyl acetate, dichloromethane, tetrahydrofuran, acetonitrile, dimethyl sulfoxide, isopropyl acetate, and toluene.

[0145] In the second method, the poor solvent is selected from a single solvent or a mixed solvent of water, isopropyl alcohol, and n-heptane.

[0146] Preferably,

[0147] In method one,

[0148] The mass volume ratio of the compound I product to the first solvent or mixed solvent is not greater than 60 mg:1 mL,

[0149] The mixed solvent is selected from methanol-n-heptane, methanol-water, methyl tert-butyl ether-n-heptane, ethanol-n-heptane, and ethanol-water;

[0150] In method 2,

[0151] The ratio of the volume of the poor solvent to the volume of the good solvent solution of Compound I product ranges from 0.25 to 5.00,

[0152] The combination of good solvent and poor solvent is selected from acetone + water, acetonitrile + water, dimethyl sulfoxide + water, butanone + water, acetone + isopropanol, tetrahydrofuran + isopropanol, acetone + n-heptane, ethyl acetate + n-heptane, tetrahydrofuran + n-heptane;

[0153] In method three,

[0154] The concentration of the good solvent solution of Compound I product at the higher temperature is greater than or equal to 120 mg / mL, the higher temperature is not less than 50°C and not greater than the boiling point of the solvent, and the lower temperature range after cooling is 0-25°C;

[0155] In method four,

[0156] The concentration range of the good solvent solution of Compound I product is 30-60 mg / mL, and the good solvent is selected from butanone, acetone, ethyl acetate, isopropyl acetate, dichloromethane, tetrahydrofuran, acetonitrile, toluene single solvent and butanone-acetonitrile, acetone-dichloromethane, ethyl acetate-toluene, isopropyl acetate-acetonitrile, and dichloromethane-acetonitrile mixed solvent.

[0157] In order to prepare high-quality compound crystalline form 1, the compound I product before preparation should have a certain purity. Therefore, it is preferred that the purity of the compound I product is greater than or equal to 95% as determined by HPLC.

[0158] The detection conditions are reverse phase chromatography, mobile phase A is an aqueous solution with added ammonium acetate, mobile phase B is acetonitrile, and mobile phase A and mobile phase B are gradient eluted; detection is performed with an ultraviolet detector at a detection wavelength of 254 nm, and the compound I product is dissolved in acetonitrile and then injected; the purity is calculated using the area normalization method.

[0159] Also provided is a pharmaceutical composition containing Compound Form 1.

[0160] In addition to containing Compound Form 1, the pharmaceutical composition should also be added with pharmaceutically acceptable excipients or excipients according to the characteristics of the drug, medicine, and preparation. The pharmaceutical composition can be in any dosage form for clinical use, such as tablets, suppositories, dispersible tablets, enteric-coated tablets, chewable tablets, orally disintegrating tablets, capsules, sugar-coated tablets, granules, dry powders, oral solutions, small needles for injection, freeze-dried powder for injection, or large infusions. Depending on the specific dosage form and mode of administration, the pharmaceutically acceptable excipients or excipients in the drug may include one or more of the following: diluents, solubilizers, disintegrants, suspending agents, lubricants, adhesives, fillers, flavoring agents, sweeteners, antioxidants, surfactants, preservatives, encapsulating agents, and pigments.

[0161] Taking oral solutions and injections of liquid preparations as an example, compound form 1 has a solubility of 7.5 mg / ml or above in butanone, acetone, N,N-dimethylformamide, N,N-dimethylacetamide, ethyl acetate, dichloromethane, tetrahydrofuran, acetonitrile, dimethyl sulfoxide, isopropyl acetate, toluene, methanol, and ethanol solvents, and can be used as a solvent excipient for the preparation of liquid preparations (injection or oral).

[0162] Furthermore, considering some special requirements of pharmaceutical excipients, butanone, acetone, ethyl acetate, dichloromethane, tetrahydrofuran, acetonitrile, dimethyl sulfoxide, isopropyl acetate, toluene, and methanol are not suitable as excipients or are highly toxic or irritating, and are not suitable for development as oral preparations or injection preparations. However, N,N-dimethylformamide, N,N-dimethylacetamide, ethanol, etc. are less toxic and are widely added to oral liquid preparations and injection preparations. Therefore, N,N-dimethylformamide, N,N-dimethylacetamide, and ethanol are preferred as solvent excipients for oral liquid preparations and injections.

[0163] More preferably, the pharmaceutical composition of Compound Form 1 is a liquid injection for injection, which contains N,N-dimethylacetamide and / or ethanol.

[0164] Liquid injections include concentrated injection solutions (injected after dilution), injection solutions (injected directly without dilution), and injection suspensions or injection emulsions.

[0165] Injectable suspension, a suspension preparation for injection made by dispersing the drug in a liquid is called a suspension injection.

[0166] Injectable emulsion contains water, organic solvent, drug, and emulsifier and other auxiliary materials when necessary, which are emulsified to form an injectable emulsion injection.

[0167] Whether preparing a concentrated injection solution (injection after dilution), an injection solution (injection without dilution), an injection suspension or an injection emulsion, N,N-dimethylacetamide or ethanol or N,N-dimethylacetamide and ethanol can be added as a solvent.

[0168] Based on the above content, we can see that

[0169] For a substance with sufficient purity (for example, HPLC test, area normalization method test with a purity greater than 95% or higher), if its specific chemical structure is unknown, it can be directly identified as Compound I by any one of the above-mentioned i, ii, and iii spectral tests.

[0170] For an unknown mixture (such as a solid oral preparation or a liquid injection preparation), its mass spectrum can be determined by separation and purification (such as HPLC-HRMS coupling technology), and then the specific substance can be separated and accumulated before any one of the above-mentioned i, ii, and iii spectral tests is performed. After such identification, it can be determined that the mixture contains compound I.

[0171] When a substance of sufficient purity is known to be Compound I, it can be determined to be Compound Form 1 after identification by any one of the above conditions 1, 2, and 3.

[0172] For a certain unknown mixture (such as a solid oral preparation or a liquid injection preparation), it has been confirmed that it contains Compound I. Since the present application has verified that Compound Form 1 will not be destroyed or transformed even through suspension stirring experiments, antisolvent precipitation experiments, cooling crystallization experiments, evaporation crystallization experiments, grinding experiments, etc., the Compound I extracted from the mixture by appropriate solvent extraction and evaporation will ultimately still be the original Compound Form 1. At this time, it can be determined that the mixture contains Compound Form 1 after identification through any one of the above conditions one, two, and three.

[0173] In particular, for a mixture that has been determined to contain a solid form of Compound 1, such as a solid oral preparation, the excipients commonly contained therein, such as talc, starch, dextrin, magnesium carbonate, and the like, may not interfere with XRPD detection, or their XRPD spectra can be subtracted during detection. In this way, the XRPD can be directly detected, and the presence of Compound Form 1 can be determined by direct comparison with the above-mentioned conditions by directly comparing the XRPD characteristic peaks or the characteristic peaks after subtracting the excipient background.

[0174] Provided is a purification method for Compound I or Compound Form 1, comprising a cooling crystallization method and a suspension stirring method, namely:

[0175] Putting Compound 1 or Compound Form 1 into methanol and cooling for crystallization;

[0176] Alternatively, Compound 1 or Compound Form 1 is added to ethanol and suspended with stirring;

[0177] Alternatively, Compound I or Compound Form 1 is added into methyl tert-butyl ether and suspended and stirred.

[0178] Before cooling crystallization, the heating temperature must not exceed the boiling point of methanol, preferably 45-60°C. For more efficient crystallization, the solution at high temperature can be as concentrated as possible, such as a saturated solution. For ethanol, a high concentration of 100-120 mg / ml is preferred. The cooling process can be rapid or slow, either to room temperature or to 5°C.

[0179] Suspension stirring is the process of putting Compound I or Compound Form 1 into a suitable solvent to form a state in which a solid is suspended in a liquid, similar to a slurry or paste. The effect of purification and refining is related to the solvent, the amount of solid added, and the stirring temperature. According to experiments, ethanol and methyl tert-butyl ether are suitable as solvents, and the amount added is preferably 40-60 mg per milliliter of solvent. The temperature cannot be higher than the boiling point of the solvent and can be appropriately higher than room temperature. Preferably, suspension stirring at room temperature in ethanol and suspension stirring at 50°C in methyl tert-butyl ether have good effects. The stirring time is generally selected to be greater than 2 hours, such as 24 hours, 48 ​​hours, or 72 hours.

[0180] Preferably, in the above purification method:

[0181] Cooling crystallization is carried out by cooling from 50°C;

[0182] Compound I or Compound Form 1 is placed in ethanol and suspended with stirring at room temperature, i.e., 25°C ± 2°C;

[0183] Compound I or Compound Form 1 was added into methyl tert-butyl ether and suspended with stirring at 50°C.

[0184] The present application provides a storage method for Compound 1 or the above-mentioned compound crystal form 1:

[0185] It is placed in a sealed, light-proof environment with a temperature not higher than 25℃±2℃ and a humidity not higher than 60%±5%RH;

[0186] or

[0187] It is placed at a temperature not higher than 2-8°C, sealed and protected from light.

[0188] The present application provides a storage method for Compound 1 or the above-mentioned compound crystal form 1:

[0189] It is placed in a sealed, light-proof environment with a temperature not higher than 25℃±2℃ and a humidity not higher than 60%±5%RH;

[0190] or

[0191] It is placed at a temperature not higher than 2-8°C, sealed and protected from light;

[0192] Under the above storage conditions, the shelf life, storage period, or validity period of Compound I or Compound Form 1 is at least 36 months.

[0193] In this application, unless otherwise specified, all numerical values ​​have an error range of 10%. BRIEF DESCRIPTION OF THE DRAWINGS

[0194] Figure 1 is the IR spectrum of compound 1;

[0195] Figure 2 is a UV spectrum of compound 1;

[0196] Figure 3 shows the structure of compound I. 1 H-NMR spectrum;

[0197] Figure 4 shows the 13 C-NMR spectrum;

[0198] FIG5 is a nuclear magnetic resonance DEPT 135 spectrum of compound 1;

[0199] Figure 6 shows the NMR of compound I. 19 F spectrum measurement results;

[0200] Figure 7 shows the NMR of compound I. 31 P NMR spectrum;

[0201] FIG8 is a CH HSQC-NMR spectrum of compound 1;

[0202] FIG9 is a CH HMBC-NMR spectrum of compound 1;

[0203] FIG10 is a HH COSY-NMR nuclear magnetic resonance spectrum of Compound 1;

[0204] FIG11 is a DSC curve of Compound 1 subjected to a heating-cooling-heating recrystallization experiment;

[0205] Figure 12 is a superimposed comparison of the XRPD test results of untreated Compound 1 and the sample subjected to the heating-cooling treatment in Example 2, wherein AST-3411-C is the untreated Compound 1 and 2020-04-21-cycle is the sample treated in Example 2;

[0206] FIG13 is a DSC curve of a sample of Compound 1 subjected to heating-cooling treatment in Example 2;

[0207] FIG14 is an XRPD spectrum of compound crystalline form 1;

[0208] FIG15 is a polarizing microscope photograph of compound Form 1;

[0209] FIG16 is a superposition of DSC and TGA curves of compound Form 1;

[0210] FIG17 is a superposition of the DSC and TGA curves of Form 1 of the same batch of compound in FIG16 tested again;

[0211] Figure 18 is a DVS spectrum of compound crystalline form 1;

[0212] FIG19 is a superimposed comparison of the XRPD test results of the compound crystalline form 1 before and after the DVS test;

[0213] FIG20 is a superimposed comparison of the XRPD results of compound Form 1 before and after the stability test;

[0214] FIG21 is a comparison chart of the XRPD results of suspension stirring experiments of compound Form 1 in different solvents at room temperature;

[0215] FIG22 is a comparison of the XRPD results of samples of Compound Form 1 obtained from suspension and stirring experiments at 50° C. in different solvents;

[0216] FIG23 is a PLM photograph of a sample of Compound Form 1 suspended and stirred in ethanol at 50° C. (scale bar: 50 μm);

[0217] FIG24 is a PLM photograph of a sample of Compound Form 1 obtained by suspension and stirring in methyl tert-butyl ether at 50° C. (scale: 25 μm);

[0218] FIG25 is a superimposed DSC and TGA curve of a sample of Compound Form 1 suspended and stirred in methyl tert-butyl ether at 50° C.;

[0219] FIG26 is a superimposed comparison of XRPD results of samples obtained from a suspension stirring experiment of Compound Form 1 in a mixed solvent at room temperature;

[0220] FIG27 is a comparison of the XRPD results of samples precipitated from Form 1 of Compound 1 in different anti-solvents;

[0221] FIG28 is a PLM photograph of a sample precipitated by anti-solvent precipitation in an acetonitrile / water system of Compound Form 1 (scale bar: 50 μm);

[0222] FIG29 is a PLM photograph of a sample of compound Form 1 precipitated by anti-solvent precipitation in a tetrahydrofuran / n-heptane system (scale bar: 25 μm);

[0223] FIG30 is a superimposed comparison of the XRPD results of the samples precipitated by fast cooling (Fast Cooling) and slow cooling (Slow Cooling) in methanol of Compound Form 1;

[0224] FIG31 is a superimposed comparison of XRPD results of samples obtained after slow evaporation of Compound Form 1 in different solvents;

[0225] FIG32 is a superimposed comparison of XRPD results of samples obtained after rapid volatilization of Compound Form 1 in different solvents;

[0226] FIG33 is a superimposed comparison of XRPD results of samples of Compound Form 1 obtained after different grinding times;

[0227] FIG34 is a superimposed comparison of XRPD and DSC results for selected samples. In the XRPD and DSC diagrams, the treatment processes from right to left are Fast evaporation in THF, RT-Slurry in Hept, Ref.Form 1, and Well-Plate, respectively. That is, the treatment processes are fast evaporation in tetrahydrofuran, slurrying in n-heptane, untreated compound Form 1, and slow evaporation in acetone / dichloromethane, respectively.

[0228] Figure 35 is a PLM photo of the selected sample (AST-3411-C, i.e., Ref. Form 1);

[0229] Figure 36 is a superimposed comparison of the XRPD test results of compound Form 1 under different storage conditions after 6 months of storage. From top to bottom in the figure, the XRPD spectra are: initial, 2-8°C, sealed, dark conditions, 25°C ± 2°C, 60% ± 5% RH, dark conditions, and 40°C ± 2°C, 75% ± 5% RH, dark conditions;

[0230] Figure 37 is a superimposed comparison of the XRPD test results of compound Form 1 under different storage conditions after 12 months of storage. From top to bottom in the figure, the XRPD spectra are: initial, 2-8°C, sealed, dark conditions, 25°C ± 2°C, 60% ± 5% RH, dark conditions;

[0231] Figure 38 is a superimposed comparison of the XRPD test results of Compound Form 1 under different storage conditions after 24 months of storage. From top to bottom in the figure, the XRPD spectra are: initial, 2-8°C, sealed, dark conditions, 25°C ± 2°C, 60% ± 5% RH, dark conditions;

[0232] Figure 39 is a superimposed comparison of the XRPD test results of compound Form 1 under different storage conditions after 36 months of storage. From top to bottom in the figure, the XRPD spectra are: initial, 2-8°C, sealed, dark conditions, 25°C ± 2°C, 60% ± 5% RH, dark conditions;

[0233] Figure 40 is a superimposed comparison of the XRPD test results of Compound Form 1 after storage for different lengths of time at 2-8°C, sealed, and protected from light. From top to bottom in the figure are the spectra at the initial stage, after 6 months, 12 months, 24 months, and 36 months respectively;

[0234] Figure 41 is a superimposed comparison of the XRPD test results of compound form 1 after storage for different lengths of time at 25°C ± 2°C, 60% ± 5% RH, and in the dark. From top to bottom in the figure, the spectra are the initial, 6 months, 12 months, 24 months, and 36 months later.

[0235] Ref. Form 1 in the accompanying drawings is the spectrum of the compound crystal form 1 in Example 3 (its XRPD spectrum is shown in Figure 14), and this batch of samples was used as a control for each XRPD spectrum measurement. DETAILED DESCRIPTION

[0236] The present invention is described below with reference to specific examples. Those skilled in the art will appreciate that these examples are only used to illustrate the present invention and are not intended to limit the scope of the present invention in any way.

[0237] The experimental methods in the following examples are conventional methods unless otherwise specified. The medicinal materials, reagents, etc. used in the following examples are commercially available products unless otherwise specified.

[0238] Unless otherwise specified, the following tests were performed using the following general analysis methods.

[0239] XRPD

[0240] The test was performed using a PIXceI 1D The X-ray powder diffractometer used as the detector was Panalytical Empyrean (Malvern Panalytical Ltd). The test conditions were Cu target Kα radiation, a step size expressed in 2θ° of 0.013°, and an X-ray tube voltage and current of 45 kV and 40 mA, respectively.

[0241] All samples were ground (1-2 minutes) and sieved (200 mesh) before testing.

[0242] Before preparing the test samples, the samples were vacuum dried at 25°C and dried with phosphorus pentoxide for 2 hours.

[0243] DSC

[0244] The test was conducted using a Discovery DSC 250 differential scanning calorimeter (TA Instruments, US). The sample was weighed and placed in a perforated, sealed aluminum pan, and the weight was accurately recorded. Unless otherwise specified, the heating program was to equilibrate the sample at 25°C and then heat it at a rate of 10°C / min to the final temperature.

[0245] TGA

[0246] The test was performed using a thermogravimetric analyzer TGA 55 (TA Instruments, US). The sample was placed in an open aluminum sample pan, automatically weighed in a heating furnace, and heated from room temperature to the final temperature at a rate of 10°C / min.

[0247] DVS

[0248] Water adsorption / desorption data were collected using a Vsorp dynamic water vapor sorption instrument (ProUmid GmbH & Co. KG, Germany). The sample was placed in a tared sample chamber and automatically weighed. The instrument parameters were set as follows.

[0249] Sample temperature: 25°C

[0250] Cycle time: 10min

[0251] Minimum time for each cycle: 50 minutes

[0252] Maximum time per cycle: 120 minutes

[0253] Weight limit: 100%

[0254] Equilibrium conditions: 0.01% / 40min

[0255] Environmental cycle #1: 0%-0%, 1 step, 40°C, 3 hours

[0256] Environmental cycle #2: 0%-90%, 9 steps, 25°C

[0257] Environmental cycle #3: 80%-0%, 8 steps, 25°C

[0258] Adsorption:0,10,20,30,40,50,60,70,80,90

[0259] Desorption:80,70,60,50,40,30,20,10,0

[0260] PLM

[0261] The instrument used for analysis was Polarizing Microscope ECLIPSE LV100POL (Nikon, JPN).

[0262] HPLC

[0263] Chromatographic column CORTECS C18+2.7um 4.6*150mm

[0264] Mobile phase A: 10mM NH4OAc in H2O B: ACN

[0265] Gradient (time / mobile phase B%): 0 / 10, 8 / 58, 18 / 58, 22 / 90, 30 / 90, 30.1 / 10

[0266] Column temperature 30℃

[0267] Detector DAD; 254 nm

[0268] Flow rate 1.0 mL / min

[0269] Injection volume 5 μL

[0270] Run time 30.1 minutes

[0271] Cleaning time 4 minutes

[0272] Diluent ACN

[0273] NMR

[0274] The tests were performed using an AVANCE III 400 MHz nuclear magnetic resonance spectrometer (Bruker), including DEPT, HSQC, HMBC, and COSY tests.

[0275] UV

[0276] The test was performed using a UV-2600 ultraviolet-visible spectrophotometer (Shimadzu Corporation, Japan).

[0277] IR

[0278] The test was performed using an IR Tracer 100 Fourier transform infrared spectrometer (Shimadzu, Japan).

[0279] HRMS

[0280] The test was performed using an Acquity Xevo G2-XS QT of UPLC / MS ultra-high performance liquid chromatography-mass spectrometry system (Waters).

[0281] Unless otherwise specified, the following experiments were performed according to the following instructions.

[0282] Rough solubility test

[0283] 5.0±0.5 mg of Compound Form 1 was accurately weighed into a sample bottle, and then a specific solvent was gradually added at room temperature (25° C.) until the solid was dissolved or the total volume reached 8 ml.

[0284] Suspension stirring experiment

[0285] Compound Form 1 (20 or 30 mg) was added to 0.5 mL of the selected solvent or mixed solvent, and the suspension was stirred at room temperature or 50° C. The sample collected by filtration was subjected to XRPD analysis.

[0286] Antisolvent precipitation experiment

[0287] Compound Form 1 (approximately 30 mg) was dissolved in 0.2-0.5 mL of a solvent (butanone, acetone, ethyl acetate, isopropyl acetate, tetrahydrofuran, acetonitrile, and dimethyl sulfoxide) at room temperature. 0.2-2.5 mL of an antisolvent (water, isopropanol, and n-heptane) was then slowly added to the solution until a solid precipitated. The suspension was stirred at room temperature for 1-20 hours. The collected solid was filtered and analyzed by XRPD.

[0288] Cooling crystallization experiment

[0289] Slowly cool to room temperature: Compound Form 1 (approximately 60 mg) was dissolved in 0.5 mL of methanol at 50°C. The clear solution (filtrate) was then transferred to room temperature for 20 hours. The solid collected by filtration was subjected to XRPD analysis.

[0290] Rapid Cooling: Compound Form 1 (approximately 60 mg) was dissolved in 0.5 mL of methanol at 50°C. The clear solution (filtrate) was then transferred to a refrigerator and rapidly cooled to ~5°C for 20 hours. The solid collected by filtration was subjected to XRPD analysis.

[0291] Evaporation crystallization experiment

[0292] Slow Evaporation: Compound Form 1 (approximately 30 mg) was dissolved in 1 mL of the selected solvent. The stock solution (filtrate) was distributed in a 96-well plate. Each well contained two 100 μL aliquots of the filtrate. The 96-well plate was covered with a perforated film and evaporated at room temperature in a laboratory fume hood.

[0293] Rapid evaporation: Compound Form 1 (approximately 30 mg) was dissolved in 200-500 μL of a selected solvent and evaporated with nitrogen to condense the solution to precipitate a solid, which was then subjected to XRPD analysis.

[0294] Stability test

[0295] Compound Form 1 was placed under two conditions: 40°C / 75% RH and 60°C / closed for one week, and then the samples were subjected to XRPD and HPLC analysis.

[0296] Abbreviations

[0297] X-ray Powder Diffraction, XRPD

[0298] Thermogravimetric Analysis, TGA

[0299] Differential Scanning Calorimetry, DSC

[0300] Dynamic Vapor Sorption is dynamic water adsorption and desorption analysis, DVS

[0301] Polarized Light Microscopy, PLM

[0302] High Performance Liquid Chromatography, HPLC

[0303] Nuclear Magnetic Resonance (spectroscopy) is nuclear magnetic resonance (spectroscopy), NMR (S)

[0304] DEPT spectroscopy (Distortionless Enhancement by Polarization Transfer), also known as distortionless polarization transfer technology, is a carbon nuclear magnetic resonance spectroscopy detection technique used to distinguish primary, secondary, tertiary, and quaternary carbons in carbon spectra. In a DEPT 45 spectrum, all methine, methylene, and methyl peaks are oriented upward; in a DEPT 90 spectrum, only methine peaks are observed. In a DEPT 135 spectrum, methyl and methine peaks are oriented upward, while methylene peaks are inverted.

[0305] HH COSY (HH correlation spectroscopy), also known as hydrogen-hydrogen correlation spectrum, is a coupling correlation spectrum between protons in the same coupling system, and the mutually coupled hydrogen nuclei give rise to cross peaks.

[0306] HSQC (heteronuclear singular quantum correlation) is a type of NMR CH COSY spectrum that gives directly connected carbon-hydrogen relationships.

[0307] HMBC (Heteronuclear Multiple Bond Correlation) is 1 The heteronuclear multi-carbon correlation spectrum of H 1 H nuclei and long-range coupling 13 C core is associated.

[0308] Ultraviolet spectrum is ultraviolet absorption spectrum, UV

[0309] Infrared Ray spectrum is infrared absorption spectrum, IR

[0310] Mass Spectrometry, MS

[0311] High Resolution Mass Spectrometry, HRMS

[0312] overlap peaks

[0313] spectroscopy

[0314] Example 1 Preparation of Compound I

[0315] According to the synthesis process of compound 16 (i.e., compound 1) in the previously disclosed document WO2020228685, the synthetic preparation route of compound 1 is as follows:

[0316] The synthesis method of compound 16 in the above literature was appropriately amplified and synthesized as follows:

[0317] "After the reaction is completed, cool to room temperature, filter with diatomaceous earth, wash the solid with DCM, concentrate the mother liquor, add 1.5 mL of anhydrous ether for crystallization, and filter to obtain pure compound No. 16." HPLC test showed a purity of 95.42%, which is greater than 95%. That is, the final product is pure enough for subsequent tests: HRMS, NMR, IR, and UV.

[0318] The specific NMR, IR, and UV spectra are shown in Figures 1 to 10.

[0319] Example 2 Heating-cooling-heating recrystallization experiment by DSC

[0320] Using DSC for heating-cooling-heating recrystallization, studies have shown that this process can detect different crystal forms in homogeneous polycrystals.

[0321] A sample of Compound I prepared in Example 1 (vacuum dried at 25°C with phosphorus pentoxide as the drying aid) was heated to 150°C (corresponding to AB in Figure 11 ) at a rate of 10°C / min, rapidly equilibrated to -20°C (corresponding to BC in Figure 11 ), then heated to 60°C (corresponding to CD in Figure 11 ) at a rate of 10°C / min, then heated to 120°C (corresponding to EF in Figure 11 ) at a rate of 0.5°C / min, and then held at this temperature for 5 minutes. Its DSC curve is shown in Figure 11 .

[0322] DSC results show peaks at 80 and 140°C:

[0323] The peak at 140 °C is presumed to be the melting endotherm, which may correspond to a crystal form (named crystal form 1);

[0324] The peak at 80°C is a shoulder peak before the melting peak, which may be a potential other crystal form (named as crystal form x).

[0325] The untreated compound I and the sample subjected to the above-mentioned DSC heating-cooling treatment were subjected to XRPD testing, and the superposition comparison of the two is shown in Figure 12.

[0326] The sample subjected to the above DSC heating-cooling treatment was subjected to DSC testing. The results are shown in FIG13 , which shows an endothermic melting peak with a peak value of 138.70° C., a corresponding onset point, i.e., a melting point, of 137.56° C., and an enthalpy value ΔH of 68.133 J / g.

[0327] The results showed that the sample subjected to the above DSC heating-cooling treatment may be Form 1, and it is very likely that most or all of the Compound I sample prepared in Example 1 above is Form 1.

[0328] Example 3 The sample of Compound I prepared in Example 1 was purified and then subjected to crystal characterization

[0329] The sample prepared in Example 1 was refined as follows:

[0330] Take an appropriate amount of compound I sample and add it to the reaction bottle. Add an appropriate amount of acetone, stir and reflux under reflux to 50°C. After complete dissolution, add an appropriate amount of water, continue heating and stirring, then cool to 5-15°C until solid precipitates and stir and crystallize for 4 hours. Filter with suction, rinse the filter cake with acetone, and collect the filter cake.

[0331] The filter cake was dried under reduced pressure at 45°C and dried with phosphorus pentoxide to obtain an off-white solid with a purity of 97.56% as determined by HPLC.

[0332] The purified samples were characterized by XRPD, DSC, TGA, DVS and PLM. The characterization data are summarized in Table 17 and Figures 14 to 19.

[0333] As shown in FIG14 , the XRPD spectrum has the characteristic peaks represented by the above 2θ°, and the relative intensities thereof are close to the values ​​shown in Table 17 below.

[0334] Table 17: Diffraction characteristic peaks, interplanar distances, and relative intensities obtained from XRPD test of compound form 1

[0335] Under a polarizing microscope, as shown in Figure 15 , the crystals are rod-shaped.

[0336] Obviously, according to the DSC of Figures 16 and 17 , it can be determined that the compound prepared after purification is the above-mentioned crystal form 1.

[0337] The DSC curve in FIG16 shows an endothermic melting peak with a peak value of 137.90° C., a corresponding onset point, or melting point, of 136.34° C., and an enthalpy value ΔH of 74.710 J / g.

[0338] The DSC curve in FIG17 shows an endothermic melting peak with a peak value of 140.19° C., a corresponding onset point, or melting point, of 138.57° C., and an enthalpy value ΔH of 83.701 J / g.

[0339] TGA showed a weight loss of 0.8% in the range of 120-150°C: the weight loss in Figure 16 was 0.774% and in Figure 17 was 0.350%, which may be due to the loss of residual solvent.

[0340] According to the DVS results in Figure 18, the moisture absorption of Compound Form 1 from 0.0% RH to 80% RH is 0.13%. Therefore, Compound Form 1 is not hygroscopic. According to Figure 19, the crystal form remains unchanged after the DVS test.

[0341] Example 4 Solubility Test of Compound Form 1

[0342] The solubility of Compound Form 1 was roughly measured by visual observation at room temperature. The results are shown in Table 1.

[0343] It can be seen that compound form 1 has high solubility (>100 mg / mL) in butanone, acetone, ethyl acetate, dichloromethane, tetrahydrofuran, acetonitrile, dimethyl sulfoxide, N,N-dimethylformamide, and N,N-dimethylacetamide, and is almost insoluble in glycerol, water, and n-heptane (<1 mg / mL).

[0344] Example 5 Solid Stability Experiment of Compound Form 1

[0345] An appropriate amount of compound Form 1 was placed under two conditions: 40°C / 75% RH and 60°C / closed for one week, and then tested by HPLC and XRPD respectively after one week.

[0346] The XRPD test results are shown in FIG20 . There is no substantial difference between the three, and all are compound crystal form 1.

[0347] The HPLC purity results are shown in Table 18 below, indicating that no degradation occurred under the test conditions.

[0348] Table 18: HPLC results of solid stability test of compound crystalline form 1

[0349] Therefore, compound Form 1 is physically and chemically stable after being placed under both 40°C / 75% RH and 60°C / closed conditions for one week.

[0350] It can be seen from Examples 1 to 5 above that compound Form 1 has a high solubility in a suitable solvent, and the formulation design of liquid preparations of different concentrations can be achieved through solvent combination during formulation development; at the same time, it is not hygroscopic and also exhibits good stability in a high temperature and high humidity environment. Therefore, compound Form 1 is suitable for later formulation development.

[0351] Taking into account the solubility and the chemical inertness, safety and non-toxicity requirements of pharmaceutical excipients, preferably, when compound Form 1 is used to prepare a liquid injection for injection, the solvent recommended is N,N-dimethylacetamide, ethanol or N,N-dimethylformamide and a mixed solvent thereof, which can be prepared as an injection concentrated solution (injected after dilution), an injection (injected directly without dilution), an injection suspension or an injection emulsion.

[0352] According to the heating-cooling-heating recrystallization experiment in Example 2, the prepared compound I may contain another crystalline form x corresponding to 80°C in the DSC curve. In order to explore whether compound form 1 can be converted into this crystalline form x and whether crystalline form x is stable, the following suspension stirring experiment, antisolvent precipitation experiment, cooling crystallization experiment, evaporation crystallization experiment, and grinding experiment were performed.

[0353] Example 6 Suspension stirring experiment (slurry)

[0354] Single solvent

[0355] About 20 mg of compound Form 1 was suspended in 0.5 mL of solvent and stirred at room temperature for 3 days; about 30 mg of compound Form 1 was suspended in 0.5 mL of solvent and stirred at 50°C for 3 days.

[0356] The solid was collected and subjected to XRPD analysis. The specific results are shown in Figures 21 and 22. Except for the suspension and stirring in MeOH solvent at 50°C, the compound crystalline form 1 was obtained in all solvents. The results are shown in Table 19.

[0357] Table 19: Results of suspension and stirring of compound form 1 in a single solvent

[0358] Some samples were subjected to PLM testing, as shown in Figures 23 and 24, which showed that most of the samples appeared as fine particles and / or agglomerates, and large crystals were observed from EtOH and MTBE, which may be potential solvents for crystallization.

[0359] The sample obtained by slurrying in MTBE at 50°C was further characterized by TGA and DSC, as shown in Figure 25: TGA showed a weight loss of 0.401% in the temperature range of 120-150°C, which may be caused by solvent encapsulation; DSC showed an endothermic melting peak with a peak value of 139.83°C, corresponding to an onset point, i.e., a melting point, of 138.97°C, and an enthalpy value ΔH of 81.341 J / g, which was attributed to the melting peak of compound form 1.

[0360] mixed solvents

[0361] An appropriate amount of starting material was weighed and suspended in a 1:1 solvent mixture by volume. A slurrying experiment was performed at room temperature for 3 days. All experiments yielded Form 1. The results are shown in Table 20 and Figure 26.

[0362] Table 20: Results of suspension and stirring of compound crystal form 1 in mixed solvents

[0363] Through the suspension stirring experiments of compound crystal form 1 in different solvents and at different temperatures, it can be seen that compound crystal form 1 cannot be converted into other crystal forms. After suspension stirring treatment in different solvents, compound crystal form 1 remains, and no crystal form x appears.

[0364] Example 7 Antisolvent precipitation experiment

[0365] Compound Form 1 was dissolved in 0.2-0.5 mL of the selected solvent, and the antisolvent was added until a solid precipitated. The results are shown in Table 21 and Figure 27. All antisolvent precipitation experiments yielded Form 1, except for the MEK / water and THF / water conditions, which produced an oil. Relatively large crystals were obtained under ACN / water and THF / Hept conditions, as shown in Figures 28 and 29.

[0366] Table 21: Antisolvent precipitation results of compound form 1

[0367] Through the anti-solvent precipitation experiment of compound crystal form 1 in different solvent combinations, it can be seen that compound crystal form 1 cannot be converted into other crystal forms. After anti-solvent precipitation treatment with different solvents, compound crystal form 1 is still obtained, and no crystal form x appears.

[0368] Example 8 Cooling crystallization experiment

[0369] Cooling crystallization was performed using different cooling rates. The experimental details and results are shown in Table 22 and Figure 30. All the crystals obtained in the experiment were Compound Form 1.

[0370] Table 22: Cooling crystallization results of compound form 1

[0371] Through the cooling crystallization experiment of compound form 1 in MeOH solution at different rates, it can be seen that compound form 1 cannot be converted into other forms. After cooling crystallization treatment at different rates, compound form 1 is still compound form 1, and no form x appears.

[0372] Example 9 Evaporation Crystallization Experiment

[0373] Slow evaporation

[0374] Evaporation crystallization was performed at different rates. Slow evaporation was performed on a 96-well plate covered with a perforated film. Samples from selected wells were subjected to XRPD analysis, and all obtained were Compound Form 1. The results are summarized in Table 23, and the XRPD results are shown in Figure 31.

[0375] Table 23: Slow evaporation experimental results of compound form 1 in different solvents

[0376] Rapid evaporation

[0377] Rapid evaporation was also used for crystallization experiments. The prepared solutions were dried using N2 blowing, and the resulting solids were subjected to XRPD analysis. All evaporation experiments produced Form 1. The results are summarized in Table 24, and the XRPD results are shown in Figure 32.

[0378] Table 24: Rapid volatilization results of compound crystal form 1 in different solvents

[0379] Through the evaporation crystallization experiments of compound form 1 at different rates in different solvent combinations, it can be seen that compound form 1 cannot be converted into other forms. After evaporation crystallization treatment at different rates, compound form 1 is still compound form 1, and no form x appears.

[0380] Example 10 Grinding Experiment

[0381] Compound Form 1 was ground for 2 minutes and 5 minutes, respectively, and then XRPD analysis was performed. The results are shown in Figure 33. The XRPD spectrum of Compound Form 1 remained essentially unchanged after grinding, with only some peak intensities weakening, indicating a decrease in crystallinity.

[0382] Through experiments on compound crystal form 1 at different grinding times, it can be seen that compound crystal form 1 cannot be converted into other crystal forms. After 2-5 minutes of grinding, compound crystal form 1 is still obtained, and no crystal form x appears; however, the crystallinity decreases. It is speculated that if the grinding time is long enough, the crystal form may disappear and turn into a non-crystalline state, such as amorphous.

[0383] Example 11 DSC, XRPD and PLM testing experiments of selected samples

[0384] DSC, XRPD, and PLM tests were performed on the selected suspension stirring, rapid evaporation, and slow evaporation samples. Compound Form 1 was heated to 300°C at a rate of 10°C / min. The results are shown in Figures 34 and 35:

[0385] Differences in the onset melting temperatures were observed for samples obtained by rapid evaporation in tetrahydrofuran.

[0386] The sample obtained by slurrying (suspending and stirring) in n-heptane was found to have a small shoulder peak before the melting peak, which may be due to the relatively low crystallinity and the presence of particles of different sizes, or the presence of other crystal forms may also cause the baseline to decrease before melting.

[0387] It can be seen from this that although different suspension stirring, rapid evaporation and slow evaporation operations on compound crystal form 1 will not lead to changes in the crystal form, they may lead to changes in crystallinity, crystal particle size and crystal particle size distribution.

[0388] Example 12 HPLC test of selected samples

[0389] The selected samples were tested by HPLC to observe the purification effect. The specific results are shown in Table 25 below.

[0390] Compound Form 1 exhibited the most significant purification effect when suspended and stirred in methyl tert-butyl ether at 50°C for three days, with purity increasing from 95.42% to 97.72%. Compound Form 1 obtained by suspension and stirring in water at 50°C for three days, as well as by cooling in methanol and stirring in ethanol at room temperature for three days, exhibited good purification effects, with purity increasing to 97.0%-97.3%.

[0391] It can be seen that the four methods of slow cooling in methanol, rapid cooling in methanol (the starting temperature of both rapid cooling and slow cooling is 50°C, see Example 8 for details), suspension stirring in ethanol at room temperature, and suspension stirring in methyl tert-butyl ether at 50°C can be used to purify and refine Compound I and Compound Form 1.

[0392] Table 25: Summary of HPLC test results of samples obtained by different treatment methods

[0393] Example 13 Long-term stability experiment of compound crystal form 1

[0394] An appropriate amount of Compound Form 1 was placed in a stability test chamber at 2-8°C in the dark (sealed in an aluminum foil bag on the outer packaging), and in a stability test chamber set at 25°C ± 2°C / 60% ± 5% RH. HPLC and XRPD were tested at the initial time T0, 3 months, 6 months, 9 months, 12 months, 18 months, 24 months, and 36 months (part of the samples were tested). The specific results are shown in Table 26. Furthermore, an appropriate amount of Compound Form 1 was placed in a stability test chamber set at 40°C ± 2°C / 75% ± 5% RH. HPLC and XRPD were tested at the initial time T0, 1 month, 3 months, and 6 months. The specific results are shown in Table 26 and Figures 36 to 41.

[0395] In the long-term stability experiments, XRPD was performed using a D2 Phaser X-ray diffractometer (Bruker). The test conditions used Cu target Kα radiation, a scanning range of 3° to 40° in 2θ, a step size of 0.02° in 2θ°, and an X-ray tube voltage and current of 30 kV and 30 mA, respectively. Water content was determined using the Karl Fischer method.

[0396] Table 26: Long-term stability test results of compound crystal form 1 stored in the dark at different temperatures

[0397] From the analysis of Figure 36, it can be seen that after 6 months of storage, the XRPD spectra of the three samples at 2-8°C, sealed, and light-proof conditions, 25°C±2°C, 60%±5%RH, and light-proof conditions, and 40°C±2°C, 75%±5%RH, and light-proof conditions have almost no obvious changes compared with the initial spectra, and the spectra differences under the three different storage conditions are not obvious.

[0398] From the analysis of Figure 37, it can be seen that after 12 months of storage, the XRPD spectra of the two samples under the conditions of 2-8°C, sealed, and protected from light, and 25°C±2°C, 60%±5%RH, and protected from light have almost no obvious changes compared with the initial spectra, and the difference in the spectra under the two different storage conditions is not obvious.

[0399] From the analysis of Figure 38, it can be seen that after 24 months of storage, the XRPD spectra of the two samples at 2-8°C, sealed, and light-proof conditions, and at 25°C±2°C, 60%±5%RH, and light-proof conditions are compared with the initial spectra. It can be found that compared with the initial spectra, the intensities of some characteristic peaks are significantly weakened or even disappear, but the corresponding characteristic peaks (2θo) 9.1±0.2, 18.5±0.2, 20.5±0.2, 21.3±0.2, and 25.8±0.2 are still clearly present; and the difference in the spectra under the two different storage conditions is not obvious.

[0400] From the analysis of Figure 39, it can be seen that after 36 months of storage, the XRPD spectra of the two samples under 2-8°C, sealed, and light-proof conditions, and 25°C±2°C, 60%±5%RH, and light-proof conditions are compared with the initial spectra. It can be found that compared with the initial spectra, the intensities of some characteristic peaks are significantly weakened or even disappear, but the corresponding characteristic peaks (2θo) 9.1±0.2, 18.5±0.2, 20.5±0.2, 21.3±0.2, and 25.8±0.2 are still clearly present; and the difference in the spectra under the two different storage conditions is not obvious.

[0401] Comparing the spectra after different storage times under the same storage conditions. Analysis of Figures 40 and 41 clearly reveals that with the extension of storage time, the intensity of certain characteristic peaks in the XRPD spectra significantly weakens, and the degree of weakening is positively correlated with storage time: the longer the storage time, the greater the weakening of the characteristic peak intensity. In comparison, Figure 41 appears to have a greater degree of weakening, which means that the high temperature and high humidity environment may promote the reduction of the crystallinity of Compound Form 1, that is, the high temperature and high humidity environment may promote the transformation of Compound Form 1 from a crystalline state to an amorphous state (such as amorphous).

[0402] The above phenomena indicate that the crystallinity of compound crystal form 1 may decrease with the extension of storage time, and the high temperature and high humidity in the storage environment will promote the transformation of compound crystal form 1 from a crystalline state to an amorphous state (such as amorphous).

[0403] There was no significant change in moisture content, purity, and XRPD. Therefore, it can be determined that compound crystal form 1 is stable when stored for 36 months at 2-8°C, sealed, and protected from light. Although XRPD shows that its crystallinity may be reduced, it is still compound crystal form 1.

[0404] There was no significant change in moisture content, purity, and XRPD. Therefore, it can be determined that compound form 1 is stable when stored for 36 months at 25°C ± 2°C, 60% ± 5% RH, and in the dark. Although XRPD shows that its crystallinity may be reduced, it is still compound form 1.

[0405] There was no significant change in moisture content, purity, and XRPD. Therefore, it can be determined that compound form 1 is stable when stored for 6 months at 40°C ± 2°C, 75% ± 5% RH, and in the dark.

[0406] The experimental data of the above embodiments can support the storage conditions of Compound I and Compound Form 1: the temperature is not higher than 25°C ± 2°C, and the humidity is not higher than 60% ± 5% RH under sealed and light-proof conditions; or it is placed at a temperature not higher than 2-8°C, and sealed and light-proof conditions, and has a stability of at least 36 months, that is, its HPLC purity and moisture content do not change significantly within 36 months: the test data in different months fluctuate, and this fluctuation is an error. In fact, observing the data curve composed of 8 data points from 0 to 36 months, it can be found that the HPLC purity and moisture content do not actually change. That is to say, Compound I and Compound Form 1 have a shelf life or storage period or validity period of at least 36 months when stored under the above conditions.

[0407] Obviously, by observing the data curve formed by connecting 8 data points from 0 to 36 months, it can be found that the HPLC purity and moisture content are actually unchanged. That is to say, based on the existing 36-month HPLC purity and moisture content data, those skilled in the art can infer that even if the storage time is further extended, such as extending it by 12 months, that is, 48 ​​months, extending it by 18 months, that is, 54 months, and extending it by 24 months, that is, 60 months, the storage method of Compound I and Compound Form 1 provided by the present invention can still ensure the stability of Compound I or Compound Form 1.

Claims

1. Compound crystal form 1, the structural formula of the compound is as shown in Formula I, and the compound crystal form 1 satisfies one of the following conditions: Condition 1: The compound crystal form 1 is tested by X-ray powder diffraction. The test conditions are Cu target Kα ray, and the scanning range of 2θ° is from 3° to 40°. The obtained spectrum shows characteristic peaks at 9.1±0.2, 18.5±0.2, 20.5±0.2, 21.3±0.2, and 25.8±0.2 in terms of 2θ°; Condition 2: The compound crystal form 1 is tested by differential scanning calorimetry. The temperature is increased from 25°C to 180°C at a rate of 10°C / min, and the heating curve is recorded. It shows an endothermic melting peak with a peak value range of 135.27±0.05°C to 140.19±0.05°C, corresponding to a starting point, i.e., a melting point range of 131.09±0.05°C to 138.97±0.05°C, and an enthalpy value ΔH range of 48.647±0.039 J / g to 83.701±0.067 J / g; Condition 3: The solubility is tested through the following operations: Accurately weigh 5.0 ± 0.5 mg of Compound Crystal Form 1 into a sample vial, and then gradually add a specific solvent at room temperature of 25 °C until the solid dissolves or add up to a total of 8 mL. The results are shown in the following table: Among them, A solubility data > 100 mg / ml means that when the volume of the added solvent is less than 0.05 ml, all of it is dissolved until no solid can be observed with the naked eye, > 50 mg / ml means that when the volume of the added solvent is greater than 0.05 ml and less than 0.10 ml, all of it is dissolved until no solid can be observed with the naked eye, < 0.6 mg / ml means that even after the volume of the added solvent exceeds the total amount of 8 ml in the sample bottle, the solid is still visible to the naked eye and cannot be completely dissolved, The others ~33, ~12.5, ~7.5, ~4.5, ~3, ~2.9, ~2 indicate that the solubility is 33±3.3 mg / ml, 12.5±1.25 mg / ml, 7.5±30.75 mg / ml, 4.5±0.45 mg / ml, 3±0.3 mg / ml, 2.9±0.29 mg / ml, 2±0.2 mg / ml, that is, after adding a specific volume of a specific solvent, all of it is dissolved until no solid can be observed with the naked eye.

2. Compound crystal form 1, the structural formula of the compound is as shown in Formula I, and this compound crystal form 1 simultaneously satisfies the following three conditions: Condition 1: The compound crystal form 1 is tested by X-ray powder diffraction. The test conditions are Cu target Kα ray, and the scanning range of 2θ° is from 3° to 40°. The obtained spectrum shows characteristic peaks at 9.1±0.2, 18.5±0.2, 20.5±0.2, 21.3±0.2, and 25.8±0.2 in terms of 2θ°; Condition 2: The compound crystal form 1 is tested by differential scanning calorimetry. The temperature is increased from 25°C to 180°C at a rate of 10°C / min, and the heating curve is recorded. It shows an endothermic melting peak with a peak value range of 135.27±0.05°C to 140.19±0.05°C, corresponding to a starting point, i.e., a melting point range of 131.09±0.05°C to 138.97±0.05°C, and an enthalpy value ΔH range of 48.647±0.039 J / g to 83.701±0.067 J / g; Condition 3: The solubility is tested through the following operations: Accurately weigh 5.0 ± 0.5 mg of Compound Crystal Form 1 into a sample vial, and then gradually add a specific solvent at room temperature of 25 °C until the solid dissolves or add up to a total volume of 8 mL. The results are shown in the following table: Among them, A solubility data > 100 mg / ml means that when the volume of the added solvent is less than 0.05 ml, all of it is dissolved until no solid can be observed with the naked eye, > 50 mg / ml means that when the volume of the added solvent is less than greater than 0.05 ml and less than 0.10 ml, all of it is dissolved until no solid can be observed with the naked eye, <0.6 mg / ml means that after adding a solvent exceeding the total volume of 8 ml of the sample vial, the solid is still visible to the naked eye and cannot be completely dissolved. Others ~33, ~12.5, ~7.5, ~4.5, ~3, ~2.9, ~2 indicate that the solubility is 33 ± 3.3 mg / ml, 12.5 ± 1.25 mg / ml, 7.5 ± 30.75 mg / ml, 4.5 ± 0.45 mg / ml, 3 ± 0.3 mg / ml, 2.9 ± 0.29 mg / ml, 2 ± 0.2 mg / ml, that is, after adding a specific volume of a specific solvent, it is completely dissolved until no solid can be observed with the naked eye.

3. The crystalline form 1 of the compound according to claim 1 or 2, wherein The spectrum in condition one expressed in 2θ° also has a characteristic peak of 17.2 ± 0.2; or The spectrum in condition one expressed in 2θ° also has characteristic peaks of 12.9 ± 0.2 and 19.5 ± 0.2; or The spectrum in condition one expressed in 2θ° also has characteristic peaks of 15.7 ± 0.2, 16.8 ± 0.2, 18.0 ± 0.2, 26.8 ± 0.2, and 29.8 ± 0.2; or The spectrum in condition one expressed in 2θ° also has a characteristic peak of 23.7 ± 0.2; or The spectrum in condition one expressed in 2θ° also has characteristic peaks of 12.9 ± 0.2, 15.7 ± 0.2, 16.8 ± 0.2, 18.0 ± 0.2, 19.5 ± 0.2, 23.7 ± 0.2, 24.6 ± 0.2, 24.9 ± 0.2, 26.8 ± 0.2, and 29.8 ± 0.

2.

4. The crystalline form 1 of the compound according to claim 1 or 2, wherein The spectrogram in Condition 1 has characteristic peaks listed in the following table: X-ray powder diffraction tests were performed using an X-ray powder diffractometer Panalytical Empyrean (Malvern Panalytical Ltd) equipped with a PIXceI 1D detector, with a step size of 0.013° expressed in 2θ°, and the X-ray tube voltage and current were 45 kV and 40 mA, respectively; or The spectrogram in Condition 1 has characteristic peaks listed in the following table and the corresponding interplanar spacing: X-ray powder diffraction tests were carried out using an X-ray powder diffractometer Panalytical Empyrean (Malvern Panalytical Ltd) equipped with a PIXceI 1D detector. The step size expressed in 2θ° was 0.013°, and the X-ray tube voltage and current were 45 kV and 40 mA respectively; or The spectrogram in Condition 1 has characteristic peaks listed in the following table, as well as the corresponding interplanar spacing and relative intensity: X-ray powder diffraction tests were carried out using an X-ray powder diffractometer Panalytical Empyrean (Malvern Panalytical Ltd) equipped with a PIXceI 1D detector. The step size expressed in 2θ° was 0.013°, and the X-ray tube voltage and current were 45 kV and 40 mA, respectively.

5. The crystalline form 1 of the compound according to claim 1 or 2, the spectrum under condition 1 is shown in Figure 14, and the X-ray powder diffraction test is carried out using an X-ray powder diffractometer Panalytical Empyrean (Malvern Panalytical Ltd) equipped with a PIXceI 1D detector. The step size expressed in 2θ° is 0.013°, and the X-ray tube voltage and current are 45 KV and 40 mA respectively.

6. The crystalline form 1 of the compound according to claim 1 or 2, wherein The difference between the peak value and the starting point in condition two, that is, the melting point, is in the range of 0.86 to 4.18 °C, preferably 0.86 to 1.86 °C; or The heating rate curves in Condition 2 are selected from the following 8 heating rate curves. All these 8 heating rate curves have endothermic melting peaks, and their peak values, corresponding starting points (i.e., melting points), and corresponding enthalpy values ΔH are listed as follows: The differential scanning calorimetry test is carried out using a differential scanning calorimeter Discovery DSC 250 (TA Instruments, US).

7. The crystalline form 1 of the compound according to claim 1 or 2, Stored at 2 - 8 °C, sealed, and protected from light, or at 25 °C ± 2 °C, 60% ± 5% RH, and protected from light for up to 36 months without crystal form transformation, and the purity and water content do not change substantially; or The crystal morphology observed by a polarized light microscope is rod-shaped, and the specific morphology is shown in Figure 15; or By thermogravimetric analysis, weight loss is observed in the range of 120 - 150 °C, and the weight loss does not exceed 0.8%.

8. The crystalline form 1 of the compound according to claim 1, which is a crystal form without water; or By dynamic moisture adsorption and desorption analysis, the desorption weight loss is 0.13% under the condition of 0 - 90% RH.

9. The crystalline form 1 of the compound according to any one of claims 1-8 shows an ion peak with a mass-to-charge ratio of m / z 556.1063 [M+H] in the high-resolution mass spectrum, and the compound I is confirmed by any one of the following spectroscopic detections: + ​ Spectral detection I, infrared spectral detection, using the KBr tablet method to collect infrared absorption spectra in the wavenumber range of 4000 - 400 cm -1 The infrared absorption spectra have the characteristic absorption wavenumbers, vibration types, and structural fragment attributions shown in the following table: Spectrum Detection II, 1H NMR spectrum, detected after dissolving in DMSO-d6, with the following chemical shifts, peak shapes and number of protons: Spectrum Detection III, Nuclear Magnetic Resonance Carbon Spectrum, detected after dissolving in DMSO-d6, with chemical shifts and carbon types as shown in the following table:

10. The preparation method of the crystalline form 1 of the compound according to any one of claims 1 - 8, which is prepared by one of the following methods: Method 1, suspension stirring method: Under the condition of 20 - 55 °C, stir the suspension of compound I product and the first solvent or mixed solvent, filter and collect the filter cake, and dry to obtain the crystalline form 1 of the compound. Method 2, anti-solvent precipitation method: Add a poor solvent to a good solvent solution of Compound I product until solid precipitation occurs, stir the suspension, filter to collect the filter cake and dry it to obtain Compound Crystal Form 1; Method 3, cooling crystallization method: Cool the good solvent solution of Compound I product, filter to collect the filter cake after solid precipitation and dry it to obtain Compound Crystal Form 1; Method 4, evaporation crystallization method: Evaporate the solvent of the good solvent solution of Compound I product, filter to collect the filter cake after solid precipitation and dry it to obtain Compound Crystal Form 1; Among them, In Method 1, the first solvent or mixed solvent is selected from a single solvent or mixed solvent of methanol, ethanol, methyl tert-butyl ether, isopropanol, 1,2-propanediol, glycerol, n-heptane, water; In Methods 2, 3, and 4, the good solvents are each independently selected from a single solvent or mixed solvent of methyl ethyl ketone, acetone, N,N-dimethylformamide, N,N-dimethylacetamide, ethyl acetate, dichloromethane, tetrahydrofuran, acetonitrile, dimethyl sulfoxide, isopropyl acetate, toluene; In Method 2, the poor solvent is selected from a single solvent or mixed solvent of water, isopropanol, n-heptane.

11. According to the preparation method described in Claim 10, In Method 1, The mass-volume ratio of Compound I product to the first solvent or mixed solvent is not greater than 60 mg: 1 mL, The mixed solvent is selected from methanol-n-heptane, methanol-water, methyl tert-butyl ether-n-heptane, ethanol-n-heptane, ethanol-water; In Method 2, The volume ratio of the poor solvent to the volume of the good solvent solution of Compound I product ranges from 0.25 to 5.00, The combination of the good solvent and the poor solvent is selected from acetone + water, acetonitrile + water, dimethyl sulfoxide + water, methyl ethyl ketone + water, acetone + isopropanol, tetrahydrofuran + isopropanol, acetone + n-heptane, ethyl acetate + n-heptane, tetrahydrofuran + n-heptane; In Method 3, The concentration of the good solvent solution of Compound I product at a higher temperature is greater than or equal to 120 mg / mL, the higher temperature is not less than 50 °C and not greater than the solvent boiling point, and the temperature after cooling, i.e., the lower temperature range, is 0 - 25 °C; In Method 4, The concentration range of the good solvent solution of Compound I product is 30 - 60 mg / mL, and the good solvents are selected from single solvents of methyl ethyl ketone, acetone, ethyl acetate, isopropyl acetate, dichloromethane, tetrahydrofuran, acetonitrile, toluene and mixed solvents of methyl ethyl ketone-acetonitrile, acetone-dichloromethane, ethyl acetate-toluene, isopropyl acetate-acetonitrile, dichloromethane-acetonitrile.

12. The preparation method according to claim 10, wherein, Compound I product is detected by HPLC, and its purity is greater than or equal to 95%, The detection conditions are a reverse chromatographic column, mobile phase A is an aqueous solution added with ammonium acetate, mobile phase B is acetonitrile, and mobile phase A and mobile phase B are gradient eluted; detected by an ultraviolet detector, the detection wavelength is 254 nm, and Compound I product is dissolved in acetonitrile for injection; the purity is calculated by the area normalization method.

13. A pharmaceutical composition containing Compound Crystal Form 1 described in any one of Claims 1 - 8.

14. According to the pharmaceutical composition described in Claim 13, it is a liquid injection for injection, and this injection contains N,N-dimethylacetamide and / or ethanol.

15. A purification method of Compound I or Crystal Form 1 of the compound according to any one of Claims 1-8, which comprises putting Compound I or Crystal Form 1 of the compound into methanol for cooling crystallization; Or suspend and stir Compound I or Compound Crystal Form 1 in ethanol; Alternatively, compound I or crystalline form 1 of the compound is added to methyl tert-butyl ether and stirred while suspended.

16. The purification method according to claim 15, wherein cooling crystallization is carried out by cooling from 50 °C; compound I or crystalline form 1 of the compound is added to ethanol and stirred while suspended at room temperature, i.e., 25 °C ± 2 °C; compound I or crystalline form 1 of the compound is added to methyl tert-butyl ether and stirred while suspended at 50 °C.

17. A storage method for compound I or crystalline form 1 of the compound according to any one of claims 1-8: it is placed under sealed and light-proof conditions with a temperature not higher than 25 °C ± 2 °C and a humidity not higher than 60% ± 5% RH; or it is placed under sealed and light-proof conditions with a temperature not higher than 2-8 °C.

18. A storage method for compound I or crystalline form 1 of the compound according to any one of claims 1-8: it is placed under sealed and light-proof conditions with a temperature not higher than 25 °C ± 2 °C and a humidity not higher than 60% ± 5% RH; or it is placed under sealed and light-proof conditions with a temperature not higher than 2-8 °C; Under the above storage conditions, the shelf life, storage period, or validity period of the compound I or crystalline form 1 of the compound is at least 36 months.

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