Crystal form b of 6-((5,6-diphenyl-1,2,4-triazin-3-yl)(isopropyl)amino)-n-(methylsulfonyl)hexanamide, and use thereof and preparation method therefor

Abstract

Disclosed in the present application are a crystal form B of compound I (6-((5,6-diphenyl-1,2,4-triazin-3-yl)(isopropyl)amino)-N-(methylsulfonyl)hexanamide) and a pharmaceutical composition thereof, the use thereof and a preparation method therefor. The crystal form B of compound I in the present application at least has diffraction peaks at the following diffraction angles: 3.36±0.2°, 10.07±0.2°, 13.90±0.2°, 16.51±0.2°, 20.20±0.2° and 21.10±0.2° 2θ in the X-ray powder diffraction pattern thereof. The crystal form B of compound I in the present application has diffraction peaks in the X-ray powder diffraction pattern thereof at least at the following diffraction angles: 3.36±0.2°, 10.07±0.2°, 13.90±0.2°, 16.51±0.2°, 20.20±0.2° and 21.10±0.2° 2θ.

Description

6-((5,6-diphenyl-1,2,4-triazin-3-yl)(isopropyl)amino)-N-(methylsulfonyl)hexanamide crystal form B and its use and preparation method

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0002] This application claims priority to Chinese patent application CN202410169645.1, filed on February 6, 2024. This application claims priority to Chinese patent application CN2024108986421, filed on July 5, 2024. This application cites the full text of the above-mentioned Chinese patent application. Technical Field

[0003] The present application belongs to the field of pharmacy, and specifically relates to a crystalline form of the compound 6-((5,6-diphenyl-1,2,4-triazin-3-yl)(isopropyl)amino)-N-(methylsulfonyl)hexanamide, its pharmaceutical composition, use and preparation method. Background Art

[0004] Pulmonary arterial hypertension (PAH) is a rare, incurable pulmonary vascular disease that can gradually lead to right heart failure and ultimately death. PAH is characterized by pulmonary microvascular remodeling, which leads to a progressive increase in pulmonary artery resistance (PVR), which in turn causes right heart failure. This makes PAH a progressive and fatal disease. 75% of PAH patients die within 5 years of diagnosis, with an average survival of 1.9 years after symptom onset. PAH is therefore often referred to as a "malignant tumor of the cardiovascular and pulmonary vascular system."

[0005] Currently, treatment options for PAH worldwide include conventional therapy and targeted therapies. Conventional therapy often only improves symptoms but cannot effectively halt disease progression. Regarding targeted therapy, there are three pathways for PAH: the nitric oxide pathway, the endothelin pathway, and the prostacyclin (PGI2) pathway. PGI2 is an important endothelial relaxing factor that stimulates cyclic adenosine monophosphate (cAMP) production, causing pulmonary vascular smooth muscle relaxation and inhibiting smooth muscle growth. PGI2 deficiency can cause pulmonary hypertension, making PGI2 drugs the most active treatment for PAH. PGI2 drugs include PGI2 analogs and PGI2 receptor agonists. PGI2 analogs, which share a native PGI2 backbone, are rapidly metabolized and have a very short biological half-life. They require frequent dosing or intravenous infusion, leading to poor patient compliance. Furthermore, PGI2 analogs have poor target selectivity, making it difficult to separate their therapeutic effects from other effects and prone to adverse reactions.

[0006] The compound of formula I, also known as Compound I, has the chemical name 6-((5,6-diphenyl-1,2,4-triazin-3-yl)(isopropyl)amino)-N-(methylsulfonyl)hexanamide, and its structural formula is shown in Formula I. It is a PGI2 receptor agonist with a novel structure and good drugability. Compound I has strong target selectivity, and its agonist activity at the prostacyclin IP receptor is more than 1,000 times that of the other seven prostacyclin receptor targets. It mainly activates the IP receptor to promote the production of cAMP in pulmonary artery smooth muscle cells, thereby inhibiting abnormal contraction of the pulmonary artery, inhibiting the proliferation of pulmonary artery smooth muscle cells, and reducing pulmonary artery pressure, thereby achieving the treatment of pulmonary hypertension. Compared with similar drugs already on the market, it has higher efficacy and safety.

[0007] At present, the crystal form of Compound I has not been disclosed. This application focuses on the crystal form research of Compound I during the drug development process, and provides a drug crystal form with good stability, as well as its pharmaceutical composition, use and preparation method. Technical issues

[0008] After extensive research, it was discovered that Compound I exists in various crystalline forms. Extensive research has been conducted on the crystalline forms of Compound I to identify and prepare crystalline forms that meet pharmaceutical requirements. Based on these studies, this application provides Compound I Form B, which is non-hygroscopic and exhibits excellent storage stability, making it suitable for formulation development. This application also provides pharmaceutical compositions and uses of Compound I Form B, as well as a method for preparing Compound I Form B, which operates under mild process conditions and is suitable for large-scale production. Technical Solutions

[0009] To achieve the purpose of this application, the following technical solutions are adopted:

[0010] One object of the present application is to provide a crystalline form B of compound I, whose X-ray powder diffraction pattern expressed in 2θ angles has diffraction peaks at 3.36±0.2°, 10.07±0.2°, 13.90±0.2°, 16.51±0.2°, 20.20±0.2°, and 21.10±0.2°.

[0011] In some embodiments, the crystalline form B of compound I has an X-ray powder diffraction pattern expressed in 2θ angles having characteristic peaks at 3.36±0.2°, 6.72±0.2°, 10.07±0.2°, 13.90±0.2°, 16.51±0.2°, 17.67±0.2°, 19.50±0.2°, 20.20±0.2°, and 21.10±0.2°.

[0012] In some embodiments, the crystalline form B of Compound I has an X-ray powder diffraction pattern expressed in 2θ angles as shown in FIG1 .

[0013] In some embodiments, when Form B of Compound 1 is characterized by TGA / DSC, its TGA graph can confirm that Form B does not contain crystalline water or solvate.

[0014] In some embodiments, when Form B of Compound I is characterized by TGA / DSC, its DSC graph shows that the melting point (extrapolated onset temperature) of Form B is 140.5±2°C.

[0015] In some embodiments, the TGA / DSC graph of Form B of Compound 1 is shown in FIG2 .

[0016] The second object of the present application is to provide a method for preparing Form B of Compound I, which comprises the following steps: dissolving Compound I by heating in a solvent, cooling to 50°C to 60°C, keeping warm until solid precipitates, continuing crystallization, separation, and drying to obtain Form B.

[0017] In some embodiments, the solvent is one or a mixed solvent of alcohols, ethers, esters, alkanes, ketones, acetonitrile, and water.

[0018] In some embodiments, the solvent is one or a mixed solvent of methanol, ethanol, isopropanol, acetonitrile, tetrahydrofuran, ethyl acetate, n-hexane, acetone, and water.

[0019] In certain preferred embodiments, the solvent is one or a mixed solvent of isopropanol, ethanol, water, tetrahydrofuran and n-hexane.

[0020] In certain preferred embodiments, the solvent is isopropyl alcohol.

[0021] In some embodiments, the mass volume ratio (g / mL) of the compound I to the solvent is 1:3.5-15.

[0022] In some embodiments, the heating and dissolving may be performed in a single solvent or a mixed solvent, or may be performed by first heating and dissolving in one solvent and then adding another one or more solvents.

[0023] In some embodiments, the heating condition is heating under reflux.

[0024] In some embodiments, the temperature is lowered to 50°C to 60°C. There are various ways to cool the temperature. The method may be to turn off the heating and cool the temperature slowly naturally, or to add other solvents to cool the temperature slowly under heating or non-heating conditions. As long as the temperature range of the cooling can be guaranteed, various cooling methods can be selected.

[0025] In some embodiments, the temperature is lowered to 50°C to 60°C, and a more preferred temperature range is 55°C to 60°C.

[0026] In some embodiments, the heat preservation is carried out until solids precipitate, and the crystallization method can be static crystallization or crystallization under stirring, such as paddle stirring, suspension stirring, etc. Stirring crystallization is preferred, as stirring can crystallize faster than static crystallization.

[0027] In some embodiments, the heat preservation is carried out until solids precipitate, and the heat preservation temperature fluctuates by 1° C. to 2° C. based on the previous cooling temperature.

[0028] In some embodiments, the crystallization is continued after the solid precipitates. There are many ways to crystallize, and conventional crystallization methods can be used. For example, the crystallization can be carried out by naturally cooling the temperature, and then the temperature can be further reduced to 0°C to 10°C and kept warm for crystallization.

[0029] Furthermore, the natural cooling crystallization is carried out at room temperature.

[0030] Furthermore, the natural cooling crystallization can be performed by standing still or by stirring, such as paddle stirring, suspension stirring, etc. Stirring crystallization is preferred, as stirring can crystallize faster than standing still.

[0031] Furthermore, the natural cooling crystallization has a crystallization time of 1 to 15 hours, preferably 1 to 2 hours.

[0032] Furthermore, the temperature is further lowered to 0°C to 10°C in a cold water bath, an ice bath, or an ice salt bath. Various cooling conditions can be selected as long as the temperature range of the cooling can be ensured.

[0033] Furthermore, the temperature is lowered to 0°C to 10°C and then kept warm for crystallization. The crystallization can be performed by standing still or under stirring, such as paddle stirring, suspension stirring, etc. Stirring crystallization is preferred, as stirring can crystallize faster than standing still.

[0034] Furthermore, the temperature is lowered to 0°C to 10°C and then kept warm for crystallization, and the crystallization time is 1 to 15 hours, preferably 1 to 2 hours.

[0035] In some embodiments, the heat preservation is continued until solid precipitates and then crystallization is continued. The crystallization method can also be to start heating, heat to 55℃~60℃, keep stirring, cool naturally to crystallize, and then continue to cool to 0℃~10℃ and keep warm to crystallize.

[0036] Furthermore, the heat-insulating stirring has a stirring time of 20 min to 3 h, preferably 30 min to 1 h.

[0037] Furthermore, the natural cooling crystallization is to turn off the heating and cool down naturally and slowly in a hot oil bath.

[0038] Furthermore, the natural cooling crystallization can be performed by standing still or by stirring, such as paddle stirring, suspension stirring, etc. Stirring crystallization is preferred, as stirring can crystallize faster than standing still.

[0039] Furthermore, the natural cooling crystallization has a crystallization time of 5 to 24 hours, preferably 5 to 8 hours.

[0040] Furthermore, the temperature is further lowered to 0°C to 10°C in a cold water bath, an ice bath, or an ice salt bath. Various cooling conditions can be selected as long as the temperature range of the cooling can be ensured.

[0041] Furthermore, the heat preservation crystallization can be performed by standing still or by stirring, such as paddle stirring, suspension stirring, etc. Stirring crystallization is preferred, as stirring can crystallize faster than standing still.

[0042] Furthermore, the crystallization time during the heat preservation is 1 to 20 hours, preferably 1 to 2 hours.

[0043] In some embodiments, the crystallization is continued after the heat preservation until solid precipitates, and the crystallization method can also be natural cooling crystallization.

[0044] Furthermore, the natural cooling crystallization can be performed by removing the oil bath and naturally cooling at room temperature, or by naturally cooling in a hot oil bath.

[0045] Furthermore, the natural cooling crystallization can be performed by standing still or by stirring, such as paddle stirring, suspension stirring, etc. Stirring crystallization is preferred, as stirring can crystallize faster than standing still.

[0046] Furthermore, the natural cooling crystallization has a crystallization time of 5 to 24 hours, preferably 10 to 20 hours.

[0047] In some embodiments, seed crystals of Compound I may be further added.

[0048] In some embodiments, the amount of the seed crystal added is 0.1% to 5% by mass of Compound I.

[0049] In some embodiments, after the heating to dissolve, during the cooling to 50°C to 60°C or during the heat preservation after cooling to 50°C to 60°C, seed crystals of Compound I can be added to further accelerate the precipitation of Form B.

[0050] Furthermore, the seed crystals may be various crystalline forms of Compound I, such as seed crystals of Form A, seed crystals of Form B, etc. The Form A of Compound I is described in the patent "6-((5,6-diphenyl-1,2,4-triazin-3-yl)(isopropyl)amino)-N-(methylsulfonyl)hexanamide Form A, its pharmaceutical composition, use and preparation method" filed by the present applicant on the same filing date.

[0051] In some embodiments, the separation can be performed by conventional methods, such as centrifugation or filtration.

[0052] In some embodiments, the drying is a conventional drying method, such as vacuum drying.

[0053] In some embodiments, the drying entails drying to constant weight.

[0054] The third object of the present application is to provide a pharmaceutical composition comprising a therapeutically effective dose of Compound I Form B and a pharmaceutically acceptable carrier and excipient.

[0055] Furthermore, the pharmaceutical composition can be formulated into a variety of dosage forms for easy administration, for example, oral preparations (such as tablets, capsules, solutions or suspensions); injectable preparations (such as injectable solutions or suspensions, or injectable dry powders that can be used immediately after adding a pharmaceutical solvent before injection).

[0056] The fourth object of the present application is to provide a therapeutically effective dose of Compound I Form B or the pharmaceutical composition for use in the preparation of a drug for preventing and / or treating a disease or condition.

[0057] In certain preferred embodiments, the disease or condition is associated with PGI2 receptor agonism.

[0058] In certain preferred embodiments, the disease or condition is selected from: pulmonary hypertension, platelet aggregation-related cardiovascular and cerebrovascular diseases, and diabetic nephropathy.

[0059] The fifth object of the present application is to provide a therapeutically effective dose of Compound I Form B or the use of the pharmaceutical composition in the preparation of PGI2 receptor agonist drugs.

[0060] Definitions and Explanations of Terms

[0061] As used herein, the term "room temperature" or "RT" refers to an ambient temperature of 20 to 25°C (68 to 77°F).

[0062] On the basis of conforming to the common sense in this field, the above-mentioned preferred conditions can be arbitrarily combined to obtain the preferred embodiments of the present application. Beneficial effects

[0063] The positive progress of this application is that the crystalline Form B of Compound I of the present application has low hygroscopicity, a higher melting point, and outstanding thermal stability. Furthermore, it is highly pure and less susceptible to impurities. It exhibits strong drug stability and crystal form stability in forced degradation tests, making it suitable for formulation development. Furthermore, its mild preparation process conditions make it suitable for large-scale production. Therefore, Form B offers significant advantages in terms of drugability and industrial production, and is of great significance for drug development. BRIEF DESCRIPTION OF THE DRAWINGS

[0064] FIG1 shows the XRPD spectrum of Compound 1 Form B prepared in Example 1;

[0065] FIG2 shows the TGA / DSC spectrum of Compound I Form B prepared in Example 1;

[0066] Figure 3 shows the crystal structure of Compound I Form B prepared in Example 1. 1 H-NMR spectrum;

[0067] FIG4 shows the IR spectrum of Compound 1 Form B prepared in Example 1;

[0068] FIG5 shows the XRPD overlay of Compound I Form B prepared in Example 1 at high temperature, high humidity, and light exposure for 30 days and on day 0;

[0069] FIG6 shows the average concentration-time curve of Compound I in plasma of Group 1 animals after intravenous administration in the bioavailability study of Experimental Example 7;

[0070] FIG7 shows the average concentration-time curve of Compound I in plasma after oral administration to the second group of animals in the bioavailability test of Experimental Example 7. Modes for Carrying Out the Invention

[0071] The present application is further described in detail below through specific implementation methods, but this is only intended to help understand the present application so that professionals in the field can implement or use the present application, and does not constitute any limitation to the present application.

[0072] Example 1

[0073] 10 g of the Compound I drug substance was dissolved in 120 mL of isopropanol by heating under reflux. The oil bath was turned off for heating, and the temperature was slowly lowered to 58° C., maintained at 58±1° C. with stirring until solid precipitated. The mixture was then naturally cooled at room temperature, stirred and crystallized for 2 h, and then further cooled to 10° C. in a cold water bath, maintained with stirring and crystallized for 1.5 h. The mixture was filtered and dried in vacuo to constant weight to obtain a sample of Compound I Form B with a yield of 93.5%.

[0074] Example 2

[0075] 10 g of Compound I API was dissolved in 80 mL of anhydrous ethanol under reflux. The oil bath was turned off and the temperature was slowly cooled to 52°C. The temperature was maintained at 52±1°C with stirring until solid precipitated. The sample was then cooled naturally at room temperature with stirring and crystallization for 1.5 hours. The sample was then cooled to 5°C in an ice bath, maintained with stirring and crystallization for 2.0 hours. The sample was filtered and vacuum dried to constant weight to obtain a sample of Compound I Form B with a yield of 85.3%. The X-ray powder diffraction pattern of the sample obtained by this method was consistent with that of the Compound I Form B sample obtained in Example 1.

[0076] Example 3

[0077] 10 g of Compound I API was dissolved in 50 mL of methanol under reflux. The oil bath was turned off and the temperature was slowly lowered to 50°C. 0.1 g of Form B seed crystals were added and the mixture was stirred at 50±1°C until solid precipitated. The mixture was then heated to 58±2°C and stirred for 30 min. The oil bath was turned off and the mixture was slowly lowered in the hot oil bath while stirring for 5 h. The mixture was then cooled to 0°C in an ice-salt bath, stirred for 1 h, and filtered. The mixture was then dried under vacuum to constant weight to obtain a Compound I Form B sample with a yield of 77.4%. The X-ray powder diffraction pattern of the sample obtained by this method was consistent with that of the Compound I Form B sample obtained in Example 1.

[0078] Example 4

[0079] 10 g of Compound I API was dissolved in 50 mL of anhydrous ethanol and 50 mL of isopropanol under reflux. The oil bath was turned off and the temperature was slowly lowered to 53°C. The temperature was maintained at 53±1°C with stirring until solid precipitated. The sample was then allowed to stand for 13 hours to crystallize. The sample was then cooled to 3°C in an ice bath, stirred at this temperature for 1 hour to crystallize, filtered, and vacuum dried to constant weight to obtain a Compound I Form B sample with a yield of 88.7%. The X-ray powder diffraction pattern of the sample obtained by this method was consistent with that of the Compound I Form B sample obtained in Example 1.

[0080] Example 5

[0081] 10 g of the Compound I drug substance was dissolved in 40 mL of ethyl acetate by heating under reflux. The oil bath was turned off, and the temperature was slowly lowered to 50°C. The mixture was then stirred at 50±1°C until solids precipitated. The heat was then turned on, the temperature was raised to 55±2°C, and stirred at this temperature for 1 hour. The heat was turned off, the temperature was slowly lowered in the hot oil bath, and the mixture was allowed to stand for crystallization for 20 hours. The mixture was then cooled to 2°C in an ice bath, allowed to stand for crystallization for 18 hours, filtered, and dried under vacuum to constant weight to obtain a Compound I Form B sample with a yield of 68.8%. The X-ray powder diffraction pattern of the sample obtained by this method was consistent with that of the Compound I Form B sample obtained in Example 1.

[0082] Example 6

[0083] 10 g of Compound I API was dissolved in 90 mL of 95% ethanol under reflux. The oil bath was turned off and the temperature was slowly lowered to 55°C. The temperature was maintained at 55±1°C with stirring until solid precipitated. The temperature was then lowered to room temperature with stirring for 1 hour to allow crystallization. The temperature was then lowered to 5°C in an ice bath, maintained with stirring for 2.0 hours to allow crystallization. The sample was filtered and dried under vacuum to constant weight to obtain a Compound I Form B sample with a yield of 86.2%. The X-ray powder diffraction pattern of the sample obtained by this method was consistent with that of the Compound I Form B sample obtained in Example 1.

[0084] Example 7

[0085] 10 g of Compound I API was dissolved in 60 mL of acetonitrile and 40 mL of water under reflux. The oil bath was turned off and the temperature was slowly lowered to 51°C. The temperature was maintained at 51±1°C with stirring until solid precipitated. The temperature was then lowered to room temperature with stirring for 1.5 hours to allow crystallization. The temperature was then lowered to 3°C in an ice bath, maintained with stirring for 1 hour to allow crystallization. The mixture was filtered and dried under vacuum to constant weight to obtain a Compound I Form B sample with a yield of 76.1%. The X-ray powder diffraction pattern of the sample obtained by this method was consistent with that of the Compound I Form B sample obtained in Example 1.

[0086] Example 8

[0087] 10 g of Compound I API was dissolved in a mixed solvent of 20 mL of acetone and 15 mL of water under reflux. The oil bath was turned off and the temperature was slowly lowered to 55°C. The temperature was maintained at 55±1°C with stirring until solid precipitated. The temperature was then lowered to room temperature with stirring for 1 hour to allow crystallization. The temperature was then lowered to 3°C in an ice bath, allowed to stand for 15 hours to allow crystallization. The sample was filtered and vacuum dried to constant weight to obtain a Compound I Form B sample with a yield of 80.3%. The X-ray powder diffraction pattern of the sample obtained by this method was consistent with that of the Compound I Form B sample obtained in Example 1.

[0088] Example 9

[0089] 10 g of Compound I API was dissolved in 30 mL of tetrahydrofuran under reflux. 120 mL of n-hexane was slowly added, and the temperature was lowered to 60°C. The mixture was then stirred at 60±2°C until solid precipitated. The oil bath was removed, and the mixture was allowed to cool naturally at room temperature. Crystallization was allowed to proceed with stirring for 16 hours, followed by filtration and vacuum drying to constant weight to obtain a Compound I Form B sample with a yield of 89.3%. The X-ray powder diffraction pattern of the sample obtained by this method was consistent with that of the Compound I Form B sample obtained in Example 1.

[0090] Experimental Example 1: Determination of the diffraction angle (2θ) of the X-ray powder diffraction peak

[0091] Regarding the Compound I Form B sample prepared in Example 1, its X-ray powder diffraction (XRPD) spectrum is shown in FIG1 , and the diffraction angle (2θ) values ​​of its X-ray powder diffraction peaks are shown in Table 1.

[0092] In addition, the physical property data of each crystal described in Experimental Example 1 were measured under the following conditions.

[0093] Testing instrument: Japan Rigaku SmartLab SE fully automatic multifunctional X-ray diffractometer.

[0094] Operating conditions: X-ray tube: cathode: copper; tube voltage: 40 kV; tube current: 30 mA; scanning mode: one-dimensional scanning; scanning rate: 10° / min; scanning axis: θ / 2θ; scanning range: 3-35°; step interval: 0.01°.

[0095] Test results: Form B has characteristic peaks at diffraction angles 2θ of 3.36±0.2°, 6.72±0.2°, 10.07±0.2°, 13.90±0.2°, 16.51±0.2°, 17.67±0.2°, 19.50±0.2°, 20.20±0.2°, and 21.10±0.2° as determined by X-ray powder diffraction.

[0096] Table 1 Diffraction angle (2θ) values ​​of X-ray powder diffraction peaks of the Compound I Form B sample prepared in Example 1

[0097] Experimental Example 2: Thermogravimetric-Differential Scanning Calorimetry Analysis

[0098] Regarding the Compound I Form B sample prepared in Example 1, its thermogravimetric-differential scanning calorimetry (TGA / DSC) spectrum is shown in FIG2 .

[0099] Testing instrument model: Synchronous thermal analyzer STA449F3

[0100] Test conditions: Temperature: 25°C, Humidity: 35% RH

[0101] Crucible: DSC / TG pan Al2O3

[0102] Atmosphere: AIR(80 / 20)-- / NITROGEN / 50 / NITROGEN / 20

[0103] Test results: The TGA graph can confirm that Form B does not contain crystalline water or solvate; the DSC graph can confirm that the melting point (extrapolated onset temperature) of Form B is 140.5±2°C.

[0104] Experimental Example 3: Nuclear Magnetic Resonance Spectroscopy ( 1 H-NMR)

[0105] Regarding the compound I crystal form B sample prepared in Example 1, its nuclear magnetic resonance ( 1 The H-NMR) spectrum is shown in Figure 3.

[0106] Measurement conditions: The data were collected on a Bruker 500 MHz NMR spectrometer with CDCl3 as the solvent.

[0107] Test results: 1 H-NMR: δ: 10.181 (br, 1H), 7.493~7.439 (m, 4H), 7.392~7.378 (m, 1H), 7.317~7.261 (m, 5H), 5.072 (m, 1 H), 3.608 (m, 2H), 3.230 (s, 3H), 2.376 (m, 2H), 1.729 (m, 4H), 1.460 (m, 2H), 1.303~1.289 (m, 6H), ppm.

[0108] Experimental Example 4: Infrared Spectroscopy (IR)

[0109] Regarding the Form B sample of Compound I prepared in Example 1, its infrared (IR) spectrum is shown in FIG4 .

[0110] Instrument model: Nicolet.

[0111] Detection method: Take an appropriate amount of this product (about 1-2 mg) and an appropriate amount of ground and dried potassium bromide, place them in an agate mortar, mix the sample and potassium bromide and grind them evenly, take an appropriate amount of the ground mixture and put it into a tablet pressing mold to press the tablet, and measure the infrared spectrum.

[0112] Test results: The infrared spectrum of the sample is at 3105±5cm -1 、2952±5cm -1 、1684±5cm -1 、1532±5cm -1 、1473±5cm -1 、1457±5cm -1 、1441±5cm -1 、1370±5cm -1 、1150±5cm -1 、1137±5cm -1 、697±5cm - 1 、688±5cm -1 There are characteristic peaks.

[0113] Experimental Example 5: Moisture-absorbing properties

[0114] Regarding the sample of Compound I Form B prepared in Example 1, its hygroscopicity was determined with reference to the 2020 edition of the Chinese Pharmacopoeia, Part IV, General Chapter 9103, Guidelines for Hygroscopicity Tests of Drugs. The specific test method is as follows:

[0115] Take a dry stoppered glass weighing bottle (outer diameter 50mm, height 15mm), place it in a suitable constant temperature and humidity desiccator at 25℃±1℃ one day before the test, and accurately weigh the weight (m1).

[0116] Take an appropriate amount of the test sample and spread it evenly in the above-mentioned weighing bottle. The thickness of the test sample is generally about 1mm, and accurately weigh the weight (m2).

[0117] Open the weighing bottle and place it with the bottle cap under the above constant temperature and humidity conditions for 24 hours. Cover the weighing bottle cap and accurately weigh the weight (m3).

[0118] Test results: As shown in Table 2, the moisture-induced weight gain of the three batches of samples was less than 0.2%, indicating that the crystal form B obtained in this application is not hygroscopic.

[0119] Table 2 Summary of hygroscopicity results

[0120] Experimental Example 6: Stability Test

[0121] A stability study was conducted on the Form B sample of Compound I prepared in Example 1. The samples were placed under high temperature (60°C), high humidity (RH90±5%), and light (4500Lx±500Lx) conditions for 30 days, and then the XRPD spectra of the samples were measured. The changes in the crystal form were determined by comparing with the XRPD spectra measured on day 0. At the same time, the purity was determined by high performance liquid chromatography to examine the changes in related substances.

[0122] The stability test results of the Form B sample at 0 day and 30 days of high temperature, high humidity and light irradiation are shown in Table 3, and the XRPD superposition diagram of the Form B sample at the end of 30 days of high temperature, high humidity and light irradiation and on day 0 is shown in Figure 5.

[0123] The results in Figure 5 show that the XRPD pattern of Form B detected under high temperature (60°C), high humidity (RH90±5%), and light (4500Lx±500Lx) conditions for 30 days is compared with the XRPD pattern detected on day 0. The crystal form remains unchanged, indicating that Form B is a stable crystal form.

[0124] The data in Table 3 show that after 30 days of exposure to high temperature, high humidity, and light, the Form B sample showed no significant changes in related substances, indicating that Form B is highly pure and stable. The stability study results indicate that this crystal form is suitable for formulation development.

[0125] Table 3 Results of the stability test of Form B samples under high temperature, high humidity and light for 30 days

[0126] Experimental Example 7: Bioavailability Test

[0127] Regarding the bioavailability test of the Compound I Form B sample prepared in Example 1, the Compound I Form B sample was first prepared into a clear solution and a suspension using conventional methods.

[0128] This study involved 12 Beagle dogs (half male and half female) randomly divided into two groups, each containing three male and three female animals. Group 1 animals were not fasting and received a single intravenous injection of 0.5 mg / kg of Compound I (clear solution). Blood samples were collected 24 hours after administration at the time of administration (pre-dose, 0.083, 0.25, 0.5, 1, 2, 3, 4, 5, 6, 8, 12, and 24 hours). Group 2 animals were fasting and received a single oral gavage of 0.5 mg / kg of Compound I (suspension). Blood samples were collected 24 hours after administration at the time of administration (pre-dose, 0.25, 0.5, 1, 2, 3, 4, 5, 6, 8, 12, and 24 hours). The homogeneity and concentration of the drug formulations were analyzed using a validated HPLC-UV analytical method. The concentration of Compound I in plasma samples was determined using a validated liquid chromatography-tandem mass spectrometry (LC-MS / MS) analytical method. Using blood drug concentration data, Phoenix 7.0 software uses a non-compartmental model to calculate its bioavailability.

[0129] The average concentration-time curves of Compound I in the plasma of Group 1 animals after intravenous administration and the average concentration-time curves of Compound I in the plasma of Group 2 animals after oral administration in the bioavailability study are shown in Figures 6 and 7, respectively.

[0130] The results showed that the bioavailability of Form B was 67.48%, which was average. However, the bioavailability reached between 60% and 90%, which could meet the basic requirements of in vivo absorption and efficacy.

[0131] The Compound I Form B samples prepared in Examples 2 to 9 have comparable properties to the Compound I Form B sample prepared in Example 1, including hygroscopicity, stability, and bioavailability.

[0132] The above description is only a preferred embodiment of the present application and is not intended to limit the present application. Any modifications, equivalent replacements or improvements made within the spirit and principles of the present application should be included in the scope of protection of the present application.

Claims

1. A crystalline form B of a compound of formula I, characterized in that: The free base crystalline form B of the compound of formula I has an X-ray powder diffraction pattern expressed in 2θ angles at 3.36±0.2°, 10.07±0.2°, 13.90±0.2°, 16.51±0.2°, 20.20±0.2°, and 21.10±0.2°.

2. The crystalline form B of the compound of formula I according to claim 1, wherein The free base crystalline form B of the compound of formula I has an X-ray powder diffraction pattern expressed in 2θ angles at 3.36±0.2°, 6.72±0.2°, 10.07±0.2°, 13.90±0.2°, 16.51±0.2°, 17.67±0.2°, 19.50±0.2°, 20.20±0.2°, and 21.10±0.2°.

3. The crystalline form B of the compound of formula I according to claim 2, characterized in that The free base crystalline form B of the compound of formula I has an X-ray powder diffraction pattern expressed in 2θ angles as shown in Figure 1.

4. The crystalline form B of the compound of formula I according to claim 1, wherein The free base form B of the compound of formula I has a melting point of 138° C.-143° C. as measured by differential scanning calorimetry.

5. The crystalline form B of the compound of formula I according to claim 1, characterized in that The free base form B of the compound of formula I has a thermogravimetric-differential scanning calorimetry analysis spectrum as shown in Figure 2.

6. The method for preparing the crystal form B according to any one of claims 1 to 5, characterized in that: include: Compound I is dissolved by heating in a solvent, cooled to 50°C to 60°C, and kept warm until solid precipitates, followed by crystallization, separation, and drying to obtain Form B.

7. The method for preparing Form B according to claim 6, wherein: The solvent is one or a mixed solvent of alcohols, ethers, esters, alkanes, ketones, acetonitrile or water.

8. The method for preparing Form B according to claim 7, wherein: The solvent is one or a mixed solvent of methanol, ethanol, isopropanol, acetonitrile, tetrahydrofuran, ethyl acetate, n-hexane, acetone or water.

9. The method for preparing Form B according to claim 8, wherein The solvent is one or a mixed solvent of isopropyl alcohol, ethanol, water, tetrahydrofuran and n-hexane.

10. The method for preparing Form B according to claim 9, wherein: The solvent is isopropyl alcohol.

11. The method for preparing Form B according to claim 6, wherein: The mass volume ratio (g / mL) of the compound of formula I to the solvent is 1:3.5-15.

12. The method for preparing Form B according to claim 6, wherein: The heating and dissolving is: heating and dissolving in a single solvent or a mixed solvent, or first heating and dissolving in one solvent and then adding another one or more solvents.

13. The method for preparing Form B according to claim 6, wherein: The temperature reduction range of the cooling to 50°C to 60°C is cooling to 55°C to 60°C.

14. The method for preparing Form B according to claim 6, wherein: The step of keeping the temperature until solid precipitates and then continuing the crystallization comprises the following steps: naturally cooling the temperature for crystallization, then continuing to cool the temperature to 0°C to 10°C, and keeping the temperature for crystallization; or starting heating, heating the temperature to 55°C to 60°C, keeping the temperature for stirring, naturally cooling the temperature for crystallization, then continuing to cool the temperature to 0°C to 10°C, and keeping the temperature for crystallization; or naturally cooling the temperature for crystallization.

15. The method for preparing Form B according to claim 6, wherein: During the preparation process, seed crystals of Compound I may be further added.

16. A pharmaceutical composition, characterized in that A pharmaceutically acceptable carrier, an excipient, and a therapeutically effective dose of the compound of formula I, Form B, according to any one of claims 1 to 5.

17. Use of a therapeutically effective dose of the crystalline form B of the compound of formula I according to any one of claims 1 to 5 or the pharmaceutical composition according to claim 16 in the preparation of a medicament for preventing and / or treating pulmonary hypertension, platelet aggregation-related cardiovascular and cerebrovascular diseases, or diabetic nephropathy.

18. Use of a therapeutically effective dose of the crystalline form B of the compound of formula I according to any one of claims 1 to 5 or the pharmaceutical composition according to claim 16 in the preparation of a PGI2 receptor agonist drug.

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