Polymorph of compound of formula i and preparation method therefor

Abstract

The present invention belongs to the technical field of chemical drugs, and provides a polymorph of a compound of formula I, a preparation method therefor, and a pharmaceutical composition containing the polymorph of the compound of formula I. In addition, the present invention also relates to the use of the polymorph of the compound of formula I or the pharmaceutical composition containing the polymorph of the compound of formula I in the preparation of a drug for treating or preventing related diseases caused by an increased level of CYP11B2 activity.

Description

A polymorph of a compound of formula I and its preparation method Technical Field

[0001] This invention belongs to the field of chemical pharmaceutical technology, and provides a polymorph of a compound of formula I and a method for preparing the same, as well as a pharmaceutical composition containing the polymorph of the compound of formula I. In addition, this invention also relates to the use of the polymorph of the compound of formula I or a pharmaceutical composition containing the polymorph of the compound of formula I of this invention in the preparation of a medicament for treating or preventing diseases caused by elevated CYP11B2 activity levels. Background Technology

[0002] Aldosterone is a steroid hormone with mineralocorticoid activity. It is primarily produced in response to angiotensin II, adrenocorticotropic hormone (ACTH), and the zona glomerulosa of the adrenal gland, which increases serum potassium levels. Its main physiological role in the kidneys is to maintain sodium and potassium balance by regulating cation exchange (Na+ reabsorption and K+ secretion) in the distal nephron. However, aldosterone has also been shown to be a pro-inflammatory and pro-fibrotic hormone in the blood vessels, heart, and kidneys. The effects of aldosterone on gene expression are regulated through binding to the mineralocorticoid receptor (MR) and typical nuclear hormone receptor pathways.

[0003] CYP11B2 (aldosterone synthase) is a cytochrome P450 enzyme known as an enzyme that catalyzes a series of reactions from 11-deoxycorticosterone (i.e., the aldosterone precursor) to aldosterone. CYP11B2 is primarily expressed in the zona glomerulosa of the adrenal cortex, and plasma aldosterone levels are regulated by the activity of this enzyme in the adrenal glands. Furthermore, aldosterone expression has been confirmed in sites outside the adrenal glands, such as the cardiovascular system, kidneys, adipose tissue, and brain. The finding that locally produced aldosterone in various organs is associated with organ dysfunction has attracted attention. Reports indicate that CYP11B2 inhibitors can inhibit aldosterone production in studies using enzymes and cultured cells, and have shown inhibitory and therapeutic effects in studies using various experimental animal models. In addition, CYP11B2 inhibitors have been confirmed to reduce plasma and urinary aldosterone levels and have an antihypertensive effect in patients with hypertension and primary aldosteronism. Finding ways to block the biosynthetic pathway of aldosterone is a highly feasible approach to establishing effective treatments for various aldosterone-related diseases.

[0004] PCT / CN2024 / 142727 discloses an aldosterone synthase inhibitor, its preparation method, and its uses, specifically disclosing compound (R)-N-(4-(2-cyanoquinoline-6-yl)-5,6,7,8-tetrahydroisoquinoline-8-yl)propionamide (i.e., compound I), whose structural formula is: However, the microstructure of its medicinal salt was not disclosed in the patent application.

[0005] It is well known in this field that drug polymorphism is a common phenomenon in drug development and an important factor affecting drug quality. Different polymorphs of the same drug may differ significantly in appearance, solubility, melting point, dissolution rate, and bioavailability, and may also have different impacts on drug stability, bioavailability, and efficacy. Therefore, the issue of drug polymorphism should be comprehensively considered in drug development. Studying the polymorphism of compounds of Formula I is also of great significance for developing drugs suitable for industrial production and with good biological activity. Summary of the Invention

[0006] In view of the problems existing in the prior art, the present invention aims to provide a polymorph of a compound of formula I and a method for preparing the same, so as to solve the problems existing in the prior art.

[0007] This invention is achieved through the following technical solution:

[0008] This invention provides a crystal form A of a compound of formula I, characterized in that, in the X-ray diffraction pattern, the crystal form A of the compound of formula I exhibits characteristic peaks at 13.45°, 13.62°, 23.21°, and 23.95° (represented by a 2θ angle), with an error of ±0.2°; preferably, the crystal form A of the compound of formula I exhibits characteristic peaks at 13.45°, 13.62°, 14.04°, 15.06°, 18.11°, 18.92°, 23.06°, 23.21°, 23.95°, and 25.06° (represented by a 2θ angle), with an error of ±0.2°; more preferably, the crystal form A of the compound of formula I exhibits characteristic peaks at 13.45°, 13.62°, 14.04°, 15.06°, 18.11°, 18.92°, 23.06°, 23.21°, 23.95°, and 25.06° (represented by a 2θ angle), in the X-ray diffraction pattern. Characteristic peaks are observed at 18.92°, 18.92°, 23.06°, 23.21°, 23.95°, 25.06°, and 25.76°, with an error of ±0.2°; more preferably, the A crystal form of the compound of formula I, represented by a 2θ angle in the X-ray diffraction pattern, exhibits characteristic peaks at 9.00°, 9.14°, 13.45°, 13.62°, 14.04°, 15.06°, 18.11°, 18.45°, 18.92°, 19.82°, 21.85°, 22.34°, 23.06°, 23.21°, 23.95°, 25.06°, and 25.76°, with an error of ±0.2°; most preferably, the A crystal form of the compound of formula I has an XRPD spectrum substantially as shown in Figure 1; the structure of the compound of formula I is as follows:

[0009] Furthermore, as a preferred embodiment of the present invention, the DSC spectrum of the A crystal form of the compound of formula I has an endothermic peak at 224.3℃±5℃; preferably, the A crystal form of the compound of formula I has a DSC spectrum substantially as shown in FIG2.

[0010] Furthermore, the present invention also provides a B-crystal form of the compound of formula I, characterized in that the B-crystal form of the compound of formula I exhibits characteristic peaks at 11.19° and 18.76° in the X-ray diffraction pattern, represented by a 2θ angle, with an error of ±0.2°; preferably, the B-crystal form of the compound of formula I exhibits characteristic peaks at 9.27°, 11.19°, 18.76°, 23.43°, and 24.79° in the X-ray diffraction pattern, represented by a 2θ angle, with an error of ±0.2°; more preferably, the B-crystal form of the compound of formula I exhibits characteristic peaks at 9.27°, 11.19°, 18.76°, 20.11°, 21.31°, 22.27°, and 24.79° in the X-ray diffraction pattern, represented by a 2θ angle. Characteristic peaks are observed at 3.00°, 23.43°, 24.79°, 25.25°, and 25.59°, with an error of ±0.2°. More preferably, the B crystal form of the compound of formula I exhibits characteristic peaks at 9.27°, 11.19°, 17.21°, 18.76°, 20.11°, 20.45°, 21.31°, 22.27°, 23.00°, 23.43°, 24.79°, 25.25°, and 25.59° in its X-ray diffraction pattern (represented by a 2θ angle), with an error of ±0.2°. Most preferably, the B crystal form of the compound of formula I has an XRPD spectrum substantially as shown in Figure 3. The structure of the compound of formula I is as follows:

[0011] Further, as a preferred embodiment of the present invention, the B crystal form of the compound of formula I contains 2.4 wt% to 5.0 wt% water; preferably, the B crystal form of the compound of formula I contains 2.46 wt% to 4.9 wt% water; more preferably, the B crystal form of the compound of formula I contains 2.46 wt%, 2.5 wt%, 3.65 wt%, 3.7 wt%, 4.81 wt%, or 4.9 wt% water; even more preferably, the B crystal form of the compound of formula I contains 2.5 wt%, 3.7 wt%, or 4.9 wt% water.

[0012] Furthermore, as a preferred embodiment of the present invention, the DSC spectrum of the B crystal form of the compound of formula I has an endothermic peak at 225.0±5℃, and further, it has endothermic peaks at 67.5±5℃, 101.0±5℃, 129.7±5℃ and 225.0±5℃; preferably, the B crystal form of the compound of formula I has a DSC spectrum basically as shown in Figure 8.

[0013] Furthermore, the present invention also provides a C-crystal form of the compound of formula I, characterized in that the C-crystal form of the compound of formula I exhibits characteristic peaks at 12.05°, 13.56°, 15.18°, and 17.74° in the X-ray diffraction pattern, represented by a 2θ angle, with an error of ±0.2°; preferably, the C-crystal form of the compound of formula I exhibits characteristic peaks at 9.08°, 12.05°, 13.56°, 15.18°, and 17.74° in the X-ray diffraction pattern, represented by a 2θ angle, with an error of ±0.2°; more preferably, the C-crystal form of the compound of formula I exhibits characteristic peaks at 8.27°, 9.08°, 11.13°, 12.05°, 12.38°, 13.56°, and 15.1° in the X-ray diffraction pattern, represented by a 2θ angle. Characteristic peaks are observed at 8°, 17.74°, 18.09°, and 21.53°, with an error of ±0.2°; more preferably, the C crystal form of the compound of formula I, represented by a 2θ angle in the X-ray diffraction pattern, exhibits characteristic peaks at 4.05°, 8.27°, 9.08°, 11.13°, 12.05°, 12.38°, 13.56°, 15.18°, 17.74°, 18.09°, 18.64°, 20.46°, 21.53°, 26.85°, 27.76°, 28.80°, and 29.30°, with an error of ±0.2°; most preferably, the C crystal form of the compound of formula I has an XRPD spectrum substantially as shown in Figure 4; the structure of the compound of formula I is as follows:

[0014] Furthermore, the present invention also provides a D-crystal form of the compound of formula I, characterized in that the D-crystal form of the compound of formula I exhibits characteristic peaks at 8.52°, 13.13°, 16.94°, 19.91°, and 24.90° in the X-ray diffraction pattern, with an error of ±0.2°; preferably, the D-crystal form of the compound of formula I exhibits characteristic peaks at 8.52°, 13.13°, 16.94°, 17.29°, 19.91°, 20.86°, 22.04°, and 24.90° in the X-ray diffraction pattern, with an error of ±0.2°; more preferably, the D-crystal form of the compound of formula I exhibits characteristic peaks at 8.52°, 13.13°, 16.94°, 17.29°, 19.91°, 20.86°, 22.04°, and 24.90° in the X-ray diffraction pattern, with an error of ±0.2°; more preferably, the D-crystal form of the compound of formula I exhibits characteristic peaks at 8.52°, 13.13°, 16.94°, 17.29°, 19.91°, 20.86°, 22.04°, and 24.90° in the X-ray diffraction pattern, with an error of ±0.2°. Characteristic peaks are observed at 19.91°, 20.44°, 20.86°, 22.04°, 22.39°, and 24.90°, with an error of ±0.2°; more preferably, the D crystal form of the compound of formula I exhibits characteristic peaks at 8.52°, 12.65°, 13.13°, 16.94°, 17.29°, 18.58°, 19.91°, 20.44°, 20.86°, 22.04°, 22.39°, 24.90°, 25.70°, 25.85°, and 26.20° in the X-ray diffraction pattern, represented by a 2θ angle, with an error of ±0.2°; most preferably, the D crystal form of the compound of formula I has an XRPD spectrum substantially as shown in Figure 5; the structure of the compound of formula I is as follows:

[0015] Furthermore, the present invention also provides an E-crystal form of a compound of formula I, characterized in that the E-crystal form of the compound of formula I has characteristic peaks at 18.80° and 24.82° in the X-ray diffraction pattern, represented by 2θ angles, with an error of ±0.2°; preferably, the E-crystal form of the compound of formula I has characteristic peaks at 9.28°, 9.46°, 18.80° and 24.82° in the X-ray diffraction pattern, represented by 2θ angles, with an error of ±0.2°; more preferably, the E-crystal form of the compound of formula I has characteristic peaks at 8.63°, 9.28°, 9.46° and 18.80° in the X-ray diffraction pattern, represented by 2θ angles, with an error of ±0.2°; Characteristic peaks are observed at 1.34°, 14.14°, 18.80°, 24.82°, and 29.31°, with an error of ±0.2°; more preferably, the E crystal form of the compound of formula I, represented by a 2θ angle in the X-ray diffraction pattern, exhibits characteristic peaks at 8.63°, 9.28°, 9.46°, 11.34°, 14.14°, 17.38°, 18.80°, 24.82°, and 29.31°, with an error of ±0.2°; most preferably, the E crystal form of the compound of formula I has an XRPD spectrum substantially as shown in Figure 6; the structure of the compound of formula I is as follows:

[0016] Furthermore, the present invention also provides an F-crystal form of the compound of formula I, characterized in that the F-crystal form of the compound of formula I has characteristic peaks at 8.27°, 13.78°, and 23.60° in the X-ray diffraction pattern, represented by a 2θ angle, with an error of ±0.2°; preferably, the F-crystal form of the compound of formula I has characteristic peaks at 3.57°, 8.27°, 13.78°, 23.60°, and 25.28° in the X-ray diffraction pattern, represented by a 2θ angle, with an error of ±0.2°; more preferably, the F-crystal form of the compound of formula I has characteristic peaks at 3.57°, 8.27°, 11.18°, 13.78°, 16.65°, 18.42°, 23.60°, 25.28°, 26.10°, and 25.28° in the X-ray diffraction pattern, represented by a 2θ angle. Characteristic peaks are observed at 6.64°, 27.14°, and 28.16°, with an error of ±0.2°; more preferably, the F crystal form of the compound of formula I, represented by a 2θ angle in the X-ray diffraction pattern, exhibits characteristic peaks at 3.57°, 6.98°, 8.27°, 9.17°, 10.42°, 11.18°, 12.08°, 13.78°, 16.65°, 17.05°, 18.42°, 22.39°, 23.60°, 24.23°, 25.28°, 26.10°, 26.64°, 27.14°, and 28.16°, with an error of ±0.2°; most preferably, the F crystal form of the compound of formula I has an XRPD spectrum substantially as shown in Figure 7; the structure of the compound of formula I is as follows:

[0017] Furthermore, the present invention also provides a G-crystal form of the compound of formula I, characterized in that the G-crystal form of the compound of formula I has characteristic peaks at 12.12°, 13.62°, 20.21°, 20.51°, 22.89° and 25.60° in the X-ray diffraction pattern, with an error of ±0.2°; preferably, the G-crystal form of the compound of formula I is represented by 2θ angles at 12.12°, 13.62°, 15.60° in the X-ray diffraction pattern. Characteristic peaks are observed at 25°, 17.72°, 20.21°, 20.51°, 21.19°, 21.37°, 22.89°, and 25.60°, with an error of ±0.2°; more preferably, the G crystal form of the compound of formula I, represented by a 2θ angle in the X-ray diffraction pattern, has characteristic peaks at 8.37°, 9.12°, 12.12°, 12.46°, 13.62°, 15.25°, 17.72°, 20.21°, 20.51°, and 25.60°. Characteristic peaks are observed at 21.19°, 21.37°, 22.89°, 23.48°, 25.60°, 26.30°, and 26.80°, with an error of ±0.2°; more preferably, the G crystal form of the compound of formula I, represented by a 2θ angle in the X-ray diffraction pattern, is at 8.37°, 9.12°, 12.12°, 12.46°, 13.62°, 15.25°, 17.72°, 18.10°, 18.56°, and 18... Characteristic peaks are observed at 0.72°, 20.21°, 20.51°, 21.19°, 21.37°, 22.89°, 23.48°, 24.81°, 25.60°, 26.30°, 26.80°, 27.72°, 28.82°, and 29.25°, with an error of ±0.2°; most preferably, the G crystal form of the compound of formula I has an XRPD spectrum substantially as shown in Figure 9; the structure of the compound of formula I is shown below:

[0018] Furthermore, as a preferred embodiment of the present invention, the DSC spectrum of the G crystal form of the compound of formula I has an endothermic peak at 225.1±5℃, and further, it has endothermic peaks at 131.6±5℃ and 225.1±5℃; preferably, the G crystal form of the compound of formula I has a DSC spectrum basically as shown in Figure 11.

[0019] Furthermore, the present invention also provides an H-crystal form of the compound of formula I, characterized in that the H-crystal form of the compound of formula I has characteristic peaks at 10.20° and 19.14° in the X-ray diffraction pattern, represented by 2θ angles, with an error of ±0.2°; preferably, the H-crystal form of the compound of formula I has characteristic peaks at 9.44°, 10.20°, 17.63°, 19.14°, 19.43°, 20.94°, and 24.93° in the X-ray diffraction pattern, represented by 2θ angles, with an error of ±0.2°; more preferably, the H-crystal form of the compound of formula I has characteristic peaks at 9.44°, 10.20°, 15.20°, 17.63°, 19.14°, 19.43°, 20.25°, 20.94°, 23.00°, and 24.93° in the X-ray diffraction pattern, represented by 2θ angles. Characteristic peaks are observed at 93° and 25.40°, with an error of ±0.2°; more preferably, the H crystal form of the compound of formula I, represented by 2θ angles in the X-ray diffraction pattern, exhibits characteristic peaks at 4.79°, 5.22°, 7.72°, 9.44°, 10.20°, 10.53°, 14.11°, 14.59°, 15.20°, 17.63°, 19.14°, 19.43°, 20.25°, 20.94°, 21.79°, 23.00°, 23.93°, 24.42°, 24.57°, 24.93°, 25.40°, and 25.73°, with an error of ±0.2°; most preferably, the H crystal form of the compound of formula I has an XRPD spectrum substantially as shown in Figure 10; the structure of the compound of formula I is shown below:

[0020] Furthermore, as a preferred embodiment of the present invention, the DSC spectrum of the H crystal form of the compound of formula I has an endothermic peak at 225.6±5℃, and further, it has endothermic peaks at 110.4±5℃, 124.8±5℃, 132.5±5℃ and 225.6±5℃; preferably, the H crystal form of the compound of formula I has a DSC spectrum basically as shown in Figure 12.

[0021] Furthermore, the present invention also provides a pharmaceutical composition, characterized in that the pharmaceutical composition comprises the crystal form of the compound of formula I as described in any of the preceding claims and one or more pharmaceutically acceptable excipients and / or carriers.

[0022] Furthermore, the present invention also provides a crystal form of the compound of formula I described in any of the preceding claims, or the use of the pharmaceutical composition of the present invention in the preparation of a medicament for treating or preventing diseases related to elevated CYP11B2 activity levels; preferably, the diseases related to elevated CYP11B2 activity levels are selected from: hypertension, chronic kidney disease, primary aldosteronism, diabetic nephropathy, congestive heart failure, or Cushing's syndrome.

[0023] For clarity, this article defines the general terminology used in the description of compounds.

[0024] Unless otherwise stated, the following terms and phrases used herein are intended to have the following meanings. A particular term or phrase should not be considered uncertain or unclear unless specifically defined, but should be understood in its ordinary sense. When a trade name appears herein, it is intended to refer to the corresponding product or its active ingredient. The term "pharmaceutically acceptable" as used herein refers to compounds, materials, compositions, and / or dosage forms that, within the bounds of reliable medical judgment, are suitable for use in contact with human and animal tissues without undue toxicity, irritation, allergic reactions, or other problems or complications, in proportion to a reasonable benefit / risk ratio.

[0025] The term "medicinal salt" refers to a salt of the compounds of the present invention, prepared by reacting a compound having specific substituents discovered in the present invention with a medicinal acid or base.

[0026] The term "excipient" generally refers to the carrier, diluent, and / or medium required to formulate an effective pharmaceutical composition.

[0027] "Pharmaceutical composition" means a mixture containing one or more of the compounds described herein or their physiologically / pharmacologically acceptable salts or prodrugs, along with other chemical components, such as physiologically / pharmacologically acceptable carriers and excipients. The purpose of a pharmaceutical composition is to facilitate administration to a living organism, thereby promoting the absorption of the active ingredient and the exertion of its biological activity.

[0028] X-ray powder diffraction (XRPD) refers to the experimentally observed diffraction pattern or parameters derived from it, characterized by peak positions (x-axis) and peak intensities (y-axis). Those skilled in the art will understand that experimental errors depend on instrument conditions, sample preparation, and sample purity. In particular, it is known to those skilled in the art that X-ray diffraction patterns typically change with instrument conditions, and appropriate error tolerances for XRPD can be: 2θ ± 0.5°; 2θ ± 0.4°; 2θ ± 0.3°; 2θ ± 0.2°. It is particularly important to note that the relative intensities of the X-ray diffraction pattern can also vary with experimental conditions, so the order of peak intensities cannot be considered the sole or decisive factor. Furthermore, the influence of experimental factors such as sample height can cause an overall shift in peak angles, which is generally permissible. Therefore, those skilled in the art will understand that any crystal form with characteristic peaks identical or similar to those of the patterns of this invention falls within the scope of this invention.

[0029] For pharmaceuticals or pharmacologically active agents, the term "effective amount" or "therapeutic effective amount" refers to a sufficient quantity of a drug or agent that is non-toxic but achieves the desired effect. For the oral dosage forms of this invention, the "effective amount" of one active substance in the composition refers to the quantity required to achieve the desired effect when used in combination with another active substance in the composition. The determination of the effective amount varies from person to person, depending on the recipient's age and general condition, as well as the specific active substance. A suitable effective amount in any given case can be determined by a person skilled in the art through routine testing.

[0030] The terms “active ingredient,” “therapeutic agent,” “active substance,” or “active agent” refer to a chemical entity that can effectively treat a target disorder, disease, or symptom.

[0031] "Optional" or "optionally" means that the event or condition described below may occur but is not required to occur, and the description includes both the scenario in which said event or condition occurs and the scenario in which said event or condition does not occur. Attached Figure Description

[0032] 1) Figure 1 is the X-ray diffraction pattern of crystal form A of the compound of formula I obtained in Example 2;

[0033] 2) Figure 2 is the DSC spectrum of crystal form A of the compound of formula I obtained in Example 2;

[0034] 3) Figure 3 is the X-ray diffraction pattern of the B crystal form of the compound of formula I obtained in Example 3;

[0035] 4) Figure 4 shows the X-ray diffraction pattern of the C-crystal form of the compound of formula I obtained in Example 4;

[0036] 5) Figure 5 shows the X-ray diffraction pattern of the D crystal form of the compound of formula I obtained in Example 5;

[0037] 6) Figure 6 is the X-ray diffraction pattern of the E crystal form of the compound of formula I obtained in Example 6;

[0038] 7) Figure 7 shows the X-ray diffraction pattern of the F-crystal form of the compound of formula I obtained in Example 7;

[0039] 8) Figure 8 is the DSC spectrum of the B crystal form of the compound of formula I obtained in Example 3;

[0040] 9) Figure 9 shows the X-ray diffraction pattern of the G crystal form of the compound of formula I obtained in Example 8;

[0041] 10) Figure 10 is the X-ray diffraction pattern of the H crystal form of the compound of formula I obtained in Example 9;

[0042] 11) Figure 11 is the DSC spectrum of the G crystal form of the compound of formula I obtained in Example 8;

[0043] 12) Figure 12 is the DSC spectrum of the H crystal form of the compound of formula I obtained in Example 9. Detailed Implementation

[0044] The present invention will be further described in detail below with reference to the embodiments, but the content of the invention is not limited to the embodiments.

[0045] The structure of the compound was determined by nuclear magnetic resonance (NMR) or mass spectrometry (MS). NMR shifts (δ) were expressed in 10⁻¹⁰ increments. -6 The unit (ppm) is given. NMR measurements were performed using a Bruker AVANCE-III NMR spectrometer. The solvents used were deuterated dimethyl sulfoxide (DMSO-d6) and deuterated chloroform (CDCl3), with tetramethylsilane (TMS) as the internal standard.

[0046] MS measurements were performed using an ISQ EC mass spectrometer (manufacturer: Thermo, model: ISQ EC).

[0047] High-performance liquid chromatography (HPLC) analysis was performed using a Thermo U3000 HPLC DAD HPLC system and an Agilent 1260 HPLC system.

[0048] The CombiFlash rapid preparation system uses CombiFlash Rf+LUMEN (TELEDYNE ISCO).

[0049] Thin-layer chromatography silica gel plates used are from Yantai Yinlong HSGF. 254 or GF 254 Silica gel plates: The silica gel plates used in thin-layer chromatography (TLC) have a size of 0.17 mm to 0.23 mm, while those used for TLC separation and purification of products have a size of 0.4 mm to 0.5 mm.

[0050] Silica gel column chromatography generally uses 100-200 mesh silica gel from Rushan Shangbang as the carrier.

[0051] Unless otherwise stated, the crystal form of the present invention was detected using the following equipment and conditions: X-ray powder diffraction (XRPD). The XRPD pattern was acquired on an X-ray powder diffractometer manufactured by PANalytacal, and the scanning parameters are shown in the table below:

[0052] X-ray powder diffraction conditions:

[0053] The X-ray powder diffraction (XRPD) pattern shown in this invention was obtained using an Empyrean X-ray powder diffractometer under the following conditions: Cu-Kα radiation, wavelength... The divergence slit is 1 / 4°, the X-ray tube voltage is 45kV, the X-ray tube current is 40mA, the scanning range is 3-40° (2θ), the step size is 0.026°, and the dwell time per step is about 30s.

[0054] Differential scanning calorimetry (DSC) was performed. The DSC images were acquired using a NETZSCH DSC 200F3 differential scanning calorimeter. The test parameters are shown in the table below:

[0055] DSC test conditions:

[0056] Dynamic moisture adsorption test conditions:

[0057] The dynamic moisture adsorption map described in this application was acquired using an Intrinsic dynamic moisture adsorption analyzer from SMS Corporation. The parameters of the dynamic moisture adsorption test method described in this invention are as follows:

[0058] Humidity range 0-95% RH

[0059] Temperature: 25℃

[0060] Protective gas and flow rate: N2, 200 ml / min

[0061] Maximum test time for each humidity level: 6 hours

[0062] Sample collection time interval: 5 seconds.

[0063] Example 1

[0064] (R)-N-(4-(2-cyanoquinolin-6-yl)-5,6,7,8-tetrahydroisoquinolin-8-yl)propionamide

[0065] The specific synthesis route is as follows:

[0066] Step A: Synthesis of ethyl 5-bromo-4-methylnicotinate

[0067] 5-Bromo-4-methylnicotinic acid (50.0 g, 231.45 mmol) and iodoethane (39.7 g, 254.59 mmol) were dissolved in 500 mL of N,N-dimethylformamide, and potassium bicarbonate (46.3 g, 462.90 mmol) was added. The mixture was degassed and protected with nitrogen, and the reaction was stirred at room temperature for 12 hours.

[0068] After the reaction was complete, the mixture was filtered, water was added to the filtrate, and the mixture was extracted with ethyl acetate (300 mL × 3 times). The combined organic phases were washed with saturated brine (500 mL), dried over anhydrous sodium sulfate, and finally concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: ethyl acetate / n-hexane = 1 / 10) to give 54.6 g of ethyl 5-bromo-4-methylnicotinate. [M+H] + =244.05.

[0069] Step B: Synthesis of methyl 4-bromo-8-oxo-5,6,7,8-tetrahydroisoquinoline-7-carboxylate

[0070] At -78°C, LDA (123 mL, 246.06 mmol, 2M) was added dropwise to a tetrahydrofuran (500 mL) solution of ethyl 5-bromo-4-methylnicotinate (54.6 g, 223.69 mmol). The mixture was stirred for 30 minutes, followed by the addition of a tetrahydrofuran (200 mL) solution of methyl acrylate (48.1 g, 559.22 mmol). The mixture was then stirred at -78°C for 2 hours.

[0071] After the reaction was complete, 400 mL of 10% acetic acid aqueous solution was added to the mixture to quench the reaction. The organic solvent was removed by evaporation, and the mixture was extracted with ethyl acetate (300 mL × 3 times). The combined organic phases were washed with saturated brine (500 mL), dried over anhydrous sodium sulfate, and finally concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: ethyl acetate / n-hexane = 1 / 10) to give 31.5 g of methyl 4-bromo-8-oxo-5,6,7,8-tetrahydroisoquinoline-7-carboxylate. [M+H] + =284.06.

[0072] Step C: Synthesis of 4-bromo-6,7-dihydroisoquinoline-8(5H)-one

[0073] 31.5 g (110.87 mmol) of methyl 4-bromo-8-oxo-5,6,7,8-tetrahydroisoquinoline-7-carboxylate was dissolved in 300 mL of hydrochloric acid (6 M), and the mixture was refluxed at 105 °C for 16 hours.

[0074] After the reaction was complete, the solvent was evaporated, 300 mL of water was added, and the pH was adjusted to approximately 9 with 1 N sodium hydroxide aqueous solution. The mixture was extracted with ethyl acetate (200 mL × 3 times), the organic phases were combined, washed with saturated brine (300 mL), dried over anhydrous sodium sulfate, and finally concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: ethyl acetate / n-hexane = 1 / 8) to give 19.6 g of 4-bromo-6,7-dihydroisoquinoline-8(5H)-one. [M+H] + =226.05.

[0075] Step D: Synthesis of (S)-N-(4-bromo-6,7-dihydroisoquinoline-8(5H)-ylidene)-2-methylpropane-2-sulfonamide

[0076] 4-Bromo-6,7-dihydroisoquinoline-8(5H)-one (10.0 g, 44.23 mmol) was dissolved in 200 mL of tetrahydrofuran, and (S)-tert-butylsulfinamide (5.9 g, 48.66 mmol) and tetraisopropyl titanate (37.7 g, 132.70 mmol) were added. The mixture was heated to 65 °C under nitrogen protection and stirred for 24 hours.

[0077] After the reaction was complete, 100 mL of water was added to quench the reaction, and the solid was filtered. The filtrate was concentrated, and 100 mL of water was added to the residue. The residue was extracted with ethyl acetate (100 mL × 3 times), and the organic phases were combined. The residue was washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, and finally concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: ethyl acetate / n-hexane = 1 / 5) to give 13.3 g of (S)-N-(4-bromo-6,7-dihydroisoquinoline-8(5H)-ylidene)-2-methylpropane-2-sulfonamide. [M+H] + =329.12.

[0078] Step E: Synthesis of (S)-N-((R)-4-bromo-5,6,7,8-tetrahydroisoquinoline-8-yl)-2-methylpropane-2-sulfonamide

[0079] Sodium borohydride (2.3 g, 60.59 mmol) was added in portions to a methanol (400 mL) solution of (S)-N-(4-bromo-6,7-dihydroisoquinoline-8(5H)-ylidene)-2-methylpropane-2-sulfonamide (13.3 g, 40.39 mmol), and the mixture was stirred at -42 °C for 1 hour.

[0080] After the reaction was complete, 100 mL of water was added to quench the reaction, the solvent was evaporated, and 100 mL of water was added to the residue. The mixture was extracted with ethyl acetate (100 mL × 3 times), the organic phases were combined, washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, and finally concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 20) to give 11.1 g of (S)-N-((R)-4-bromo-5,6,7,8-tetrahydroisoquinoline-8-yl)-2-methylpropane-2-sulfonamide. [M+H] + =331.06. 1HNMR(400MHz, CDCl3)δ8.58(s,1H),8.57(s,1H),4.59–4.51(m,1H),3.41(d,J=10.0Hz,1H) ,2.83–2.68(m,2H),2.38–2.28(m,1H),2.05–1.95(m,2H),1.94–1.84(m,1H),1.29(s,9H).

[0081] Step F: Synthesis of (R)-4-bromo-5,6,7,8-tetrahydroisoquinoline-8-amine

[0082] In a solution of (S)-N-((R)-4-bromo-5,6,7,8-tetrahydroisoquinoline-8-yl)-2-methylpropane-2-sulfonamide (11.1 g, 9.86 mmol) in dichloromethane (100 mL), 40 mL of hydrogen chloride-dioxane solution (4 M) was added, and the mixture was stirred at room temperature for 5 hours.

[0083] After the reaction was complete, the solvent was evaporated from the mixture, and 100 mL of water was added to the residue. The pH was adjusted to 9 with sodium hydroxide solution (1 M), and the mixture was extracted with ethyl acetate (100 mL × 3 times). The organic phases were combined, washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, and finally concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 10) to give 7.2 g of (R)-4-bromo-5,6,7,8-tetrahydroisoquinoline-8-amine. [M+H] + =227.11.

[0084] Step G: Synthesis of (R)-N-(4-bromo-5,6,7,8-tetrahydroisoquinoline-8-yl)propionamide

[0085] (R)-4-bromo-5,6,7,8-tetrahydroisoquinoline-8-amine (7.2 g, 31.70 mmol) and triethylamine (8.8 mL, 63.41 mmol) were dissolved in dichloromethane (100 mL), and propionyl chloride (3.1 mL, 34.87 mmol) was added dropwise at 0 °C. The mixture was stirred at room temperature for 5 minutes.

[0086] After the reaction was complete, water was added to the mixture, and the mixture was extracted with dichloromethane (100 mL × 3 times). The organic phases were combined, washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, and finally concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 20) to give 8.5 g of (R)-N-(4-bromo-5,6,7,8-tetrahydroisoquinoline-8-yl)propionamide. [M+H] + =283.12.

[0087] Step H: Synthesis of (R)-N-(4-(2-cyanoquinoline-6-yl)-5,6,7,8-tetrahydroisoquinoline-8-yl)propionamide

[0088] (R)-4-bromo-5,6,7,8-tetrahydroisoquinoline-8-amine (100 mg, 0.35 mmol) and pinacol ester of 2-cyanoquinoline-6-borate (119 mg, 0.42 mmol) were dissolved in a mixed solvent of 5.0 mL dioxane and 1.0 mL water. Sodium carbonate (76 g, 0.71 mmol) and tetrakis(triphenylphosphine)palladium (8 mg, 0.0071 mmol) were added. The mixture was reacted under nitrogen protection at 85 °C for 6 hours.

[0089] After the reaction was completed, the resulting suspension was filtered, the filter cake was washed with dichloromethane, the filtrate was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (eluent: methanol / dichloromethane = 1 / 20) to give 109 mg (R)-N-(4-(2-cyanoquinoline-6-yl)-5,6,7,8-tetrahydroisoquinoline-8-yl)propionamide.

[0090] [M+H] + =357.00. NMR data: 1 HNMR (400MHz, DMSO-d6) δ8.74(d,J=8.5Hz,1H),8.45(s,1H),8.39(s,1H),8.34(d,J=8.4Hz,1H),8.25(d,J=8.7Hz,1H),8.18(d,J=2.0Hz,1H),8.14( d,J=8.4Hz,1H),7.99(dd,J=8.7,2.0Hz,1H),5.15(q,J=6.5Hz,1H),2.71– 2.63(m,2H),2.26–2.12(m,2H),1.99–1.66(m,4H),1.09(t,J=7.6Hz,3H).

[0091] Example 2

[0092] Preparation of crystal form A of compound I:

[0093] a) The product (1 g) from step H of Example 1 was added to a reaction flask, followed by acetone (30 mL). The mixture was heated to 50–60 °C and stirred until the solid dissolved completely. The mixture was then concentrated to dryness under vacuum. After concentration, ethyl acetate (10 mL) was added to the reaction flask at room temperature, and the mixture was stirred for 60 min. Then, methyl tert-butyl ether (10 mL) was added, and the mixture was cooled to 0–10 °C and stirred for approximately 1 h. The mixture was filtered, and the filter cake was washed with methyl tert-butyl ether. The resulting solid was dried under vacuum at 80–90 °C for approximately 12 h to obtain the A crystal form of compound I.

[0094] The X-ray diffraction pattern of the A-crystal form of the obtained Formula I compound is shown in Figure 1. The specific characteristic peaks are shown in Table 1 below, with an error of ±0.2°. The DSC spectrum of the A-crystal form of the Formula I compound has an endothermic peak at 224.3℃±5℃; the DSC spectrum of the A-crystal form of the Formula I compound is shown in Figure 2.

[0095] Table 1. X-ray diffraction data of crystal form A of compound I.

[0096] Preparation of crystal form A of compound I:

[0097] b) Add 50 mg of the product from step H of Example 1 to 1 ml of ethanol and stir at room temperature until dissolved. Take 0.5 ml of the solution, add 2 ml of isopropyl ether, stir at room temperature for 3 days, filter, and dry to obtain a solid.

[0098] Example 3

[0099] Preparation of crystal form B of compound I:

[0100] a) Weigh 1g of the A crystal form of the compound of formula I prepared in Example 2, add 5mL of ethanol and 5mL of water, stir at room temperature for 1 day, filter, and dry the solid under vacuum at 35°C to obtain the B crystal form, with a yield of 99%.

[0101] The X-ray diffraction pattern of the B crystal form of the obtained compound of formula I is shown in Figure 3. The specific characteristic peaks are shown in Table 2 below, with an error of ±0.2°.

[0102] The DSC spectrum of the B crystal form of the compound of formula I has endothermic peaks at 67.5±5℃, 101.0±5℃, 129.7±5℃ and 225.0±5℃; the DSC spectrum of the B crystal form of the compound of formula I is shown in Figure 8.

[0103] Table 2. X-ray diffraction data of crystal form B of compound I.

[0104] Preparation of crystal form B of compound I:

[0105] b) Weigh 2g of the A crystal form of the compound of formula I prepared in Example 2, add 10mL of water, stir at room temperature for 1 day, filter, dry the solid under vacuum at 40°C for 1 day, and then dry at 50°C for 1 day to obtain the B crystal form.

[0106] Example 4

[0107] Preparation of the C-crystal form of compound I:

[0108] Weigh 1g of the A crystal form of the compound of formula I prepared in Example 2, add 10mL of butyl formate, stir at room temperature for 1 day, filter, and dry the solid under vacuum at 35°C to obtain the C crystal form, with a yield of 81%.

[0109] The X-ray diffraction pattern of the C crystal form of the obtained compound of formula I is shown in Figure 4. The specific characteristic peaks are shown in Table 3 below, with an error of ±0.2°.

[0110] Table 3. X-ray diffraction data of the C-crystal form of compound I.

[0111] Example 5

[0112] Preparation of the D-crystal form of compound I:

[0113] Weigh 50 mg of the A crystal form of compound I prepared in Example 2, add 1 mL of acetic acid, stir at room temperature for 1 day, then add 4 mL of isopropyl ether, continue stirring for 1 day, filter, and then dry the solid under vacuum to obtain the D crystal form.

[0114] The X-ray diffraction pattern of the D crystal form of the obtained compound of formula I is shown in Figure 5. The specific characteristic peaks are shown in Table 4 below, with an error of ±0.2°.

[0115] Table 4. X-ray diffraction data of the D crystal form of compound I.

[0116] Example 6

[0117] Preparation of the E crystal form of compound I:

[0118] Weigh 1g of the A crystal form of the compound of formula I prepared in Example 2, add 10mL of acetic acid, stir at room temperature for 30 minutes, add 5mL of isopropyl ether, stir for 10 minutes, then add 25mL of isopropyl ether, and continue stirring for 1 day. Filter, and dry the solid under vacuum at 35°C to obtain the E crystal form, with a yield of 88%.

[0119] The X-ray diffraction pattern of the E crystal form of the obtained compound of formula I is shown in Figure 6. The specific characteristic peaks are shown in Table 5 below, with an error of ±0.2°.

[0120] Table 5. X-ray diffraction data of the E crystal form of compound I.

[0121] Example 7

[0122] Preparation of the F-crystal form of compound I:

[0123] Weigh 50 mg of the A crystal form of compound I prepared in Example 2, add 1 mL of isopropanol, stir at room temperature for 1 day, then filter using a 0.22 μm filter membrane. The filtrate slowly evaporates at room temperature, and after 35 days, a white solid is obtained. The solid is dried under vacuum at 50 °C to obtain the F crystal form.

[0124] The X-ray diffraction pattern of the F crystal form of the obtained compound of formula I is shown in Figure 7. The specific characteristic peaks are shown in Table 6 below, with an error of ±0.2°.

[0125] Table 6. X-ray diffraction data of the F crystal form of compound I.

[0126] Example 8

[0127] Preparation of the G crystal form of compound I:

[0128] Weigh 2g of the A crystal form of compound I prepared in Example 2, add 10mL of butyl formate, stir at room temperature for 10 minutes, then add 5mL of butyl formate, filter, stir at room temperature for one day, filter again, and dry the solid under vacuum at 45°C to obtain the G crystal form.

[0129] The X-ray diffraction pattern of the G crystal form of the obtained compound of formula I is shown in Figure 9. The specific characteristic peaks are shown in Table 7 below, with an error of ±0.2°.

[0130] The DSC spectrum of the G crystal form of the compound of formula I has endothermic peaks at 131.6±5℃ and 225.1±5℃; the DSC spectrum of the G crystal form of the compound of formula I is shown in Figure 11.

[0131] Table 7. X-ray diffraction data of the G crystal form of compound I.

[0132] Example 9

[0133] Preparation of the H crystal form of compound I:

[0134] Weigh 2g of the A crystal form of compound I prepared in Example 2, add 5mL of acetic acid, stir at room temperature for 20 minutes, then add 10mL of isopropyl ether, continue stirring at room temperature for 1 day, filter, and dry the solid under vacuum at 45°C to obtain the H crystal form.

[0135] The X-ray diffraction pattern of the H crystal form of the obtained compound of formula I is shown in Figure 10. The specific characteristic peaks are shown in Table 8 below, with an error of ±0.2°.

[0136] The DSC spectrum of the H crystal form of the compound of formula I has endothermic peaks at 110.4±5℃, 124.8±5℃, 132.5±5℃ and 225.6±5℃; the DSC spectrum of the G crystal form of the compound of formula I is shown in Figure 12.

[0137] Table 8. X-ray diffraction data of the H crystal form of compound I.

[0138] Example 10 Bioactivity Assessment

[0139] Detection methods

[0140] In this paper, the inventors used the H295R Steroidogenesis Assay System to test the enzyme activities of human CYP11B1, human CYP11B2, etc. The in vitro H295R Steroidogenesis Assay System utilizes the human adrenal cancer cell line (NCI-H295R cells) to construct a level 2 "in vitro assay, providing mechanistic data" for screening and prioritization purposes. The development and standardization of this method were carried out in a multi-step process for screening the chemical effects of steroidogenesis. The H295R assay method has been optimized and validated according to the OECD test guideline (Test Guideline No. 456 H295R Steroidogenesis Assay).

[0141] Inhibition of aldosterone synthase

[0142] NCI-H295R cells can be purchased from ATCC. After culturing H295R cells from the original ATCC batch, the cells should be cultured for five generations (i.e., the cells divide four times), and then the cells that have been passaged five times should be frozen and stored in liquid nitrogen.

[0143] H295R cells were cultured in a 37°C, 5% CO2 incubator, with the culture medium changed 2-3 times per week. Cells were passaged when they reached approximately 85-90% confluence. The culture medium was then aspirated and replaced with DPBS (calcium-free). 2+ Mg 2+ Wash three times, digest with trypsin for 1-3 min, add 3 mL of culture medium to stop digestion and blow off the cells, then wash off the remaining cells with 1 mL of culture medium and add to a 15 mL centrifuge tube. Centrifuge at 800 rpm for 5 min at room temperature, discard the supernatant, resuspend the pellet with 3 mL of culture medium, and count the cell suspension. Discard the edge wells of the 96-well plate, and seed 50,000 cells into each of the remaining wells. Add 100 μL of 10% FBSDMEM:F12 (1:1) basal medium per well and incubate overnight. Replace with 150 μL of basal medium containing 10 μM Forskolin and incubate for 48 h. After 48 h, replace with basal medium containing 10 μM deoxycorticosterone. The compound is dissolved in DMSO to prepare a 100 mM stock solution. Starting at 100 mM, perform a 3-fold serial dilution in DMSO to obtain 10 concentration points. Ten concentration points were further diluted 10-fold with DMEM:F12 (1:1) blank medium, with an initial concentration of 10 mM. 1.5 μL of each concentration of the compound was added to the cells, with a final DMSO concentration of 0.1% and an initial compound concentration of 100 μM. After incubation for 48 h, 40 μL of cell supernatant was collected, and aldosterone and cortisol levels were analyzed using LCMS.

[0144] Cell viability assay

[0145] After collecting the supernatant, add 100 μL of 10% CCK8 assay reagent to each well, incubate at 37°C for 10 min, mix thoroughly by tapping, and then measure the OD value at 405 nm using a microplate reader. A 70% methanol group was set as a negative control, and DMSO solvent controls were set as a positive control. The following formula was used to calculate:

[0146] %viable cells=(OD cmpd–OD Avg MeOH[=100%dead])÷(OD Avg SCs[=100%viability]–OD Avg MeOH[=100%dead])

[0147] Wells with cell viability below 80% should not be included in the final data analysis. In cases of cytotoxicity approaching 20%, inhibition of steroid production should be carefully evaluated to ensure that cytotoxicity is not the cause of inhibition. Furthermore, data with cell viability exceeding 120% should be labeled to identify potential false positives.

[0148] The inhibition rate was calculated using the following formula in the results analysis:

[0149] Inhibition rate % = (PeakArea Avg SCs - PeakArea cmpd) / (PeakArea Avg SCs - Peak Areablank) × 100

[0150] Plotting the logarithm of compound concentration on the x-axis and inhibition rate on the y-axis, a nonlinear regression curve was fitted using Graphpad 9.0 to calculate the IC50 value (Y = Bottom + (Top - Bottom) / (1 + 10^((LogIC50 - X) * HillSlope))), Ki = IC50 / (1 + [S] / Km). The test results are shown in Table 9. Unless otherwise specified, it is assumed that all proteases are competitively inhibited. Selectivity = CYP11B1Ki(nM) / CYP11B2 Ki(nM); where A represents a selectivity value between 0 and 50, B represents a selectivity value between 51 and 100, C represents a selectivity value between 101 and 150, and D represents a selectivity value above 151.

[0151] Table 9. Inhibitory effects of compounds on CYP11B2

[0152] As shown in Table 9, the experimental results of this invention have a good inhibitory effect on CYP11B2, which is better than that of the control compound Baxdrostat. In addition, the compound of this invention has excellent selectivity for CYP11B2, which can selectively inhibit CYP11B2 while weakly inhibiting CYP11B1.

[0153] Example 11: Rat Pharmacokinetic Study

[0154] Experimental materials

[0155] SD rats: male, 180-250g, purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd.

[0156] Reagents: DMSO (dimethyl sulfoxide), PEG-400 (polyethylene glycol 400), physiological saline, heparin, acetonitrile, formic acid, and propranolol (internal standard) are all commercially available.

[0157] Instrument: AB SCIEX QTRAP 5500+.

[0158] Experimental methods

[0159] The compounds from Examples 1-49 of this invention were weighed and dissolved in a DMSO-PEG-400-physiological saline (5:60:35, v / v / v) system. After intravenous or gavage administration to rats, 200 μL of venous blood was collected at 15 min, 30 min, 1 h, 2 h, 5 h, 7 h, and 24 h (an additional 5 min for the IV group) into EDTA-K2 anticoagulant tubes. The tubes were centrifuged at 12000 rpm for 2 min, and the plasma was stored at -80℃ for later analysis. A precise amount of the test sample was dissolved in DMSO to a concentration of 2 mg / mL to prepare a stock solution. An appropriate amount of the stock solution was accurately pipetted and diluted with acetonitrile to prepare a series of standard solutions. Accurately pipette 10 μL of each of the above standard series solutions, add 90 μL of blank plasma, vortex to mix, and prepare plasma samples with concentrations equivalent to 1, 3, 5, 10, 30, 100, 300, 1000, and 3000 ng / mL. Perform dual-sample analysis for each concentration to establish a standard curve. Take 30 μL of plasma (diluted 5-fold after intravenous administration at 5 min, 15 min, and 30 min), add 150 μL of acetonitrile solution of propranolol (50 ng / mL) as internal standard, vortex to mix, add 100 μL of purified water, vortex again, centrifuge at 4000 rpm for 5 min, and analyze the supernatant by LC-MS. The LC-MS detection conditions are as follows:

[0160] Chromatographic column: YMC Triart C18, 50*3.0mm, 2.1μm.

[0161] Mobile phase: water (0.1% formic acid) - acetonitrile. Gradient elution is performed according to the table below.

[0162] Data processing

[0163] After LC-MS detection of blood drug concentration, pharmacokinetic parameters were calculated using WinNonlin 6.1 software and the non-compartmental model method. The test results are shown in Table 10.

[0164] Table 10 Pharmacokinetics of the compounds of the present invention in rats

[0165] As shown in Table 10, the compounds of this invention exhibit good pharmacokinetic characteristics in SD rats, with C60 levels observed after intravenous and gavage administration. max and AUC last All were superior to the positive control, with good absorption, high absolute bioavailability, and half-life comparable to or better than the control compound.

[0166] Example 12: Pharmacokinetic Study of Compounds in Crab-Eating Monkeys

[0167] Experimental materials

[0168] Crab-eating macaque: Male, 180-250g, purchased from Guangxi Xiongsen Primate Experimental Animal Breeding and Development Co., Ltd.

[0169] Reagents: DMSO (dimethyl sulfoxide), PEG400, physiological saline, heparin, acetonitrile, formic acid, and propranolol (internal standard) were all commercially available.

[0170] Instrument: AB SCIEX 7500.

[0171] Experimental methods

[0172] The compound was dissolved in a DMSO-PEG-400-physiological saline (5:60:35, v / v / v) system. After administration by gavage to cynomolgus monkeys, 200 μL of venous blood was collected at 30 min, 60 min, 90 min, 2 h, 3 h, 5 h, 8 h, and 24 h in EDTA-K2 anticoagulant tubes. The tubes were centrifuged at 12000 rpm for 2 min, and the plasma was stored at -80℃ for later analysis. A precise amount of the test sample was dissolved in DMSO to a concentration of 2 mg / mL to prepare a stock solution. An appropriate amount of the stock solution was accurately pipetted and diluted with acetonitrile to prepare a series of standard solutions. Accurately pipette 10 μL of each of the above standard series solutions and add 90 μL of blank plasma. Vortex to mix, preparing plasma samples with concentrations equivalent to 0.3, 1, 3, 10, 30, 100, 300, 1000, and 3000 ng / mL, and quality control samples with concentrations of 2.4, 120, and 2400 ng / mL. Perform dual-sample analysis for each concentration and establish a standard curve. Take 30 μL of plasma and add 150 μL of acetonitrile solution containing propranolol (50 ng / mL) as internal standard. Vortex to mix, then add 100 μL of purified water, vortex again, centrifuge at 4000 rpm for 5 min, and analyze the supernatant by LC-MS. The LC-MS detection conditions are as follows:

[0173] Chromatographic column: YMC Triart C18, 50*3.0mm, 2.1μm.

[0174] Mobile phase: water (0.1% formic acid) - acetonitrile. Gradient elution is performed according to the table below.

[0175] Data processing

[0176] After LC-MS detection of blood drug concentration, pharmacokinetic parameters were calculated using WinNonlin 6.1 software and the non-compartmental model method. The test results are shown in Table 11.

[0177] Table 11 Pharmacokinetic results of the compounds of this invention in cynomolgus monkeys

[0178] As shown in Table 11, the experimental results of this invention indicate that all compounds exhibit good pharmacokinetic characteristics in cynomolgus monkeys. After gavage administration, the post-absorption exposure levels were higher or comparable to the positive control Baxdrostat. max and AUC last All are superior to the positive control, with a better half-life, better absorption, and higher absolute bioavailability.

[0179] Example 13 Stability Study:

[0180] Stability Study of Influencing Factors: The A crystal form of compound I prepared in Example 2, the B crystal form of compound I prepared in Example 3, and the H crystal form of compound I prepared in Example 9 were laid out in open positions and their stability was investigated under high temperature (60℃ / 80℃) and high humidity (RH 92.5%) conditions. The sampling period was 5 or 6 days. The results are shown in Table 12.

[0181] Table 12 Results of the stability study

[0182] Note: " / " indicates that it has not been tested.

[0183] As can be seen from the results in Table 12, the A and B crystal forms of the present invention have excellent stability under both high temperature and high humidity conditions. The H crystal form has good stability under high temperature conditions, but it will undergo crystal transformation under high humidity conditions.

[0184] Example 14 Solubility test:

[0185] Stock solution of test sample: Take an appropriate amount of the test sample (excessive until it cannot be dissolved) and place it in a 25 mL volumetric flask. Accurately measure 5 mL of each pH buffer solution and pour it into the volumetric flask. Incubate at 37°C with constant temperature shaking (135 rpm) for 24 h. After the time is up, remove the flask and filter while hot (nylon, 0.22 μm).

[0186] Test solution: While still hot, accurately measure 1 mL of the filtered test solution into a volumetric flask, dilute to the mark with diluent, and shake well to obtain the test solution.

[0187] Table 13 Solubility Results

[0188] As can be seen from the results in Table 13, at pH 1.0, both crystal forms of compound I, A and B, have good solubility, while at pH 6.8 mg / mL, the solubility of crystal form A of compound I is slightly better than that of crystal form B.

[0189] Example 15 Moisture Adsorption Test

[0190] The sample was placed on an SMS Intrinsic dynamic moisture adsorption analyzer to collect data. Starting from 0 RH, a humidity gradient of 5% RH was used, with the final humidity at 95% RH. The weight change was collected every 5 seconds, and the weight change under 95% RH conditions was taken as the final moisture absorption of the product. The results are shown in Table 14.

[0191] Table 14. Moisture absorption of the crystal forms of compounds of Formula I of the present invention

[0192] As can be seen from the results in Table 14, the hygroscopicity of both crystal forms A and B of the compound of Formula I of the present invention is low, both lower than that of the compound of Formula I in Example 1. Among them, the hygroscopicity of crystal form A of the compound of Formula I is the lowest.

[0193] Example 16: Pharmacokinetic study of the crystal form of compound of formula I in beagle dogs.

[0194] 1. Experimental Materials

[0195] Beagle: Male, 8-13kg, Beijing Mars Biotechnology Co., Ltd.

[0196] Test substances: Crystal form of compound A of formula I, crystal form of compound B of formula I, and crystal form of compound H of formula I.

[0197] Reagents: Methanol, formic acid, and propranolol (internal standard) are all commercially available. Compound I and Compound I-D5 (internal standard) were prepared by our company.

[0198] Instrument: AB SCIEX TripleQuad5500+ triple quadrupole mass spectrometer.

[0199] 2. Experimental Methods

[0200] Beagle dogs were used for capsule administration experiments of each test substance. The dose of each test substance was 50 mg based on the free amount of compound I. Three dogs were administered each test substance. Venous blood samples of approximately 0.5 mL were collected at 15 min, 30 min, 1 h, 2 h, 4 h, 6 h, 8 h, and 24 h post-administration in anticoagulant tubes containing EDTA-K2. The samples were centrifuged at 4000 rpm for 10 min at 2–8 °C, and the plasma was stored at -80 °C for later analysis. A precise amount of compound I was dissolved in DMSO to a concentration of 2 mg / mL to prepare a stock solution. The stock solution of compound I was accurately pipetted and diluted with acetonitrile-water to prepare a series of standard solutions. 10 μL of each of the above standard solutions was accurately pipetted and added to 90 μL of blank plasma. The mixture was vortexed to prepare plasma samples with concentrations equivalent to 1–10000 ng / mL. Two-sample analysis was performed for each concentration to establish a standard curve. Take 30 μL of plasma, add 200 μL of acetonitrile methanol solution containing internal standard, vortex to mix, centrifuge at 6000 rpm for 10 min at 2–8 °C, take 150 μL of supernatant, add 150 μL of purified water, vortex to mix again, and detect compound I using LC-MS / MS.

[0201] 3. Data Processing

[0202] After detecting blood drug concentrations by LC-MS, pharmacokinetic parameters were calculated using WinNonlin software and a non-compartmental model method.

[0203] 4. Experimental Results

[0204] Table 15 shows the comparison of the absorption of different crystal forms of compound I administered as gastric-soluble capsules in beagle dogs.

[0205] Table 15 Comparison of different crystal forms of the compound of Formula I in beagle pk-dog administration

[0206] As can be seen from the results in Table 15, the A and B crystal forms of the compound of formula I of this invention have good pharmacokinetic properties, and C... max and AUC last Both are superior to the H crystal form of the compound of formula I.

[0207] It should be understood that the above embodiments are preferred embodiments of the present invention, but the embodiments of the present invention are not limited to the above embodiments. Those skilled in the art can make improvements or modifications based on the above description, and all such improvements and modifications should fall within the protection scope of the appended claims of the present invention.

Claims

1. A crystalline form of the compound of Formula I, characterized by, The A crystalline form of the compound of Formula I has characteristic peaks in an X-ray diffraction pattern expressed in degrees of 2 theta (2Q) at 13.45°, 13.62°, 23.21°, and 23.95°, with a + / - 0.2° error; preferably, the A crystalline form of the compound of Formula I has characteristic peaks in an X-ray diffraction 2. The A crystalline form of the compound of formula I according to claim 1, characterized in that, The DSC pattern of the A crystalline form of the compound of formula I has an endothermic peak at 224.3 °C ± 5 °C; preferably, the A crystalline form of the compound of formula I has a DSC pattern substantially as shown in Figure 2.

3. A B crystalline form of a compound of Formula I, characterized by, The B crystalline form of the compound of Formula I has characteristic peaks in an X-ray diffraction pattern, expressed in terms of 2 theta (2Q) angles, at 11.19° and 18.76°, with a + / - 0.2° error; preferably, the B crystalline form of the compound of Formula I has characteristic peaks in an X-ray diffraction pattern, expressed in terms of 2 theta (2Q) angles, at 9.27°, 11.19°, 18.76°, 23.43°, and 24.79°, with a + / - 0.2° error; more preferably, the B crystalline form of the compound of Formula I has characteristic peaks in an X-ray diffraction pattern, expressed in terms of 2 theta (2Q) angles, at 9.27°, 11.19°, 18.76°, 20.11°, 21.31°, 22.27°, 23.00°, 23.43°, 24.79°, 25.25°, and 25.59°, with a + / - 0.2° error; most preferably, the B crystalline form of the compound of Formula I has an XRPD pattern substantially as shown in FIG. 3; the structure of the compound of Formula I is shown below:

4. The B crystalline form of the compound of formula I according to claim 3, characterized in that, The DSC pattern of the B crystalline form of the compound of formula I has an endothermic peak at 225.0 ± 5 °C; preferably, the B crystalline form of the compound of formula I has a DSC pattern substantially as shown in Figure 5. A crystalline form of the compound of formula I, characterized by, The C crystalline form of the compound of Formula I has characteristic peaks in an X-ray diffraction pattern expressed in terms of 2 theta (2Q) angle at 12.05°, 13.56°, 15.18°, and 17.74°, with a margin of error of ± 0.2°; preferably, the C crystalline form of the compound of Formula I has characteristic peaks in an X-ray diffraction pattern expressed in terms of 2 theta (2Q) angle at 9.08°, 12.05°, 13.56°, 15.18°, and 17.74°, with a margin of error of ± 0.2°; more preferably, the C crystalline form of the compound of Formula I has characteristic peaks in an X- ray diffraction pattern expressed in terms of 2 theta (2Q) angle at 8.27°, 9.08°, 11.13°, 12.05°, 12.38°, 13.56°, 15.18°, 17.74°, 18.09°, and 21.53°, with a margin of error of ± 0.2°; most preferably, the C crystalline form of the compound of Formula I has an XRPD pattern substantially as shown in FIG. 4; the structure of the compound of Formula I is shown below:

6. A crystalline form of the compound of formula I, characterized by, The D crystalline form of the compound of Formula I has characteristic peaks in an X- ray diffraction pattern, expressed in terms of 2 theta (2Q) angles, at 8.52°, 13.13°, 16.94°, 19.91°, and 24.90°, with a + / - 0.2° error; preferably, the D crystalline form of the compound of Formula I has characteristic peaks in an X-ray diffraction pattern, expressed in terms of 2 theta (2Q) angles, at 8.52°, 7. An E crystalline form of a compound of Formula I, characterized by, The E crystalline form of the compound of Formula I has characteristic peaks in an X- ray diffraction pattern expressed in terms of 2 theta (2Q) angle at 18.80° and 24.82°, with a + / - 0.2° error; preferably, the E crystalline form of the compound of Formula I has characteristic peaks in an X-ray diffraction pattern expressed in terms of 2 theta (2Q) angle at 9.28°, 9.46°, 18.80° and 24.82°, with a + / - 0.2 ° error; more preferably, the E crystalline form of the compound of Formula I has characteristic peaks in an X- ray diffraetion pattern expressed in terms of 2 theta (2Q) angle at 8.63°, 9.28°, 9.46°, 11.34°, 14.14°, 18.80°, 24.82° and 29.31°, with a + / - 0.2° error; most preferably, the E crystalline form of the compound of Formula I has an XRPD pattern substantially as shown in Figure 6; the structure of the compound of Formula I is shown below:

8. A crystalline form of the compound of Formula I, characterized by: The F crystalline form of the compound of Formula I has characteristic peaks in an X- ray diffraction pattern, expressed in terms of 2 theta (2Q) angles, at 8.27°, 13.78°, and 23.60°, ± 0.2°; preferably, the F crystalline form of the compound of Formula I has characteristic peaks in an X-ray diffraction pattern, expressed in terms of 2 theta (2Q) angles, at 3.57°, 8.27°, 13.78°, 23.60°, and 25.28°, ± 0.2°; more preferably, the F crystalline form of the compound of Formula I has characteristic peaks in an X- ray diffraetion pattern, expressed in terms of 2 theta (2Q) angles, at 3. 57°, 8.27°, 11.18°, 13.78°, 16.65°, 18.42°, 23.60°, 25.28°, 26.10°, 26.64°, 27.14°, and 28.16°, ± 0.2°; most preferably, the F crystalline form of the compound of Formula I has an XRPD pattern substantially as shown in FIG. 7; the structure of the compound of Formula I is shown below:

9. A G crystalline form of a compound of Formula I, characterized by, The G crystalline form of the compound of Formula I has characteristic peaks in an X- ray diffraction pattern expressed in terms of 2 theta (2Q) angle at 12.12°, 13.62°, 20.21°, 20.51°, 22.89°, and 25.60°, with a + / - 0.2° error; preferably, the G crystalline form of the compound of Formula I has characteristic peaks in an X-ray diffraction pattern expressed in terms of 2 theta (2Q) angle at 12. 12°, 13.62°, 15.25°, 17.72°, 20.21°, 20.51°, 21.19°, 21.37°, 22.89°, and 25.60°, with a + / - 02° error; more preferably, the G crystalline form of the compound of Formula I has characteristic peaks in an X- ray diffraetion pattern expressed in terms of 2 theta (2Q) angle at 8.37°, 9.12°, 12.12°, 12.46°, 13.62°, 15.25°, 17.72°, 10. A crystalline form of the compound of Formula I, characterized by: The H crystalline form of the compound of Formula I has characteristic peaks in an X-ray diffraction pattern expressed in degrees of 2 theta (2Q) at 10.20° and 19.14°, with an error of ± 0.2°; preferably, the H crystalline form of the compound of Formula I has characteristic peaks in an X-ray diffraction pattern expressed in degrees of 2 theta (2Q) at 9.44°, 10.20°, 17.63°, 19.14°, 19.43°, 20.94°, and 24.93°, with an error of ± 0.2°; more preferably, the H crystalline form of the compound of Formula I has characteristic peaks in an X- ray diffraction pattern expressed in degrees of 2 theta (2Q) at 9.44 °, 10.20°, 15.20°, 17.63°, 19.14°, 19. 43°, 20.25°, 20.94°, 23.00°, 24.93°, and 25.40°, with an error of ± 0.2°; most preferably, the H crystalline form of the compound of Formula I has an XRPD pattern substantially as shown in FIG. 10; the structure of the compound of Formula I is shown below:

11. A pharmaceutical composition, characterized by, The pharmaceutical composition comprises a crystalline form according to any one of claims 1 to 10 and one or more pharmaceutically acceptable excipients and / or carriers.

12. Use of a crystalline form according to any one of claims 1 to 10 or of a pharmaceutical composition according to claim 11 for the manufacture of a medicament for the treatment or prevention of a disease associated with elevated CYP11B2 activity levels; preferably, the disease associated with elevated CYP11B2 activity levels is selected from the group consisting of hypertension, chronic kidney disease, primary aldosteronism, diabetic nephropathy, congestive heart failure or Cushing's syndrome.

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