Crystal form of pyridinephenyl compound and preparation method thereof

Abstract

A crystalline form of a pyridinephenyl compound and a method for preparing the same, including the use of the crystalline form in preparing drugs for treating related diseases. #imgabs0#

Description

[0001] This application claims priority to

[0002] CN202011044638.7, application date: September 28, 2020. Technical Field

[0003] The present invention relates to a crystal form of a pyridinephenyl compound and a preparation method thereof, and also relates to application of the crystal form in preparing medicines for treating related diseases. Background Art

[0004] Dry eye, also known as keratoconjunctivitis sicca, refers to a broad range of conditions characterized by abnormal tear quality, quantity, or dynamics, resulting in decreased tear film stability and accompanied by ocular discomfort or ocular surface pathology. Specific symptoms include eye irritation, visual impairment, and tear film instability. Some cases of this syndrome are caused by ocular surface inflammation, leading to lacrimal gland dysfunction. Systemic autoimmunity is also associated with this condition.

[0005] Because some toxic aldehydes are produced by the body or eye tissues and organs through metabolic mechanisms, such as malondialdehyde (MDA) and 4-hydroxy-2-nonenal (4HNE), these aldehydes are highly reactive with proteins, carbohydrates, oils and DNA, leading to chemical modification of biological molecules and activation of inflammatory molecule regulators such as NF-kappaB, thereby causing damage to different organs. This is one of the causes of dry eye.

[0006] Through research, the present invention has discovered that a small molecule drug, administered as eye drops to the site of ocular inflammation, reacts with aldehydes in the body to reduce aldehyde toxicity and inflammation, thereby achieving the therapeutic effect of dry eye. The compound of formula (II) has excellent chemical stability, with one hydrate molecule being water of crystallization, arranged in a dense crystalline form. The compound of formula (II) can effectively complex active aldehyde molecules both in vivo and in vitro, and can effectively penetrate into the cornea, thereby achieving an anti-inflammatory effect in treating dry eye. After eye drops were administered to animals, the highest concentration of the compound of formula (II) eye drops did not cause any adverse reactions in the animals, indicating that it is safe and effective for potential drug development. Summary of the Invention

[0007] The present invention provides a crystal form A of a compound of formula (I), characterized in that its X-ray powder diffraction pattern has characteristic diffraction peaks at the following 2θ angles: 7.14±0.20°, 12.46±0.20°, and 18.87±0.20°.

[0008]

[0009] In some embodiments of the present invention, the above-mentioned crystal form A has an X-ray powder diffraction pattern having characteristic diffraction peaks at the following 2θ angles: 7.14±0.20°, 12.46±0.20°, 15.99±0.20°, 17.06±0.20°, 18.87±0.20°, 20.25±0.20°, 21.41±0.20°, and 25.00±0.20°.

[0010] In some embodiments of the present invention, the above-mentioned crystal form A has an X-ray powder diffraction pattern having characteristic diffraction peaks at the following 2θ angles: 7.14±0.20°, 10.11±0.20°, 12.46±0.20°, 14.35±0.20°, 15.99±0.20°, 17.06±0.20°, 18.87±0.20°, 20.25±0.20°, 21.41±0.20°, and 25.00±0.20°.

[0011] The present invention provides a crystal form A of a compound of formula (I), characterized in that its X-ray powder diffraction pattern has characteristic diffraction peaks at the following 2θ angles: 7.142±0.200°, 12.456±0.200°, and 18.868±0.200°.

[0012]

[0013] In some embodiments of the present invention, the above-mentioned crystal form A has an X-ray powder diffraction pattern having characteristic diffraction peaks at the following 2θ angles: 7.142±0.200°, 12.456±0.200°, 15.993±0.200°, 17.057±0.200°, 18.868±0.200°, 20.249±0.200°, 21.413±0.200°, and 24.995±0.200°.

[0014] In some embodiments of the present invention, the above-mentioned crystal form A has an X-ray powder diffraction pattern having characteristic diffraction peaks at the following 2θ angles: 7.142±0.200°, 12.456±0.200°, 14.347±0.200°, 15.993±0.200°, 17.057±0.200°, 18.868±0.200°, 20.249±0.200°, 21.413±0.200°, 24.995±0.200°, 25.897±0.200°, 29.566±0.200°, and 30.442±0.200°.

[0015] In some embodiments of the present invention, the above-mentioned crystal form A has an X-ray powder diffraction pattern having characteristic diffraction peaks at the following 2θ angles: 7.142°, 8.234°, 10.112°, 11.302°, 12.056°, 12.377°, 12.456°, 14.086°, 14.347°, 15.993°, 16.530°, 16.834°, 17.057°, 17.482°, 18.868°, 20.249°, 21.413°, 22.599°, 22.721°, 23.750°, 23.941°, 24.191°, 24.763°, 24.995°, 25.897°, 27.931°, 29.566°, 30.442°, 31.373°, 31.582°, 32.162°, 32.936°, 33.830°, 34.537°, 34.774°, 35.312°, 36.107°, 36.401°, 37.575°.

[0016] In some embodiments of the present invention, the above-mentioned crystal form A has an XRPD pattern substantially as follows Figure 1 shown.

[0017] In some embodiments of the present invention, the XRPD pattern analysis data of the above-mentioned Form A are shown in Table 1:

[0018] Table 1 XRPD pattern analysis data of the crystal form of compound A of formula (I)

[0019]

[0020]

[0021] In some embodiments of the present invention, the differential scanning calorimetry curve of the above-mentioned crystal form A has an endothermic peak starting point at 81.03±3.0°C, 113.62±3.0°C and 151.37±3.0°C, respectively.

[0022] In some embodiments of the present invention, the DSC spectrum of the above-mentioned crystal form A is substantially as follows Figure 2 shown.

[0023] In some embodiments of the present invention, the thermogravimetric analysis curve of the above-mentioned crystal form A shows a weight loss of 2.046% at 96.01°C±3.0°C and a weight loss of 7.708% at 163.66°C±3.0°C.

[0024] In some embodiments of the present invention, the TGA spectrum of the above-mentioned crystal form A is substantially as follows Figure 2 shown.

[0025] The present invention provides a compound of formula (II),

[0026]

[0027] The present invention provides a crystal form B of a compound of formula (II), characterized in that its X-ray powder diffraction pattern has characteristic diffraction peaks at the following 2θ angles: 8.231±0.200°, 17.058±0.200°, and 18.955±0.200°.

[0028] In some embodiments of the present invention, the above-mentioned B crystal form has an X-ray powder diffraction pattern with characteristic diffraction peaks at the following 20 angles: 8.231±0.200°, 17.058±0.200°, 18.955±0.200°, 21.712±0.200°, and 25.678±0.200°.

[0029] In some embodiments of the present invention, the above-mentioned B crystal form has an X-ray powder diffraction pattern having characteristic diffraction peaks at the following 2θ angles: 8.231±0.200°, 12.068±0.200°, 16.505±0.200°, 17.058±0.200°, 18.955±0.200°, 21.712±0.200°, 24.242±0.200°, and 25.678±0.200°.

[0030] In some embodiments of the present invention, the above-mentioned B crystal form has an X-ray powder diffraction pattern having characteristic diffraction peaks at the following 2θ angles: 7.083±0.200°, 8.231±0.200°, 11.294±0.200°, 12.068±0.200°, 14.091±0.200°, 16.505±0.200°, 17.058±0.200°, 18.955±0.200°, 21.712±0.200°, 24.242±0.200°, 25.678±0.200°, and 30.869±0.200°.

[0031] The present invention provides a crystalline form B of a compound of formula (II), characterized in that its X-ray powder diffraction pattern has characteristic diffraction peaks at the following 2θ angles: 8.231±0.200°, 17.058±0.200°, and / or 18.955±0.200°, and / or 7.083±0.200°, and / or 11.294±0.200°, and / or 12.068±0.200°, and / or 14.091±0.200°, and / or 14.806±0.200°, and / or 16.505±0.200°, and / or 18.202±0.200°, and / or 21.712±0.200°, and / or 22. .182±0.200°, and / or 23.903±0.200°, and / or 24.242±0.200°, and / or 24.653±0.200°, and / or 25.350±0.200°, and / or 25.678±0.200°, and / or 26.270±0.200°, and / or 27.001±0.200°, and / or 27.658±0.200°, and / or 29.052±0.200°, and / or 29.721±0.200°, and / or 30.869±0.200°, and / or 34.644±0.200°, and / or 35.009±0.200°.

[0032] In some embodiments of the present invention, the above-mentioned B crystal form has an X-ray powder diffraction pattern having characteristic diffraction peaks at the following 2θ angles: 7.083°, 8.231°, 11.294°, 12.068°, 14.091°, 14.806°, 16.505°, 17.058°, 18.202°, ​​18.955°, 21.712°, 22.182°, 23.903°, 24.242°, 24.653°, 25.350°, 25.678°, 26.270°, 27.001°, 27.658°, 29.052°, 29.721°, 30.869°, 34.644°, and 35.009°.

[0033] In some embodiments of the present invention, the XRPD pattern of the above-mentioned Form B is substantially as follows Figure 3 shown.

[0034] In some embodiments of the present invention, the XRPD pattern analysis data of the above-mentioned Form B are shown in Table 2:

[0035] Table 2 XRPD pattern analysis data of the crystal form of compound B of formula (11)

[0036]

[0037] In some embodiments of the present invention, the differential scanning calorimetry curve of the above-mentioned B crystal form has an endothermic peak starting point at 101.7±3.0°C and 158.7±3.0°C, respectively.

[0038] In some embodiments of the present invention, the DSC spectrum of the above-mentioned B crystal form is substantially as follows Figure 4 shown.

[0039] In some embodiments of the present invention, the thermogravimetric analysis curve of the above-mentioned Form B shows a weight loss of 5.477% at 120.00°C±3.0°C.

[0040] In some embodiments of the present invention, the TGA spectrum of the above-mentioned B crystal form is substantially as follows Figure 5 shown.

[0041] The present invention provides a crystal form C of a compound of formula (I), characterized in that its X-ray powder diffraction pattern has characteristic diffraction peaks at the following 2θ angles: 12.871±0.200°, 17.488±0.200°, and 19.079±0.200°.

[0042]

[0043] In some embodiments of the present invention, the above-mentioned C crystal form has an X-ray powder diffraction pattern with characteristic diffraction peaks at the following 2θ angles: 12.871±0.200°, 17.488±0.200°, 18.403±0.200°, 19.079±0.200°, and 20.853±0.200°.

[0044] In some embodiments of the present invention, the above-mentioned C crystal form has an X-ray powder diffraction pattern having characteristic diffraction peaks at the following 2θ angles: 12.871±0.200°, 13.274±0.200°, 14.294±0.200°, 17.488±0.200°, 18.403±0.200°, 19.079±0.200°, 20.853±0.200°, and 21.468±0.200°.

[0045] In some embodiments of the present invention, the above-mentioned C crystal form has an X-ray powder diffraction pattern having characteristic diffraction peaks at the following 2θ angles: 10.105±0.200°, 12.871±0.200°, 13.274±0.200°, 14.294±0.200°, 17.488±0.200°, 18.403±0.200°, 19.079±0.200°, 20.853±0.200°, 21.468±0.200°, 22.647±0.200°, 23.977±0.200°, and 24.409±0.200°.

[0046] In some embodiments of the present invention, the above-mentioned C crystal form has an X-ray powder diffraction pattern having characteristic diffraction peaks at the following 2θ angles: 9.128°, 10.105°, 12.871°, 13.274°, 13.933°, 14.294°, 16.420°, 17.488°, 18.403°, 19.079°, 20.853°, 21.468°, 22.647°, 23.042°, 23.507°, 23.977°, 24.409°, 24.798°, 25.861°, 26.309°, 28.698°, 30.293°, and 37.464°.

[0047] In some embodiments of the present invention, the above-mentioned crystal form C has an XRPD pattern substantially as follows Figure 6 shown.

[0048] In some embodiments of the present invention, the XRPD pattern analysis data of the above-mentioned Form C are shown in Table 3:

[0049] Table 3 XRPD pattern analysis data of the crystal form of compound C of formula (I)

[0050]

[0051] In some embodiments of the present invention, the differential scanning calorimetry curve of the above-mentioned crystal form C has an endothermic peak starting point at 157.36±3.0°C.

[0052] In some embodiments of the present invention, the DSC spectrum of the above-mentioned crystal form C is substantially as follows Figure 7 shown.

[0053] In some embodiments of the present invention, the thermogravimetric analysis curve of the above-mentioned Form C shows a weight loss of 1.589% at 120.00°C±3.0°C.

[0054] In some embodiments of the present invention, the TGA spectrum of the above-mentioned C crystal form is substantially as follows Figure 7 shown.

[0055] The present invention also provides a compound of formula (III),

[0056]

[0057] Wherein, m is selected from 0-1, preferably 0, 0.25, 0.5 or 1.

[0058] In some embodiments of the present invention, the above compound has the structure of formula (III-1):

[0059]

[0060] The present invention provides a D crystal form of a compound of formula (III-1), characterized in that its X-ray powder diffraction pattern has characteristic diffraction peaks at the following 2θ angles: 6.600±0.200°, 17.775±0.200°, and 19.138±0.200°.

[0061] In some embodiments of the present invention, the above-mentioned D crystal form has an X-ray powder diffraction pattern with characteristic diffraction peaks at the following 2θ angles: 6.600±0.200°, 13.178±0.200°, 17.775±0.200°, 19.138±0.200°, and 25.798±0.200°.

[0062] In some embodiments of the present invention, the above-mentioned D crystal form has an X-ray powder diffraction pattern having characteristic diffraction peaks at the following 20 angles: 6.600±0.200°, 13.178±0.200°, 17.303±0.200°, 17.775±0.200°, 18.667±0.200°, 19.138±0.200°, 21.245±0.200°, and 25.798±0.200°.

[0063] In some embodiments of the present invention, the above-mentioned D crystal form has an X-ray powder diffraction pattern having characteristic diffraction peaks at the following 2θ angles: 6.600±0.200°, 11.004.200°, 13.178±0.200°, 15.521±0.200°, 16.592±0.200°, 17.303±0.200°, 17.775±0.200°, 18.667±0.200°, 19.138±0.200°, 21.245±0.200°, 25.798±0.200°, and 27.353±0.200°.

[0064] In some embodiments of the present invention, the above-mentioned D crystal form has an X-ray powder diffraction pattern with characteristic diffraction peaks at the following 2θ angles: 6.600°, 8.659°, 9.558°, 11.004°, 12.463°, 13.178°, 13.728°, 14.871°, 15.521°, 16.592°, 17.303°, 17.775°, 18.379°, 18.667°, 1 9.138°, 20.572°, 21.245°, 21.989°, 22.247°, 22.561°, 23.940°, 25.087°, 25.798°, 26.686°, 27.353°, 28.044°, 28.859°, 29.645°, 30.375°, 32.351°, 34.603°, 35.456°, 39.550°.

[0065] In some embodiments of the present invention, the XRPD pattern of the above-mentioned D crystal form is substantially as follows Figure 8 shown.

[0066] In some embodiments of the present invention, the XRPD pattern analysis data of the above-mentioned D crystal form are shown in Table 4:

[0067] Table 4 XRPD pattern analysis data of the crystal form of compound D of formula (III-1)

[0068]

[0069]

[0070] In some embodiments of the present invention, the differential scanning calorimetry curve of the above-mentioned D crystal form has an endothermic peak starting point at 116.98±3.0°C, 133.04±3.0°C and 154.86±3.0°C, respectively.

[0071] In some embodiments of the present invention, the DSC spectrum of the above-mentioned D crystal form is substantially as follows Figure 9 shown.

[0072] In some embodiments of the present invention, the thermogravimetric analysis curve of the above-mentioned D crystal form shows a weight loss of 6.939% at 150.00°C±3.0°C.

[0073] In some embodiments of the present invention, the TGA spectrum of the above-mentioned D crystal form is substantially as follows Figure 9 shown.

[0074] The present invention provides a compound of formula (IV),

[0075]

[0076] Wherein, n is selected from 0-1, preferably 0, 0.25, 0.5 or 1.

[0077] In some embodiments of the present invention, the above compound has the structure of formula (IV-1):

[0078]

[0079] The present invention provides a crystalline form E of a compound of formula (IV-1), characterized in that its X-ray powder diffraction pattern has characteristic diffraction peaks at the following 2θ angles: 18.097±0.200°, 18.691±0.200°, and 20.693±0.200°.

[0080] In some embodiments of the present invention, the above-mentioned E crystal form has an X-ray powder diffraction pattern with characteristic diffraction peaks at the following 2θ angles: 17.465±0.200°, 18.097±0.200°, 18.691±0.200°, 19.1790.200°, and 20.693±0.200°.

[0081] In some embodiments of the present invention, the above-mentioned E crystal form has an X-ray powder diffraction pattern having characteristic diffraction peaks at the following 2θ angles: 6.675±0.200°, 16.119±0.200°, 17.465±0.200°, 18.097±0.200°, 18.691±0.200°, 19.179±0.200°, 20.693±0.200°, and 26.658±0.200°.

[0082] In some embodiments of the present invention, the above-mentioned E crystal form has an X-ray powder diffraction pattern having characteristic diffraction peaks at the following 2θ angles: 6.675±0.200°, 8.741±0.200°, 11.391±0.200°, 13.762±0.200°, 16.119±0.200°, 17.465±0.200°, 18.097±0.200°, 18.691±0.200°, 19.179±0.200°, 20.693±0.200°, 23.386±0.200°, and 26.658±0.200°.

[0083] In some embodiments of the present invention, the X-ray powder diffraction pattern of the above-mentioned E crystal form has characteristic diffraction peaks at the following 2θ angles: 6.675°, 8.741°, 9.365°, 11.391°, 12.205°, 13.310°, 13.762°, 15.302°, 16.119°, 16.407°, 17.465°, 18.097°, 18.691°, 19.17 9°, 20.693°, 21.329°, 21.860°, 22.674°, 23.386°, 24.819°, 25.311°, 25.757°, 26.658°, 26.980°, 29.110°, 29.540°, 30.536°, 31.639°, 33.068°, 33.972°, 36.724°, 38.646°.

[0084] In some embodiments of the present invention, the above-mentioned E crystal form has an XRPD pattern substantially as follows Figure 10 shown.

[0085] In some embodiments of the present invention, the XRPD pattern analysis data of the above-mentioned E crystal form are shown in Table 5:

[0086] Table 5 XRPD pattern analysis data of the crystal form of compound E of formula (IV-1)

[0087]

[0088]

[0089] In some embodiments of the present invention, the differential scanning calorimetry curve of the above-mentioned E crystal form has an endothermic peak starting point at 106.62±3.0°C, 134.32±3.0°C and 158.33±3.0°C, respectively.

[0090] In some embodiments of the present invention, the DSC spectrum of the above-mentioned E crystal form is substantially as follows Figure 11 shown.

[0091] In some embodiments of the present invention, the thermogravimetric analysis curve of the above-mentioned E crystal form shows a weight loss of 6.673% at 150.00°C±3.0°C.

[0092] In some embodiments of the present invention, the TGA spectrum of the above-mentioned E crystal form is substantially as follows Figure 11 shown.

[0093] The present invention provides a compound of formula (V),

[0094]

[0095] The present invention provides a crystal form F of a compound of formula (V), characterized in that its X-ray powder diffraction pattern has characteristic diffraction peaks at the following 2θ angles: 14.012±0.200°, 16.936±0.200°, and 17.424±0.200°.

[0096] In some embodiments of the present invention, the above-mentioned F crystal form has an X-ray powder diffraction pattern having characteristic diffraction peaks at the following 2θ angles: 14.012±0.200°, 16.936±0.200°, 17.424±0.200°, 17.954±0.200°, and 22.043±0.200°.

[0097] In some embodiments of the present invention, the above-mentioned F crystal form has an X-ray powder diffraction pattern having characteristic diffraction peaks at the following 2θ angles: 12.567±0.200°, 14.012±0.200°, 15.666±0.200°, 16.936±0.200°, 17.424±0.200°, 17.954±0.200°, 20.717±0.200°, and 22.043±0.200°.

[0098] In some embodiments of the present invention, the above-mentioned F crystal form has an X-ray powder diffraction pattern having characteristic diffraction peaks at the following 2θ angles: 6.104±0.200°, 12.567±0.200°, 14.012±0.200°, 15.666±0.200°, 16.936±0.200°, 17.424±0.200°, 17.954±0.200°, 20.717±0.200°, 22.043±0.200°, and 25.436±0.200°.

[0099] In some embodiments of the present invention, the above-mentioned F crystal form has an X-ray powder diffraction pattern having characteristic diffraction peaks at the following 20 angles: 6.104°, 9.825°, 12.567°, 14.012°, 15.666°, 16.936°, 17.424°, 17.954°, 19.298°, 20.717°, 21.106°, 22.043°, 25.436°, 26.230°, 26.94°, and 29.721°.

[0100] In some embodiments of the present invention, the XRPD pattern of the above-mentioned F crystal form is substantially as follows Figure 12 shown.

[0101] In some embodiments of the present invention, the XRPD pattern analysis data of the above-mentioned Form F are shown in Table 6:

[0102] Table 6 XRPD pattern analysis data of the crystal form F of the compound of formula (V)

[0103]

[0104] In some embodiments of the present invention, the differential scanning calorimetry curve of the above-mentioned F crystal form has an endothermic peak starting point at 83.44±3.0°C and 154.65±3.0°C, respectively.

[0105] In some embodiments of the present invention, the DSC spectrum of the above-mentioned F crystal form is substantially as follows Figure 13 shown.

[0106] In some embodiments of the present invention, the thermogravimetric analysis curve of the above-mentioned F crystal form shows a weight loss of 2.972% at 120.00°C±3.0°C.

[0107] In some embodiments of the present invention, the TGA spectrum of the above-mentioned F crystal form is substantially as follows Figure 13 shown.

[0108] The present invention also provides use of the above-mentioned compound or crystal form A, crystal form B, crystal form C, crystal form D, crystal form E or crystal form F in the preparation of a drug for treating dry eye.

[0109] Definition and Description

[0110] Unless otherwise indicated, the following terms and phrases used herein are intended to have the following meanings. A particular phrase or term should not be construed as ambiguous or unclear unless specifically defined, but rather should be understood in accordance with its ordinary meaning. When a trade name appears herein, it is intended to refer to the corresponding commercial product or its active ingredient.

[0111] The intermediate compounds of the present invention can be prepared by a variety of synthetic methods well known to those skilled in the art, including the specific embodiments listed below, embodiments formed by combining them with other chemical synthesis methods, and equivalent replacement methods well known to those skilled in the art. Preferred embodiments include but are not limited to the examples of the present invention.

[0112] The chemical reactions of the present invention are carried out in suitable solvents that are compatible with the chemical transformations of the present invention and the reagents and materials required. To obtain the compounds of the present invention, it may sometimes be necessary for those skilled in the art to modify or select synthetic steps or reaction schemes based on existing embodiments.

[0113] The structures of the compounds of the present invention can be confirmed by conventional methods well known to those skilled in the art. If the present invention relates to the absolute configuration of the compounds, the absolute configuration can be confirmed by conventional techniques in the art. For example, single crystal X-ray diffraction (SXRD) is used to collect diffraction intensity data on the cultured single crystal using a Bruker D8 venture diffractometer, using CuKα radiation as the light source and scanning mode: After scanning and collecting relevant data, the crystal structure is further analyzed using the direct method (Shelxs97) to confirm the absolute configuration.

[0114] The present invention will be described in detail below through examples, which are not intended to limit the present invention in any way.

[0115] All solvents used in the present invention were commercially available and used without further purification.

[0116] The solvents used in the present invention are commercially available. The present invention uses the following abbreviations: DCM stands for dichloromethane; DMF stands for N,N-dimethylformamide; DMSO stands for dimethyl sulfoxide; EtOH stands for ethanol; and MeOH stands for methanol.

[0117] Technical Effects

[0118] The compound of the present invention has good crystal stability and is easy to formulate into medicine. The crystal eye drops of compound B of formula (II) can have a good therapeutic effect on a mouse dry eye model induced by scopolamine hydrobromide solution, mainly improving the tear secretion and corneal damage of the dry eye model mice. The crystal eye drops of compound B of formula (II) can have a good therapeutic effect on a rat dry eye model induced by a hypertonic sodium chloride solution, mainly improving the tear secretion, corneal damage and tear film rupture time of the dry eye model rats.

[0119] X-ray powder diffractometer (XRPD) method of the present invention

[0120] Instrument model: Bruker D8 Advance X-ray diffractometer

[0121] Test method: Approximately 10 mg of sample was used for XRPD analysis.

[0122] The detailed XRPD parameters are as follows:

[0123] Plain copper tube / K-Alphal

[0124] Voltage 40 kilovolts (kV)

[0125] Current 40 milliamperes (mA)

[0126] Speed ​​15 [r / min]

[0127] Scanning range from 3 to 39.9978[°]

[0128] Step size 0.020428946300[°]

[0129] Time / step (s): 0.12 [s]

[0130] Total time: 234.72 seconds

[0131] Differential Scanning Calorimeter (DSC) method of the present invention

[0132] Instrument model: DISCOVERY DSC-2500 differential scanning calorimeter

[0133] Test method: Take a sample (0.5mg~1mg) and place it in a DSC aluminum pan for testing. Under 50mL / min N2 conditions, heat the sample from room temperature to 250℃ at a heating rate of 10℃ / min.

[0134] Thermogravimetric analysis (TGA) method of the present invention

[0135] Instrument model: DISCOVERY TGA 5500 thermogravimetric analyzer

[0136] Test method: Take a sample (2-5 mg) and place it in a TGA platinum pan for testing. Under 25 mL / min N2 conditions, heat the sample from room temperature to 300°C or 20% weight loss at a heating rate of 10°C / min. BRIEF DESCRIPTION OF THE DRAWINGS

[0137] Figure 1 The XRPD spectrum of the crystal form A of the compound of formula (1) under Cu-Kα radiation.

[0138] Figure 2 The DSC and TGA spectra of the crystal form A of compound of formula (I) are shown.

[0139] Figure 3 The XRPD spectrum of the crystal form B of the compound of formula (II) is obtained using Cu-Kα radiation.

[0140] Figure 4 The DSC spectrum of the crystal form B of compound of formula (II) is shown in FIG.

[0141] Figure 5 This is the TGA spectrum of the crystal form B of compound of formula (II).

[0142] Figure 6 The XRPD spectrum of the crystal form C of the compound of formula (I) is obtained using Cu-Kα radiation.

[0143] Figure 7 The DSC and TGA spectra of the crystal form C of compound of formula (I) are shown.

[0144] Figure 8 This is the XRPD spectrum of the crystal form D of the compound of formula (III-1) using Cu-Kα radiation.

[0145] Figure 9 The DSC and TGA spectra of the crystal form D of compound (III-1) are shown.

[0146] Figure 10 This is the XRPD spectrum of the crystal form E of the compound of formula (IV-1) using Cu-Kα radiation.

[0147] Figure 11 These are the DSC and TGA spectra of the crystalline form E of compound (IV-1).

[0148] Figure 12 The XRPD spectrum of the crystalline form F of the compound of formula (V) is obtained using Cu-Kα radiation.

[0149] Figure 13 The DSC and TGA spectra of the crystalline form F of the compound of formula (V) are shown.

[0150] Figure 14 It is a three-dimensional structural ellipsoid diagram of the crystal form B of compound of formula (II).

[0151] Figure 15 This is the nuclear magnetic spectrum of the crystal form D of compound (III-1).

[0152] Figure 16 This is the nuclear magnetic spectrum of the crystal form E of compound (IV-1).

[0153] Figure 17 The tear secretion volume of the scopolamine dry eye model in mice.

[0154] Figure 18 This is corneal fluorescence staining of the mouse scopolamine dry eye model.

[0155] Figure 19 The tear secretion volume of rat hyperosmolar dry eye model animals.

[0156] Figure 20 This is corneal fluorescence staining of rat hyperosmotic dry eye model animals.

[0157] Figure 21 Tear film breakup time in rat hyperosmolar dry eye model. DETAILED DESCRIPTION

[0158] In order to better understand the content of the present invention, further description is given below in conjunction with specific embodiments, but the specific implementation methods are not intended to limit the content of the present invention.

[0159] Example 1: Preparation of Crystalline Form B of Compound (II)

[0160]

[0161] Synthesis route:

[0162]

[0163] Step 1: Preparation of compound 2

[0164] Compound 1 (30 g, 130.4 mmol, 1 eq), bis-naphthalene borate (66.23 g, 260.80 mmol, 2 eq), [1,1-bis(diphenylphosphino)ferrocene]dichloropalladium(II) dichloromethane adduct (5.32 g, 6.52 mmol, 0.1 eq), and potassium acetate (25.60 g, 260.80 mmol, 2 eq) were added to toluene (500 mL). The atmosphere was purged with nitrogen three times, and the reaction solution was stirred at 110°C for 15 hours. After completion of the reaction, the reaction solution was filtered through a pad of celite, the filtrate was concentrated, and the residue was purified by column chromatography (petroleum ether:ethyl acetate = 0 to 100:6) to obtain compound 2.

[0165] 1 H NMR (400MHz, CDCl3) δ7.84 (d, J=8.0Hz, 1H), 7.06 (s, 1H), 7.04 (d, J=8.0Hz, 1H), 5.65 (brs, 2H), 3.87 (s, 3H), 1.35 (s, 12H).

[0166] Step 2: Preparation of compound 4

[0167] Compound 3 (100 g, 460.79 mmol, 1 eq) was dissolved in anhydrous ethanol (1 L), and concentrated sulfuric acid (225.97 g, 2.30 mol, 122.81 mL, 5 eq) and anhydrous sodium sulfate Na2SO4 (65.45 g, 460.79 mmol, 46.75 mL, 1 eq) were added. The reaction solution was stirred at 85°C for 48 hours. After the reaction was completed, the reaction solution was cooled to room temperature. Saturated sodium bicarbonate aqueous solution (1 L) was added dropwise to the reaction solution. A large amount of solid was formed, which was filtered and the filter cake was washed with water (500 mL). The resulting solid was vacuum dried to obtain compound 4.

[0168] 1 H NMR (400MHz, CDCl3) δ 8.10 (d, J=1.8Hz, 1H), 7.26 (s, 1H), 4.47 (q, J=7.1Hz, 2H), 1.46 (t, J=7.2Hz, 3H).

[0169] Step 3: Preparation of compound 5

[0170] Compound 4 (70.00 g, 285.63 mmol, 1 eq) was dissolved in tetrahydrofuran (1 L) and cooled to -78°C under nitrogen. Methyllithium (1.6 M, 892.59 mL, 5 eq) was slowly added dropwise to the reaction solution, and the reaction solution was stirred at -78°C for 3 hours. After the reaction was completed, water (100 mL) was slowly added dropwise to quench the reaction. The solution was warmed to room temperature and diluted with saturated aqueous ammonium chloride (500 mL). The solution was extracted with ethyl acetate (500 mL*3). The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to obtain a crude product. The crude product was slurried with n-heptane (500 mL), filtered, and dried to obtain compound 5.

[0171] 1 H NMR (400MHz, CDCl3) δ7.86 (d, J=1.9Hz, 1H), 6.98 (d, J=1.9Hz, 1H), 4.57 (brs, 2H), 1.57 (s, 6H).

[0172] Step 4: Preparation of compound 6

[0173] Compound 5 (10 g, 43.27 mmol, 1 eq), compound 2 (23.98 g, 86.55 mmol, 2 eq), [1,1-bis(diphenylphosphino)ferrocene]dichloropalladium(II) dichloromethane adduct (1.77 g, 2.16 mmol, 0.05 eq), and cesium carbonate (28.20 g, 86.55 mmol, 2 eq) were added to dioxane (300 mL) and water (75 mL). The atmosphere was purged with nitrogen three times, and the reaction solution was stirred at 80°C for 5 hours. After completion of the reaction, the reaction solution was concentrated, and the residue was purified by column chromatography (petroleum ether:tetrahydrofuran = 0 to 100:40) to obtain crude compound 6. This crude product was heated to 80°C with tetrahydrofuran (4 mL / g), cooled, recrystallized, stirred at 25°C for 15 hours, filtered, and the filter cake dried to obtain compound 6.

[0174] 1 H NMR (400MHz, DMSO-d6) δ7.94 (d, J=2.0Hz, 1H), 7.77 (d, J=8.0Hz, 1H), 7.17 (d, J=2.0Hz, 1H), 6. 99 (d, J=1.6Hz, 1H), 7.77-7.75 (111, 3H), 5.69 (s, 2H), 5.50 (s, 1H), 3.81 (s, 3H), 1.52 (s, 6H).

[0175] Step 5: Preparation of the compound of formula (1)

[0176] Compound 6 (8.78 g, 29.14 mmol, 1 eq) was dissolved in tetrahydrofuran (80 mL) and cooled to 0°C under nitrogen. Methylmagnesium bromide (3 M, 97.12 mL, 10 eq) was added dropwise to the reaction solution, and the mixture was stirred at 0°C for 1 hour. After completion of the reaction, saturated aqueous ammonium chloride (400 mL) was slowly added to quench the reaction, and the mixture was extracted with ethyl acetate (400 mL x 2). The organic phase was concentrated under reduced pressure, and the crude residue was purified with dichloromethane (3 mL / g) at 25°C and filtered to obtain the product.

[0177] 1 H NMR (400MHz, DMSO-d6) δ7.88 (d, J=1.8Hz, 1H), 7.15-7.01 (m, 2H), 6.82 (d, J=1.6Hz, 1H), 6.69 (dd, J=1.5, 8.0Hz, 1H), 5.59 (br s, 2H), 5.51 (br s, 2H), 5.44 (s, 1H), 5.23 (s, 1H), 1.51 (d, J=3.6Hz, 12H).

[0178] Weigh 11.9 g of the above product into a round-bottom flask, add 150 mL of methyl tert-butyl methyl ether, stir at 50°C for 12 hours, then cool to 25°C and stir for 4 hours. Filter and dry to obtain a solid. XRPD analysis confirmed Form A.

[0179] Approximately 50 mg of Form A was weighed and added to a 2.0 mL glass vial. An appropriate amount of solvent or solvent mixture was added to form a suspension. After adding a magnet, the sample was stirred on a magnetic stirrer (25°C / 50°C) for one week. After centrifugation, the resulting solid sample was placed in a 40°C vacuum drying oven and dried overnight to obtain Form B of Compound (II).

[0180] serial number solvent Volume ratio Crystal form 1 water / B crystal form 2 Acetonitrile / water 3 / 1 B crystal form 3 Methanol / water 3 / 1 B crystal form 4 Tetrahydrofuran / water 3 / 1 B crystal form 5 Ethanol / water 3 / 1 B crystal form 6 Isopropyl alcohol / water 3 / 1 B crystal form

[0181] Example 2: Structural confirmation of the compound of formula (II)

[0182]

[0183] Single crystal growth process: Dissolve approximately 5 mg of Form B of Compound (II) in 3 mL of dichloromethane / methanol (2:1) at room temperature. Place the sample solution in a 4 mL semi-sealed sample vial and slowly evaporate at room temperature. Colorless, transparent crystals were obtained on the fifth day. The crystal size for the diffraction experiment was: 0.20 × 0.18 × 0.15 mm. 3 .

[0184] Single crystal diffraction instrument: Bruker D8 venture

[0185] The measured compound information is: molecular formula C 17 H 25 N3O3, crystal system Monoclinic, space group C2 / c, unit cell parameters are β=90.01°, α=γ=90°, volume Figure 14 The ellipsoid diagram of the three-dimensional structure of the compound of formula (II) crystal form B. The crystal form of the single crystal was also measured and confirmed to be crystal form B.

[0186] Example 3: Preparation of various crystal forms

[0187]

[0188] Approximately 50 mg of Form A was weighed and added to a 2.0 mL glass vial. An appropriate amount of methanol or ethanol was added to form a suspension. After adding a magnet, the sample was stirred on a magnetic stirrer (50°C) for one week. After centrifugation, the resulting solid sample was placed in a 40°C vacuum drying oven and dried overnight to obtain Form C of Compound of Formula (I).

[0189]

[0190] Weigh approximately 50 mg of Form A into a 2.0 mL glass vial and add an appropriate amount of ethyl acetate to form a suspension. After adding a magnet, place the sample on a magnetic heating stirrer (25°C / 50°C) and stir for one week. After centrifugation, place the resulting solid sample in a 40°C vacuum drying oven and dry overnight to obtain Form D of Compound (III-1). NMR data are shown in Figure 15 .

[0191]

[0192] Weigh approximately 50 mg of Form A into a 2.0 mL glass vial and add an appropriate amount of acetonitrile to form a suspension. After adding a magnet, the sample was placed on a magnetic heating stirrer (25°C / 50°C) and stirred for one week. After centrifugation, the resulting solid sample was placed in a 40°C vacuum drying oven and dried overnight to obtain Form E of Compound (IV-1). NMR data are shown in Figure 16 .

[0193]

[0194] Approximately 50 mg of Form A was weighed and added to a 2.0 mL glass vial. An appropriate amount of dichloromethane was added to form a suspension. After adding a magnet, the sample was stirred on a magnetic stirrer (25°C) for one week. After centrifugation, the resulting solid sample was placed in a 40°C vacuum drying oven and dried overnight to obtain Form F of Compound (V).

[0195] Example 4: Preliminary stability test of the crystal form of compound B of formula (ID

[0196] Approximately 50 mg of Form B of Compound (II) was accurately weighed and placed in a clean, dry glass vial. The sample was spread into a thin layer and used as the test sample. The sample was then placed under the influencing factor test conditions (40°C, 25°C / 92.5% RH, light, and light control) and accelerated conditions (40°C / 75% RH and 60°C / 75% RH). The sample was fully exposed. Samples were analyzed after 5 and 10 days for the 40°C, 25°C / 92.5% RH, light, and light control conditions, and after 1, 2, and 3 months for the accelerated conditions.

[0197]

[0198] Experimental conclusion: The crystals of compound B of formula (II) were stable when placed under the influencing factor test conditions (40°C, 25°C / 92.5% RH, light, light control) for 10 days, 40°C for 1 month, and long-term accelerated conditions (40°C / 75% RH and 60°C / 75% RH) for 3 months.

[0199] Example 5: 25°C water activity competition experiment between the crystal form B of the compound of formula (ID) and the crystal form C of the compound of formula (I)

[0200] Weigh approximately 15 mg of the crystalline form of Compound (I) C and add an appropriate amount of the solvent system to form a saturated solution at room temperature. If the solution is clear, continue adding the compound until a saturated solution is obtained. Filter the solution through a 0.45 μm nylon needle filter into a liquid phase vial. Then, add approximately equal amounts of the crystalline form of Compound (II) B and the crystalline form of Compound (I) C to the liquid phase vial to form a suspension. After adding a magnet, the suspension sample is placed on a thermomixer (25°C, 700 rpm) and shaken.

[0201]

[0202] The results showed that when the water activity of Compound B (II) and Compound C (I) was suspended and slurried for 5 days at a water activity of 0, the resulting product was Compound C (I). However, as the water activity increased, after 5 days of suspension and slurrying, all the products obtained were Compound B (II). The products obtained with water activities of 0.9 and 1.0 were dried and characterized, and the two were almost identical. Therefore, Compound B (II) is more stable than Compound C (I).

[0203] Biological test data

[0204] Example 1: Experimental study on the effect of the crystal form eye drops of compound B of formula (II) on the dry eye model in mice

[0205] Purpose of the experiment:

[0206] The dry eye model of C57BL / 6 mice was induced by subcutaneous injection of scopolamine hydrobromide solution into the lower limbs to investigate the therapeutic effect of the crystal form eye drops of compound B of formula (II) on the model.

[0207] Experimental process:

[0208] Animals were randomly and evenly divided into five groups based on tear secretion: a negative control group (normal saline), a model control group (vehicle), and groups treated with low (1 mg / mL), medium (2.5 mg / mL), and high (5 mg / mL) concentrations of the crystalline form of Compound B of Formula (II). Each group consisted of eight female animals. The day of grouping was designated D0. Following grouping, the animals were modeled, eye drops were administered, corneal fluorescence staining was scored, and tear secretion was measured according to the experimental protocol.

[0209] Experimental results:

[0210] The crystal form solutions of compound B of formula (II) (1 mg / mL, 2.5 mg / mL, 5 mg / mL) all had a good therapeutic effect on the dry eye model of mice induced by scopolamine hydrobromide solution, mainly improving the tear secretion and corneal damage of the dry eye model mice. Figure 17 ) and corneal fluorescence staining (Table 8, Figure 18 ) scoring results, the therapeutic effect of the crystalline form eye drops of compound B of formula (II) is optimal at a concentration of 5 mg / mL.

[0211] Table 7 Effect of the crystal solution of compound B of formula (II) on the tear secretion of dry eye model mice

[0212]

[0213] Note: Compared with D0, #P<0.05; ##P<0.01; compared with G1 group, *P<0.05; **P<0.01; compared with G2 group, &P<0.05; &&P<0.01.

[0214] Table 8 Effect of the crystal form solution of compound B of formula (II) on corneal fluorescence staining scores in dry eye model mice

[0215]

[0216] Note: Compared with D0, #P<0.05; ##P<0.01; compared with G1 group, *P<0.05; **P<0.01; compared with G2 group, &P<0.05; &&P<0.01.

[0217] Experimental conclusion:

[0218] The crystal form eye drops of compound B of formula (II) have a good therapeutic effect on the dry eye model of mice induced by scopolamine hydrobromide solution, and mainly improve the tear secretion and corneal damage of the dry eye model mice.

[0219] Example 2: Experimental study on the effect of the crystal form eye drops of compound B of formula (II) on the rat hyperosmotic dry eye model

[0220] Twenty female SD rats that passed the adaptive observation were selected for binocular corneal fluorescence staining scoring and tear secretion measurement. Animals with abnormal corneal fluorescence staining and large differences in tear secretion between the two eyes were eliminated.

[0221] Use a pipette to draw 20 μL of sodium chloride solution (osmotic pressure of 500mOsmol / L) and instill it into the conjunctival sac of both eyes of the animal, 5 times / day, 20 μL / time, each time interval is about 2 hours, for 28 consecutive days, and the animal's eyelids are passively closed for about 90 seconds after instillation. On the 14th day of the modeling period, the animals were scored for corneal fluorescence staining and tear secretion was measured. The data were compared with the baseline data, and animals with significant differences in tear secretion between the two eyes were selected for grouping. According to the mean tear secretion volume of the two eyes of the animals, the animals were randomly and balancedly divided into 4 groups, namely the model control group, the low and high concentration groups of the crystal form of compound B of formula (II) (1 mg / mL and 5 mg / mL), each group had 4 animals and 8 eyes, and the day of grouping was recorded as D0.

[0222] Animals in all groups were administered eye drops on Day 1 (D1) at a rate of 10 μL / eye, four times daily, with dosing intervals of approximately 3 hours, for a total of 14 days. Animals were weighed weekly during the dosing period. Tear production was measured approximately 30 minutes after the second dose on Days 7 and 14, respectively. Corneal fluorescein staining was performed approximately 30 minutes after the third dose on Days 7 and 14. After completion of these measurements on Day 14, animals were euthanized by carbon dioxide inhalation.

[0223] Experimental results: see Tables 9, 10 and Figure 19 、 20 and 21.

[0224] Table 9 Effect of the crystal solution of compound B of formula (II) on the tear secretion of dry eye model rats

[0225]

[0226] Note: Compared with group G1, *P<0.05; **P<0.01.

[0227] Table 10 Effect of the crystal form solution of compound B of formula (II) on corneal fluorescence staining scores in dry eye model mice

[0228]

[0229]

[0230] Note: Compared with group G1, *P<0.05; **P<0.01.

[0231] Experimental conclusion: The formula (ID) compound B crystalline form eye drops have a good therapeutic effect on the rat dry eye model induced by hypertonic sodium chloride solution, which mainly improves the tear secretion volume, corneal damage and tear film breakup time of the dry eye model rats.

Claims

1. Crystal form B of the compound of formula (II), characterized in that Its X-ray powder diffraction pattern has characteristic diffraction peaks at the following 2θ angles: 7.083±0.200°，8.231±0.200°，11.294±0.200°，12.068±0.200°，14.091±0.200°，16.505±0.200°，17.058±0.200°，18.955±0.200°，21.712±0.200°，24.242±0.200°，25.678±0.200°，30.869±0.200° 2. The crystal form B according to claim 1, wherein the X-ray powder diffraction pattern has characteristic diffraction peaks at the following 2θ angles: 7.083°, 8.231°, 11.294°, 12.068°, 14.091°, 14.806°, 16.505°, 17.058°, 18.202°, ​​18.955°, 21.712°, 22.182°, 23.903°, 24.242°, 24.653°, 25.350°, 25.678°, 26.270°, 27.001°, 27.658°, 29.052°, 29.721°, 30.869°, 34.644°, and 35.009°.

3. The Form B according to claim 2, has an XRPD pattern substantially as shown in FIG3 .

4. The crystal form B according to any one of claims 1 to 3, wherein the differential scanning calorimetry curve thereof has an endothermic peak starting point at 101.7±3.0°C and 158.7±3.0°C, respectively.

5. The crystal form B according to claim 4, whose DSC spectrum is substantially as shown in Figure 4.

6. The crystal form B according to any one of claims 1 to 3, wherein the thermogravimetric analysis curve thereof shows a weight loss of 5.477% at 120.00°C ± 3.0°C.

7. The crystal form B according to claim 6, whose TGA spectrum is substantially as shown in Figure 5.

8. Use of the crystal form B according to any one of claims 1 to 7 in the preparation of a medicament for treating dry eye.

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