Salt forms and crystal forms thereof as Cdc7 inhibitors

Abstract

Disclosed are a salt and a crystal form thereof as a Cdc7 inhibitor, specifically a compound of formula (II) and a crystal form thereof, as well as a preparation method thereof and an application thereof in preparing a drug for preventing or treating tumors.

Description

[0001] Citation of Related Applications

[0002] This disclosure claims priority and benefits of Chinese Patent Application No. 202011383418.7 filed with the State Intellectual Property Office of China on November 30, 2020, and the contents of the patent application disclosure are incorporated herein by reference in their entirety. Technical Field

[0003] The present disclosure belongs to the field of pharmaceutical chemistry, and specifically relates to a salt form and a crystal form thereof as a Cdc7 inhibitor, more specifically to a compound of formula (II) and a crystal form thereof, and also to a method for preparing the salt form and the crystal form and their use in preparing a drug for preventing or treating tumors. Background Art

[0004] Cdc7 is a serine / threonine kinase that binds to ASK (activator of S phase kinase, also known as DBF4) in the cell nucleus. On the one hand, it phosphorylates the minichromosome maintenance protein (MCM protein), an important element of DNA replication initiation, to activate MCM to promote the formation of replication initiation complex; on the other hand, it also participates in the ATR / Chk1 pathway as an important regulatory factor of the S phase checkpoint of the cell cycle to promote cell cycle arrest and DNA repair. Cdc7 is abnormally highly expressed in many human tumor cells. This abnormally high expression shows a high correlation with tumor proliferation, metastasis and resistance to chemotherapy drugs. Therefore, Cdc7 has become an important marker and target for current tumor research. Summary of the invention

[0005] In one aspect, the present disclosure provides a compound of formula (II)

[0006]

[0007] In another aspect, the present disclosure also provides a crystalline form of the compound of formula (II).

[0008] In some embodiments of the present disclosure, the crystalline form of the compound of formula (II) is selected from:

[0009] The crystalline form A of the compound of formula (II), whose X-ray powder diffraction pattern comprises characteristic peaks at 2θ values ​​of 11.46±0.20°, 24.03±0.20° and 25.16±0.20°;

[0010] The crystalline form B of the compound of formula (II), whose X-ray powder diffraction pattern comprises characteristic peaks at 2θ values ​​of 5.86±0.20°, 17.64±0.20° and 24.89±0.20°;

[0011] The crystalline form C of the compound of formula (II) has an X-ray powder diffraction pattern comprising characteristic peaks at 2θ values ​​of 7.40±0.20°, 11.21±0.20° and 22.18±0.20°.

[0012] On the other hand, the present disclosure also provides Form A of the compound of formula (II), whose X-ray powder diffraction pattern comprises characteristic peaks at 2θ values ​​of 11.46±0.20°, 24.03±0.20° and 25.16±0.20°.

[0013] In some embodiments of the present disclosure, the above-mentioned crystal form A has an X-ray powder diffraction pattern using Cu Kα radiation having diffraction peaks at the following 2θ values: 11.46±0.20°, 24.03±0.20° and 25.16±0.20°.

[0014] In some embodiments of the present disclosure, the above-mentioned Form A has an X-ray powder diffraction pattern comprising characteristic peaks at 2θ values ​​of 11.46±0.20°, 12.06±0.20°, 16.96±0.20°, 17.60±0.20°, 18.48±0.20°, 19.55±0.20°, 24.03±0.20° and 25.16±0.20°.

[0015] In some embodiments of the present disclosure, the above-mentioned crystalline form A has an X-ray powder diffraction pattern comprising 2θ values ​​of 6.46°±0.20°, 9.36°±0.20°, 11.46°±0.20°, 12.06°±0.20°, 12.39°±0.20°, 12.89°±0.20°, 13.37°±0.20°, 13.87°±0.20°, 14.09 °±0.20°、16.10°±0.20°、16.96°±0.20°、17.19°±0.20°、17.60°±0.20°、18.48°±0.20°、18.75°±0.20°、19.37°±0.20°、19.55°±0.20°、21.21°±0.20°、21.65°±0.20°、22.36 ° ± 0.20°, 23.06° ± 0.20°, 23.42° ± 0.20°, 23.74° ± 0.20°, 24.03° ± 0.20°, 25.16° ± 0.20°, 25.47° ± 0.20°, 26.59° ± 0.20°, 27.32° ± 0.20°, 28.11° ± 0.20°, 28.77° ± 0.20°, 29.73 The characteristic peaks are at 31.81°±0.20°, 33.09°±0.20°, 33.80°±0.20°, 34.19°±0.20°, 35.49°±0.20°, 35.99°±0.20°, 37.66°±0.20°, 38.14°±0.20°, 38.77°±0.20° and 39.38°±0.20°.

[0016] In some embodiments of the present disclosure, the above-mentioned Form A has an X-ray powder diffraction pattern comprising 2θ values ​​of 6.46°, 9.36°, 11.46°, 12.06°, 12.39°, 12.89°, 13.37°, 13.87°, 14.09°, 16.10°, 16.96°, 17.19°, 17.60°, 18.48°, 18.75°, 19.37°, 19.55°, 21.21°, 21. Characteristic peaks at 65°, 22.36°, 23.06°, 23.42°, 23.74°, 24.03°, 25.16°, 25.47°, 26.59°, 27.32°, 28.11°, 28.77°, 29.73°, 31.81°, 33.09°, 33.80°, 34.19°, 35.49°, 35.99°, 37.66°, 38.14°, 38.77° and 39.38°.

[0017] In some embodiments of the present disclosure, the above-mentioned crystal form A has an X-ray powder diffraction pattern as shown in Figure 1 shown.

[0018] In some embodiments of the present disclosure, the X-ray powder diffraction pattern of Form A of the compound of formula (II) is shown in Table 1.

[0019] Table 1 XRPD diffraction pattern analysis data of the crystalline form A of the compound of formula (II)

[0020]

[0021] In some embodiments of the present disclosure, the DSC curve of the above-mentioned crystal form A has a starting point of an endothermic peak at 230.7°C±3°C.

[0022] In some embodiments of the present disclosure, the above-mentioned crystal form A has a DSC curve as shown in Figure 4 shown.

[0023] In some embodiments of the present disclosure, the TGA curve of the above-mentioned Form A has a weight loss of 0.94±0.2% at 200.0°C±3°C.

[0024] In some embodiments of the present disclosure, the TGA curve of the above-mentioned crystal form A is as follows Figure 5 shown.

[0025] In some embodiments of the present disclosure, the above-mentioned crystal form A has an infrared spectrum at 3033.88 cm -1 ±5cm -1 、3225.69cm -1 ±5cm -1 、3087.88cm -1 ±5cm -1 、2945.00cm -1 ±5cm- 1 、2925.62cm- 1 ±5cm -1 、2837.85cm -1 ±5cm -1 、1680.20cm- 1 ±5cm -1 、1662.75cm -1 ±5cm -1 、1582.38cm -1 ±5cm -1 、1551.77cm -1 ±5cm -1 、1475.47cm -1 ±5cm -1 and 1438.43cm -1 ±5cm- 1 Characteristic absorption peak at .

[0026] In some embodiments of the present disclosure, the above-mentioned crystal form A has an infrared absorption spectrum as shown in Figure 6 shown.

[0027] In some embodiments of the present disclosure, the above-mentioned crystal form A has characteristic absorption peaks at 259.0nm±3nm, 214.4nm±3nm and 322.6nm±3nm in its ultraviolet absorption spectrum.

[0028] In some embodiments of the present disclosure, the above-mentioned crystal form A has an ultraviolet absorption spectrum as shown in Fig.11 shown.

[0029] On the other hand, the present disclosure also provides Form B of the compound of formula (II), whose X-ray powder diffraction pattern comprises characteristic peaks at 2θ values ​​of 5.86±0.20°, 17.64±0.20° and 24.89±0.20°.

[0030] In some embodiments of the present disclosure, the above-mentioned Form B has an X-ray powder diffraction pattern using Cu Kα radiation having diffraction peaks at the following 2θ values: 5.86±0.20°, 17.64±0.20° and 24.89±0.20°.

[0031] In some embodiments of the present disclosure, the above-mentioned Form B has an X-ray powder diffraction pattern comprising characteristic peaks at 2θ values ​​of 5.86±0.20°, 10.17±0.20°, 11.73±0.20°, 15.83±0.20°, 17.64±0.20°, 23.59±0.20°, 24.89±0.20° and 25.80±0.20°.

[0032] In some embodiments of the present disclosure, the above-mentioned Form B has an X-ray powder diffraction pattern comprising 2θ values ​​of 5.86°±0.20°, 10.17°±0.20°, 11.73°±0.20°, 12.57°±0.20°, 14.15°±0.20°, 14.40°±0.20°, 15.23°±0.20°, 15.83°±0.20°, 16.26 ° ± 0.20 °, 16.71 ° ± 0.20 °, 17.64 ° ± 0.20 °, 18.04 ° ± 0.20 °, 18.73 ° ± 0.20 °, 19.99 ° ± 0.20 °, 20.57 ° ± 0.20 °, 21.08 ° ± 0.20 °, 23.59 ° ± 0.20 °, 24.36 ° ± 0.20 °, 24.89 ° ± 0.20 °, 25.41 ° ± 0.20°, 25.80° ± 0.20°, 27.20° ± 0.20°, 27.90° ± 0.20°, 28.90° ± 0.20°, 29.39° ± 0.20°, 29.70° ± 0.20°, 30.52° ± 0.20°, 30.81° ± 0.20°, 31.99° ± 0.20°, 34.29° ± 0.20°, 35.83 The characteristic peaks are at 39.64°±0.20°, 28.90°±0.20°, 29.39°±0.20°, 29.70°±0.20°, 30.52°±0.20°, 30.81°±0.20°, 31.99°±0.20°, 34.29°±0.20°, 35.83°±0.20° and 39.64°±0.20°.

[0033] In some embodiments of the present disclosure, the above-mentioned Form B has an X-ray powder diffraction pattern comprising 2θ values ​​of 5.86°, 10.17°, 11.73°, 12.57°, 14.15°, 14.40°, 15.23°, 15.83°, 16.26°, 16.71°, 17.64°, 18.04°, 18.73°, 19.99°, 20.57°, 21.08°, Characteristic peaks at 23.59°, 24.36°, 24.89°, 25.41°, 25.80°, 27.20°, 27.90°, 28.90°, 29.39°, 29.70°, 30.52°, 30.81°, 31.99°, 34.29°, 35.83°, 39.64°, 28.90°, 29.39°, 29.70°, 30.52°, 30.81°, 31.99°, 34.29°, 35.83° and 39.64°.

[0034] In some embodiments of the present disclosure, the above-mentioned crystal form B has an X-ray powder diffraction pattern as shown in Figure 2 shown.

[0035] In some embodiments of the present disclosure, the X-ray powder diffraction pattern of Form B of the compound of formula (II) is shown in Table 2.

[0036] Table 2 XRPD spectrum analysis data of Form B of the compound of formula (II)

[0037]

[0038] In some embodiments of the present disclosure, the DSC curve of the above-mentioned Form B has onset points of endothermic peaks at 65.1°C±3°C, 113.7°C±3°C, 208.8°C±3°C and 221.1°C±3°C.

[0039] In some embodiments of the present disclosure, the above-mentioned crystal form B has a DSC curve as shown in Figure 7 shown.

[0040] In some embodiments of the present disclosure, the TGA curve of the above-mentioned Form B has a weight loss of 3.58±0.2% at 150.0°C±3°C.

[0041] In some embodiments of the present disclosure, the TGA curve of the above-mentioned crystal form B is as follows Figure 8 shown.

[0042] In another aspect, the present disclosure provides Form C of the compound of formula (II), whose X-ray powder diffraction pattern comprises characteristic peaks at 2θ values ​​of 7.40±0.20°, 11.21±0.20° and 22.18±0.20°.

[0043] In some embodiments of the present disclosure, the above-mentioned crystal form C has an X-ray powder diffraction pattern using Cu Kα radiation having diffraction peaks at the following 2θ values: 7.40±0.20°, 11.21±0.20° and 22.18±0.20°.

[0044] In some embodiments of the present disclosure, the above-mentioned Form C has an X-ray powder diffraction pattern comprising characteristic peaks at 2θ values ​​of 7.40±0.20°, 11.21±0.20°, 13.95±0.20°, 15.01±0.20°, 15.72±0.20°, 20.61±0.20°, 22.18±0.20° and 23.82±0.20°.

[0045] In some embodiments of the present disclosure, the above-mentioned crystal form C has an X-ray powder diffraction pattern comprising 2θ values ​​of 7.40°±0.20°, 10.50°±0.20°, 11.21°±0.20°, 11.81°±0.20°, 13.05°±0.20°, 13.95°±0.20°, 15.01°±0.20°, 15.72°±0.20°, 16.28°±0.20°, 17.64°±0.20°, 18.35°±0.20°, 18.73°±0.20°, 19.53 The characteristic peaks are at 20.1°±0.20°, 20.14°±0.20°, 20.61°±0.20°, 22.18°±0.20°, 22.51°±0.20°, 23.82°±0.20°, 24.37°±0.20°, 25.49°±0.20°, 26.36°±0.20°, 27.19°±0.20°, 28.93°±0.20°, 30.70°±0.20°, 31.60°±0.20°, 32.50°±0.20° and 34.33°±0.20°.

[0046] In some embodiments of the present disclosure, the above-mentioned Form C has an X-ray powder diffraction pattern comprising characteristic peaks at 2θ values ​​of 7.40°, 10.50°, 11.21°, 11.81°, 13.05°, 13.95°, 15.01°, 15.72°, 16.28°, 17.64°, 18.35°, 18.73°, 19.53°, 20.14°, 20.61°, 22.18°, 22.51°, 23.82°, 24.37°, 25.49°, 26.36°, 27.19°, 28.93°, 30.70°, 31.60°, 32.50° and 34.33°.

[0047] In some embodiments of the present disclosure, the above-mentioned crystal form C has an X-ray powder diffraction pattern as shown in Figure 3 shown.

[0048] In some embodiments of the present disclosure, the X-ray powder diffraction pattern of Form C of the compound of formula (II) is shown in Table 3.

[0049] Table 3 XRPD spectrum analysis data of the crystalline form C of the compound of formula (II)

[0050]

[0051] In some embodiments of the present disclosure, the DSC curve of the above-mentioned crystal form C has a starting point of an endothermic peak at 219.9°C±3°C.

[0052] In some embodiments of the present disclosure, the above-mentioned crystal form C has a DSC curve as shown in Fig. 9 shown.

[0053] In some embodiments of the present disclosure, the above-mentioned crystal form C, wherein the TGA curve of the crystal form has a weight loss of 0.72±0.2% at 170.0°C±3°C.

[0054] In some embodiments of the present disclosure, the TGA curve of the above-mentioned crystal form C is as follows Fig.10 shown.

[0055] In some embodiments of the present disclosure, the X-ray powder diffraction pattern is an X-ray powder diffraction pattern of Cu Kα radiation.

[0056] In the present disclosure, the compound of formula (I) has the structure shown below:

[0057]

[0058] In another aspect, the present disclosure provides a method for preparing a crystalline form of the compound of formula (II), the method comprising the following steps:

[0059] (1) mixing the compound of formula (I) shown below with methanol;

[0060] (2) adding maleic acid to the mixture of step (1);

[0061] (3) filtering and drying to obtain the above-mentioned Form A or Form C; and

[0062] (4) mixing the above-mentioned crystal form A with acetonitrile and water, and evaporating the solvent to dryness after the solid is dissolved to obtain the above-mentioned crystal form B,

[0063]

[0064] In some embodiments of the present disclosure, the method for preparing the crystalline form of the compound of formula (II) is the method for preparing the crystalline form A, which comprises the following steps:

[0065] (1) mixing the compound of formula (I) with methanol at 20-30° C.;

[0066] (2) After cooling to 5° C. under stirring, maleic acid was added to the mixture of step (1), and stirred at 5-13° C. for 6 hours;

[0067] (3) Filter and dry to obtain the above-mentioned crystal form A.

[0068] In some embodiments of the present disclosure, the method for preparing the crystalline form of the compound of formula (II) is the method for preparing the crystalline form B, which comprises the following steps: mixing the crystalline form A with acetonitrile and water, and after the solid is dissolved, evaporating the solvent to dryness at 40°C to obtain the crystalline form B.

[0069] In some embodiments of the present disclosure, the method for preparing the crystalline form of the compound of formula (II) is the method for preparing the crystalline form C, which comprises the following steps:

[0070] (1) mixing the compound of formula (I) with methanol at 20-30° C.;

[0071] (2) adding maleic acid to the mixture of step (1) under stirring at 75° C. and stirring at 75° C. for 0.5 hour, stirring at 40° C. for 1 hour, lowering the temperature to 25° C. and continuing stirring for 2 hours;

[0072] (3) Filter and dry to obtain the above-mentioned crystal form C.

[0073] In another aspect, the present disclosure provides a crystalline composition comprising Form A of the compound of formula (II), Form B of the compound of formula (II) or Form C of the compound of formula (II), wherein the Form A of the compound of formula (II), Form B of the compound of formula (II) or Form C of the compound of formula (II) accounts for more than 50% by weight of the crystalline composition, preferably more than 75%, more preferably more than 90%, and most preferably more than 95%. Each of the crystalline compositions may also contain a small amount of other crystalline or non-crystalline forms of the compound of formula (II).

[0074] In yet another aspect, the present disclosure provides a pharmaceutical composition comprising the above compound or the above crystal form, and optional pharmaceutically acceptable excipients.

[0075] In yet another aspect, the present disclosure also provides use of the above compound, the above crystal form, or the above pharmaceutical composition in the preparation of a Cdc7 inhibitor.

[0076] The present disclosure also provides use of the above compound, the above crystal form, the above crystalline composition, or the above pharmaceutical composition in preparing a drug for preventing or treating a disease mediated by Cdc7 kinase (eg, tumor).

[0077] The present disclosure also provides a method for preventing or treating a disease mediated by Cdc7 kinase, comprising administering a therapeutically effective amount of the above compound or the above crystal form or the above crystalline composition or the above pharmaceutical composition to a mammal, preferably a human, in need thereof.

[0078] The present disclosure also provides use of the above-mentioned compound, the above-mentioned crystal form, the above-mentioned crystalline composition, or the above-mentioned pharmaceutical composition in preventing or treating a disease mediated by Cdc7 kinase.

[0079] The present disclosure also provides the above compound or the above crystal form or the above crystalline composition or the above pharmaceutical composition for preventing or treating a disease mediated by Cdc7 kinase.

[0080] The present disclosure also provides the above-mentioned compound or the above-mentioned crystal form or the above-mentioned crystalline composition or the above-mentioned pharmaceutical composition for use as a Cdc7 inhibitor.

[0081] The present disclosure also provides the above-mentioned compound or the above-mentioned crystal form or the above-mentioned crystalline composition or the above-mentioned pharmaceutical composition for use as a drug for preventing or treating a disease mediated by Cdc7 kinase.

[0082] In some embodiments of the present disclosure, the above-mentioned disease mediated by Cdc7 kinase refers to a tumor, such as colorectal cancer or pancreatic cancer.

[0083] In some embodiments of the present disclosure, the above-mentioned Cdc7 inhibitor refers to a drug for treating tumors, such as a drug for treating colorectal cancer or pancreatic cancer.

[0084] Technical Effects

[0085] The preparation process of the disclosed compound and crystal form is simple, and the crystal form is relatively stable, less affected by heat and humidity, and convenient for preparation. The disclosed compound and its crystal form have good inhibitory activity of Cdc7, among which, it has a strong inhibitory effect on the enzyme CDC7 / DBF4, and has an inhibitory effect on the proliferation of colorectal cells COLO205 with high expression of Cdc7. It can be seen that the disclosed compound and crystal form are a potential new and effective drug for clinical treatment of tumors.

[0086] The crystal form disclosed in the present invention has advantages in terms of drug activity, hygroscopicity, stability, purity, and ease of preparation, so as to meet the needs of drug production, storage, transportation, and formulation.

[0087] Definition and Description

[0088] Unless otherwise specified, the following terms and phrases used herein are intended to have the following meanings. A particular phrase or term should not be considered to be uncertain or unclear in the absence of a special definition, but should be understood according to its ordinary meaning. When a trade name appears in this article, it is intended to refer to its corresponding commercial product or its active ingredient.

[0089] The word "comprise" or "comprises" and its English variations such as comprises or comprising or equivalents should be understood as an open, non-exclusive meaning, that is, "including but not limited to", meaning that in addition to the listed elements, components and steps, other unspecified elements, components and steps may also be included.

[0090] The compounds of the present invention may exist in specific geometric or stereoisomeric forms. The present invention contemplates all such compounds, including cis and trans isomers, (-)- and (+)-enantiomers, (R)- and (S)-enantiomers, diastereomers, (D)-isomers, (L)-isomers, and racemic mixtures and other mixtures thereof, such as enantiomerically or diastereomerically enriched mixtures, all of which are within the scope of the present invention. Additional asymmetric carbon atoms may be present in substituents such as alkyl. All of these isomers and their mixtures are included within the scope of the present invention.

[0091] Unless otherwise indicated, "(D)" or "(+)" indicates dextrorotatory, "(L)" or "(-)" indicates levorotatory, and "(DL)" or "(±)" indicates racemic.

[0092] Unless otherwise specified, the key is a solid wedge. and dotted wedge key To indicate the absolute configuration of a stereocenter, use a straight solid bond. and straight dashed key To indicate the relative configuration of a stereocenter, use a wavy line Indicates a wedge-shaped solid key or dotted wedge key Or use a wavy line Represents a straight solid bond and straight dashed key

[0093] The term "pharmaceutically acceptable" refers to those compounds, materials, compositions and / or dosage forms that are suitable for use in contact with human and animal tissues, without excessive toxicity, irritation, allergic reactions or other problems or complications, and are commensurate with a reasonable benefit / risk ratio, within the scope of sound medical judgment. "Pharmaceutically acceptable excipients" refer to inert substances that are administered together with the active ingredient and are conducive to the administration of the active ingredient, including but not limited to any glidant, sweetener, diluent, preservative, dye / colorant, flavor enhancer, surfactant, wetting agent, dispersant, disintegrant, suspending agent, stabilizer, isotonic agent, solvent or emulsifier approved by the State Food and Drug Administration for use in humans or animals (e.g., livestock). Non-limiting examples of the excipients include calcium carbonate, calcium phosphate, various sugars and various types of starch, cellulose derivatives, gelatin, vegetable oils and polyethylene glycol.

[0094] The term "pharmaceutical composition" refers to a mixture of one or more compounds of the present disclosure or their crystalline forms and pharmaceutically acceptable excipients. The purpose of a pharmaceutical composition is to facilitate administration of the compounds of the present disclosure to an organism.

[0095] The pharmaceutical composition of the present disclosure can be prepared by combining the compound of the present disclosure or its crystal form with suitable pharmaceutically acceptable excipients, for example, it can be formulated into solid, semi-solid, liquid or gaseous preparations, such as tablets, pills, capsules, powders, granules, ointments, emulsions, suspensions, suppositories, injections, inhalants, gels, microspheres and aerosols.

[0096] Typical routes of administration of the compounds of the present disclosure, or crystalline forms thereof, or pharmaceutical compositions thereof include, but are not limited to, oral, rectal, topical, inhalation, parenteral, sublingual, vaginal, intranasal, intraocular, intraperitoneal, intramuscular, subcutaneous, and intravenous administration.

[0097] The pharmaceutical composition of the present disclosure can be manufactured by methods well known in the art, such as conventional mixing methods, dissolving methods, granulating methods, making dragees, grinding methods, emulsifying methods, freeze-drying methods, and the like.

[0098] In some embodiments, the pharmaceutical composition is in oral form. For oral administration, the pharmaceutical composition can be formulated by mixing the active compound with pharmaceutically acceptable excipients well known in the art. These excipients enable the compounds of the present disclosure to be formulated into tablets, pills, lozenges, dragees, capsules, liquids, gels, slurries, suspensions, etc., for oral administration to patients.

[0099] The therapeutic dosage of the disclosed compound or its crystalline form may be determined, for example, based on the specific use of the treatment, the mode of administration of the compound, the health and condition of the patient, and the judgment of the prescribing physician. The ratio or concentration of the disclosed compound or its crystalline form in the pharmaceutical composition may not be fixed, depending on a variety of factors, including dosage, chemical properties (e.g., hydrophobicity), and route of administration.

[0100] The term "treatment" means administering a compound of the present disclosure or a crystalline form thereof or a pharmaceutical composition thereof to improve or eliminate a disease or one or more symptoms associated with the disease, and includes:

[0101] (i) inhibiting a disease or disease state, i.e. arresting its development;

[0102] (ii) ameliorating a disease or condition, even if causing regression of the disease or condition.

[0103] The term "prevention" means administering a compound of the present disclosure or a crystalline form thereof or a pharmaceutical composition thereof to prevent a disease or one or more symptoms associated with the disease, and includes preventing a disease or disease state from occurring in a mammal, particularly when such mammal is susceptible to the disease state but has not yet been diagnosed as having the disease state.

[0104] "Mammal" includes humans and domestic animals such as laboratory mammals and household pets (e.g., cats, dogs, pigs, sheep, cattle, goats, horses, rabbits), and non-domesticated mammals, such as wild mammals, etc.

[0105] With respect to a drug or pharmacologically active agent, the term "therapeutically effective amount" refers to a sufficient amount of the drug or agent that is non-toxic but can achieve the desired effect. The determination of the effective amount varies from person to person, depending on the age and general condition of the recipient, and also on the specific active substance. The appropriate effective amount in each case can be determined by a person skilled in the art based on routine experiments.

[0106] Herein, singular terms include plural referents and vice versa unless the context clearly dictates otherwise.

[0107] Herein, unless otherwise indicated, each parameter value (including 2θ values, reaction conditions) is deemed to be modified by the term "about" to reflect the measurement error that exists for each value, for example, there is an error of ±5% relative to a given value.

[0108] For the purpose of description and disclosure, all patents, patent applications and other identified publications are expressly incorporated herein by reference. These publications are provided only because their disclosure precedes the filing date of the present disclosure. All statements regarding the dates of these documents or the representations of the contents of these documents are based on the information available to the applicant and do not constitute any admission as to the correctness of the dates of these documents or the contents of these documents. Furthermore, any reference to these publications herein does not constitute an admission that the publications are part of the common general knowledge in the art in any country.

[0109] The intermediate compounds disclosed herein 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 substitutions well known to those skilled in the art. Preferred embodiments include but are not limited to the examples disclosed herein.

[0110] The structure of the compounds disclosed in the present invention can be confirmed by conventional methods known to those skilled in the art. If the present invention involves the absolute configuration of the compound, the absolute configuration can be confirmed by conventional technical means in the art. For example, single crystal X-ray diffraction (SXRD) is used to collect diffraction intensity data of the cultured single crystal using a Bruker D8venture diffractometer, the light source is CuKα radiation, and the scanning mode is φ / scan. After collecting relevant data, the direct method (Shelxs97) is further used to analyze the crystal structure, and the absolute configuration can be confirmed.

[0111] The chemical reactions of the specific embodiments of the present disclosure are carried out in a suitable solvent, which must be suitable for the chemical changes of the present disclosure and the reagents and materials required. In order to obtain the compounds of the present disclosure, it is sometimes necessary for those skilled in the art to modify or select the synthesis steps or reaction processes based on the existing embodiments.

[0112] The present disclosure will be described in detail below through examples, which are not intended to limit the present disclosure in any way.

[0113] All solvents used in this disclosure were commercially available and used without further purification.

[0114] The solvents used in the present disclosure are commercially available.

[0115] The present disclosure uses the following abbreviations: DMF-DMA: N,N-dimethylformamide dimethyl acetal; DHP: 3,4-dihydropyran; DMF: N,N-dimethylformamide; MeOH: methanol; THF: tetrahydrofuran.

[0116] It should be noted that in the X-ray powder diffraction pattern, the position of the peak or the relative intensity of the peak may vary due to factors such as the measuring instrument, measuring method / conditions, etc. For any specific crystal form, there may be an error in the position of the peak, and the measurement error of the 2θ value may be ±0.2°. Therefore, when determining each crystal form, this error should be taken into account, and the error is also within the scope of the present disclosure.

[0117] It should be noted that for the same crystal form, the position of the endothermic peak of DSC may be different due to factors such as the measuring instrument, measuring method / conditions, etc. For any specific crystal form, the position of the endothermic peak may have an error, which may be ±5°C or ±3°C. Therefore, when determining each crystal form, this error should be taken into account, and the error is also within the scope of the present disclosure.

[0118] It should be noted that for the same type of crystal, the position of the weight loss temperature and the weight loss value of TGA may be different due to factors such as the measuring instrument, measuring method / conditions, etc. For any specific crystal, there may be errors in the position of the weight loss temperature and the weight loss value, and the weight loss temperature error may be ±5°C, and the weight loss value error may be ±0.2%. Therefore, when determining each crystal, this error should be taken into account, and the error is also within the scope of the present disclosure.

[0119] It should be noted that in the infrared spectrum, the position of the peak may vary due to factors such as the measuring instrument, measuring method / conditions, etc. For any specific crystal form, there may be an error in the position of the peak. The measurement error of the peak position can be ±5cm -1Therefore, this error should be taken into account when determining each crystal form, and the error is also within the scope of the present disclosure.

[0120] It should be noted that in the ultraviolet absorption spectrum, the position of the peak may be different due to factors such as the measuring instrument, measuring method / conditions, etc. For any specific crystal form, there may be an error in the position of the peak, and the measurement error of the wavelength may be ±5nm or ±3nm. Therefore, when determining each crystal form, this error should be taken into account, and the error is also within the scope of the present disclosure.

[0121] Herein, unless otherwise specified, "room temperature" in the present disclosure refers to normal temperature, generally 20-30°C.

[0122] Instruments and analytical methods

[0123] 1.1 X-ray powder diffractometer (XRPD)

[0124] Instrument model: X'Pert3 X-ray diffractometer from PANalytical.

[0125] Test method: About 10 mg of sample was used for XRPD detection.

[0126] The detailed XRPD parameters are as follows:

[0127] Light pipe: Cu, kα

[0128] Light tube voltage: 45kV, light tube current: 40mA

[0129] Divergence slit: 0.60mm

[0130] Detector slit: 10.50mm

[0131] Anti-scatter slit: 7.10mm

[0132] Scanning range: 3-40 degrees

[0133] Step diameter: 0.0263deg

[0134] Step size: 0.12 seconds

[0135] Sample plate speed: 15rpm

[0136] 1.2 Differential Scanning Calorimeter (DSC)

[0137] Instrument model: TA 2500 differential scanning calorimeter

[0138] Test method: Take a sample (0.5-1 mg) and place it in a DSC aluminum pan for testing. Under 50 mL / min N2 conditions, heat the sample from room temperature (25°C) to 300°C or 350°C at a heating rate of 10°C / min.

[0139] 1.3 Thermogravimetric analysis (TGA)

[0140] Instrument model: TA Q5500 / Q5000 thermogravimetric analyzer

[0141] 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 (25°C) to 300°C or 350°C at a heating rate of 10°C / min, and lose 20% of its weight.

[0142] 1.4 Criteria for judging hygroscopicity:

[0143] Table 4 Judgment criteria for hygroscopicity

[0144] Moisture Classification Weight gain due to moisture absorption* deliquescence Absorb enough water to form a liquid Highly hygroscopic The weight gain due to moisture absorption is not less than 15% Hygroscopic The weight gain due to moisture absorption is less than 15% but not less than 2% Slightly hygroscopic The weight gain due to moisture absorption is less than 2% but not less than 0.2% No or almost no hygroscopicity The weight gain due to moisture absorption is less than 0.2%

[0145] *Weight gain at 25℃ / 80%RH.

[0146] 1.5 High Performance Liquid Chromatograph (HPLC)

[0147] Instrument model: Agilent 1200 HPLC

[0148] The analysis method is as follows:

[0149] Table 5 HPLC analysis method for testing the content of related substances

[0150] BRIEF DESCRIPTION OF THE DRAWINGS

[0151] Figure 1 It is the XRPD spectrum of Form A of the compound of formula (II).

[0152] Figure 2 It is the XRPD spectrum of Form B of the compound of formula (II).

[0153] Figure 3 It is the XRPD spectrum of Form C of the compound of formula (II).

[0154] Figure 4 It is the DSC spectrum of the crystalline form A of the compound of formula (II).

[0155] Figure 5 It is the TGA spectrum of the crystalline form A of the compound of formula (II).

[0156] Figure 6 This is the infrared absorption spectrum of the crystalline form A of the compound of formula (II).

[0157] Figure 7 It is the DSC spectrum of the crystalline form B of the compound of formula (II).

[0158] Figure 8 It is the TGA spectrum of the crystalline form B of the compound of formula (II).

[0159] Fig. 9 It is the DSC spectrum of Form C of the compound of formula (II).

[0160] Fig.10 It is the TGA spectrum of the crystalline form C of the compound of formula (II).

[0161] Fig.11 It is the ultraviolet absorption spectrum of the crystalline form A of the compound of formula (II).

[0162] Fig.12 It is the three-dimensional structure ellipsoid diagram of the compound of formula (I). DETAILED DESCRIPTION

[0163] The present disclosure is described in detail below by way of examples, but it is not intended to impose any adverse limitations on the present disclosure. The present disclosure has been described in detail herein, and specific embodiments thereof are also disclosed therein. It will be apparent to those skilled in the art that various changes and modifications can be made to the specific embodiments of the present disclosure without departing from the spirit and scope of the present disclosure.

[0164] Example 1: Compound of formula (I)

[0165]

[0166] Step 1: Synthesis of Intermediate 1-B

[0167] At room temperature, DMF-DMA (19.05 mol, 1.0 eq, 2.53 L) was added to a 30 L high and low temperature reactor, and raw material 1-A (2403.52 g, 19.05 mol, 1.0 eq) was added in batches under stirring. The reaction was slightly exothermic, and the dropping speed was controlled to ensure that the temperature of the reaction liquid did not exceed 35 ° C. After the addition was completed, the temperature was raised to 85-90 ° C and stirred for 3 hours. The reaction liquid was cooled to 50-55 ° C, and the solvent methanol (2.4 L) was added, and then hydrazine hydrate (17.15 mol, 98% purity, 0.9 eq, 850 mL) was slowly added dropwise. The reaction was exothermic, and the feeding speed was controlled to prevent the material from rushing. After the addition was completed, the reaction was carried out at 50-60 ° C for 0.5 hours. The reaction mixture was cooled to room temperature, and the reaction solution was concentrated under reduced pressure at 50°C and -0.08 MPa. The solid obtained by concentration was added to ethyl acetate (2.4 L) for slurrying, stirred for 0.5 hours, filtered, and the filter cake was collected. The filtrate was concentrated under reduced pressure to obtain a crude product. The crude product was added to ethyl acetate (1.2 L) for slurrying, stirred for 0.5 hours at room temperature, filtered, and the filter cake was collected. The combined filter cakes were vacuum dried at 45-50°C and -0.08 MPa to constant weight to obtain 1-B. LCMS (ESI) m / z: 151 (M+1); 1 H NMR (DMSO-d6, 400MHz) δppm 13.02 (s, 1H), 7.82 (s, 1H), 2.91-2.96 (m, 2H), 2.56-2.61 (m, 2H), 1.85-1.91 (m, 2H), 1.78-1.84 (m, 2H).

[0168] Step 2: Synthesis of Intermediate 1-C

[0169] At room temperature, add solvent dichloromethane (11.07L) to a 50L high and low temperature reactor, add intermediate 1-B (2214.90g, 14.74mol, 1.0eq) and N,N-dimethylformamide (3.40L, 44.23mol, 3.0eq) in batches under stirring, and then slowly add oxalyl chloride (3.87L, 44.23mol, 3.0eq) dropwise, and control the dropwise addition speed so that the temperature of the reaction solution does not exceed 30°C. The reaction solution is stirred at 20-30°C for 12 hours. The reaction solution is concentrated to dryness under reduced pressure at 40°C and -0.08MPa. The reduced pressure concentrated crude product is added to ice water (22.14L) in batches under stirring at room temperature, and filtered after stirring for 0.5 hours. The filter cake is collected and washed with water (2L*3). The filter cake is vacuum dried at 35-40°C and -0.08MPa to obtain 1-C. LCMS (ESI) m / z: 197 (M+1); 1H NMR (DMSO-d6, 400MHz) δppm 10.2 (s, 1H), 8.03 (s, 1H), 2.9-3.0 (m, 2H), 2.6-2.6 (m, 2H), 1.8-1.8 (m, 2H).

[0170] Step 3: Synthesis of Intermediate 1-D

[0171] At room temperature, add solvent THF (14.65L) to a 50L high and low temperature reactor, add intermediate 1-C (2929.34g, 14.84mol, 1.0eq) and concentrated ammonia (4.57L, 29.69mol, 2.0eq) in batches under stirring, and slowly add iodine (7540.23g, 29.69mol, 2.0eq) in batches, and control the iodine addition rate so that the temperature of the reaction solution does not exceed 35°C. The reaction solution was stirred and reacted at 20-35°C for 12 hours. The above reaction solution was added to a saturated sodium sulfite aqueous solution containing 120L under stirring, and filtered after stirring for 0.5 hours. The filter cake was collected, washed with water (3L*3), and vacuum dried at 45-50°C and -0.08MPa to constant weight to obtain 1-D. LCMS (ESI) m / z: 194 (M+1); 1 H NMR (DMSO-d6, 400MHz) δppm 13.06-13.27(m,1H), 7.71-8.11(m,1H), 2.90-2.96(m,2H), 2.67(m,2H), 1.90(m,2H).

[0172] Step 4: Synthesis of Intermediate 1-E

[0173] At room temperature, THF (9.70 L) was added to a 50 L high and low temperature reactor, followed by the addition of intermediate 1-D (1.94 Kg, 9.82 mol, 98.061% purity, 1 eq) and DHP (2.48 kg, 29.47 mol, 2.69 L, 3.0 eq, 1000 mL / min) to the reactor. The reaction solution was stirred at 75-80 ° C for 65 hours. The reaction solution was concentrated under reduced pressure at 50 ° C and -0.08 MPa to obtain an oil. At room temperature, the solvent DMF (15.90L) was added to a 50L high and low temperature reactor, and then the raw materials obtained above were added to the reactor in batches (3180.26g, 11.45mol, 1eq), mercaptoacetamide (1252.14g, 13.74mol, 1.2eq) and potassium carbonate (3323.28kg, 24.05mol, 2.1eq) in batches, without exothermic phenomenon. The reaction solution was reacted at 75-80°C for 1 hour (keep the system unobstructed and a slight nitrogen flow, ensure continuous stirring, tail gas needs sodium hypochlorite solution to absorb, and there is a buffer safety bottle between the tail gas absorption device and the reactor). The reaction solution was cooled to room temperature and ethyl acetate (15.90L) was added. The above reaction solution was slowly poured into 160L water under stirring. After the addition was completed, the reaction solution was a suspension. After stirring at room temperature for 12 hours, it was filtered (solid precipitation was slow), and the filter cake was collected and washed with water (3L*3). Add 15.90 L of methanol to the filter cake and stir at 65-70°C for 1 hour, cool to room temperature and filter, collect the filter cake (if the purity is not enough, repeat the above purification process), and vacuum dry at 45-50°C and -0.08 MPa to obtain 1-E. LCMS (ESI) m / z: 333 (M+1); 1 H NMR(DMSO-d6,400MHz)δppm7.52-7.92(m,1H),6.62-6.89(m,2H),6.29-6.50(m,2H),5.22-5.52(m,1H),3.83 -3.99(m,1H),3.55-3.70(m,1H),2.84-3.18(m,2H),2.60-2.71(m,2H),1.86-2.35(m,5H),1.43-1.75(m,3H).

[0174] Step 5: Synthesis of Intermediate 1-G

[0175] At room temperature, the raw material 1-F (906.50 g, 4.73 mol, 1 eq, HCl) was added to dichloromethane (9.6 L), and oxalyl chloride (331.13 g, 2.61 mol, 228.37 mL, 5 eq) was added dropwise at 10-15 ° C, and stirred for 72 hours. The reaction solution was concentrated to dryness under reduced pressure, and 5 L of DCM was added to concentrate under reduced pressure to remove the remaining oxalyl chloride, and the reaction was repeated once. The obtained 1-G was directly used for the next step reaction.

[0176] Step 6: Synthesis of intermediate 1-H

[0177] At room temperature, 9.2L of dichloromethane was added to a 30L high and low temperature reactor. 1-G (983.28g, 4.68mol, 1.7eq, HCl) was added to the reactor under stirring. The temperature was lowered to -20--15°C. 1-E (920.19g, 2.77mol, 1eq) was slowly added to the reactor in batches, and the internal temperature was controlled not to exceed -10°C. The reaction was stirred at -20--10°C for 0.5 hours. HPLC detected that the content of 1-E was <5%. Triethylamine (192mL, 1.39mol, 0.5eq) was added dropwise to the reactor at -20--10°C. Then, a sample was taken for testing. HPLC showed that the content of 1-E was <1%. The remaining triethylamine (960mL, 6.93mol, 2.5eq) was added dropwise to the reactor. The mixture was concentrated under reduced pressure, and the solid was about 3 kg. 15 L of n-heptane was added, stirred and slurried to remove dichloromethane. The mixture was filtered, and the solid was collected and dried under reduced pressure (45°C, -0.1 MPa) to obtain 1-HM.

[0178] 8.7L of methanol and 4.3L of water (methanol: water = 2:1) were added to a high-low temperature reactor, and sodium hydroxide (1107.27g, 27.7mol, 10eq) was added and the temperature was raised to 50°C. The dried 1-HM was added to the reactor in batches, and the reaction was stirred for 0.5 hours. HPLC detected that the reaction was complete, cooled to 0°C, 13L of water was added, and stirred at 0-5°C for 0.5 hours, and the solid was collected by filtration. The filtrate was adjusted to pH 7-8 with 6N hydrochloric acid, and most of the methanol was concentrated under reduced pressure. The pH was adjusted to 10-11 with 2N sodium hydroxide at 10-15°C, and a large amount of solid precipitated. Stirring was continued for 0.5 hours, and the filter cake was collected by filtration. The two filter cakes were combined and washed with water (4L*2). After filtration, the filter cake was dried under reduced pressure (55°C, -0.08MPa) to obtain 1-H. LCMS (ESI) m / z: 452 (M+1); 1H NMR(DMSO-d6+D2O,400MHz)δppm 7.66-8.08(m,1H),5.30-5.43(m,1H),3.85-4.00(m,1H),3.55-3.76(m,3H),2.89-3.12(m,6H),2.74-2.87(m ,1H),2.37-2.45(m,1H),2.05-2.22(m,1H),1.86-2.05(m,4H),1.76-1.84(m,1H),1.61-1.75(m,2H),1.60(br s,5H),1.28-1.41(m,1H).

[0179] Step 7: Synthesis of Intermediate 1-I

[0180] 1-H (1197.21 g, 2.65 mol, 1 eq) was added to 4N hydrogen chloride ethyl acetate solution (11.97 L, 47.88 mol, 18.1 eq) in batches under stirring at 20-25°C. After the addition was completed, the mixture was stirred at 10-15°C for 0.5 hours and filtered. The filter cake was washed with ethyl acetate (1 L*2) and dried under reduced pressure (40-45°C, -0.1 MPa) for 16 hours to obtain 1-IM. 1-IM was added to a mixed solvent of water (9.6 L) and methanol (1.2 L), and the pH was adjusted to 8-9 with concentrated ammonia water. The mixture was stirred for 0.5 hours, and the filter cake was collected by filtration. The filter cake was washed with water (5 L*2) and filtered, and dried under reduced pressure (55°C, -0.1 MPa) to obtain 1-I. LCMS (ESI) m / z: 368 (M+1); 1 H NMR(DMSO-d6+D2O,400MHz)δppm 7.46(s,1H),3.64-3.78(m,1H),2.89-3.03(m,5H),2.72-2.89(m,3H),2.30-2.36(m,1H),1.73-1.88(m,4H),1.35-1.54(m,4H).

[0181] Step 8: Synthesis of compound of formula (I)

[0182] First split:

[0183] At room temperature, add 10.1 L of methanol to a 30 L high and low temperature reactor, add 1-I (807.89 g, 2.20 mol, 1 eq) to the reactor, add glacial acetic acid (4 L) and water (404 mL) in sequence under stirring. When the reactor temperature rises to 60°C, add 3.7 L of methanol dissolved with (+)-di-p-toluoyl-D-tartaric acid (374.59 g, 0.97 mmol, 0.44 eq) in batches, and stir at 60-65°C for 12 hours. The temperature was gradually decreased to 20°C at a rate of 10°C / 30min, and then filtered. The filter cake was washed with methanol (4.6L). After filtration, the filter cake was dispersed in a mixed solvent of water (8L) and methanol (1L), and the pH was adjusted to 8-9 with concentrated aqueous ammonia. The mixture was stirred for 0.5 hours, and the filter cake was collected by filtration. The filter cake was washed twice by slurrying with water, and the filter cake was collected by filtration. After drying under reduced pressure (55°C, -0.1MPa), 1-J-M1 (89.116% ee) was obtained.

[0184] Second split:

[0185] At room temperature, 3.78L of methanol and 1-J-M1 (302.67g, 0.824mol, 1eq) were added to a 10L high and low temperature reactor. Glacial acetic acid (1.5L) and water (151mL) were added under stirring. When the reactor temperature rose to 60°C, 2.74L of methanol dissolved with (+)-di-p-toluoyl-D-tartaric acid (273.67g, 0.708mmol, 0.86eq) was added at once. The reaction solution was stirred at 60-65°C and under a slight nitrogen flow for 12 hours. After cooling to room temperature at a rate of 10°C / 30min, the filter cake was filtered, and the filter cake was slurried with methanol (3.2L). After filtering, it was dispersed into a mixed solvent of water (8.52L) and methanol (1.07L), and the pH was adjusted to 8-9 with concentrated ammonia water. After stirring for 0.5 hours, the filter cake was collected by filtration, and the filter cake was washed twice with water and collected by filtration. The filter cake was dried under reduced pressure (55°C, -0.1 MPa) to obtain 1-J-M2 (98.040% ee).

[0186] The third split:

[0187] At room temperature, 3.14L of methanol and 1-J-M2 (251.36g, 0.68mol, 1eq) were added to a 10L high and low temperature reactor, and glacial acetic acid (1.25L) and water (125mL) were added in sequence under stirring. When the kettle temperature rose to 60°C, 2.39L of methanol dissolved with (+)-di-p-toluoyl-D-tartaric acid (239.22g, 0.62mmol, 0.91eq) was added at one time. The reaction solution was stirred for 12 hours at 60-65°C and a slight nitrogen flow. After cooling to room temperature at a rate of 10°C / 30min, the filter cake was filtered, and the filter cake was slurried with methanol (3L). After filtration, it was dispersed in a mixed solvent of water (4.8L) and methanol (0.6L), and the pH was adjusted to 8-9 with concentrated ammonia water. Stir for 0.5 hours, filter and collect the filter cake, and wash the filter cake twice with water slurry, and collect the filter cake after filtration. The filter cake was dried under reduced pressure (55°C, -0.1 MPa) to obtain the compound of formula (I) (99.76% ee). LCMS (ESI) m / z: 368 (M+1); 1 H NMR (DMSO-d6, 400MHz): δ (ppm) 7.92 (m, 1H), 3.86-3.92 (m, 1H), 3.03-3.13 (m, 5H), 2.82-2.92 (m, 1H), 2.55-2.67 (m, 2 H),2.29-2.39(m,1H),1.93-2.05(m,2H),1.83-1.92(m,1H),1.70-1.81(m,1H),1.50-1.62(m,2H),1.37-1.49(m,2H).

[0188] Example 2: Crystalline Form A of the Compound of Formula (II)

[0189]

[0190] At room temperature, methanol (1042 mL) was added to a 3L reaction bottle, and then the compound of formula (I) (45.00 g, 122.46 mmol, 1 eq) was added to the above reaction bottle, cooled to 5°C under stirring, and maleic acid (21.32 g, 183.69 mol, 1.5 eq) was added thereto, and stirred at 5-13°C for 6 hours. Filter and collect the filter cake. The filter cake was washed with methanol (500 mL*2) and then dried under reduced pressure (45°C, -0.1 MPa) to obtain the crystalline form A of the compound of formula (II). LCMS (ESI) m / z: 368 (M+1); 1H NMR (heavy water, 400 MHz): δ ppm 7.46 (s, 1H), 6.16 (s, 2H), 4.45-4.55 (m, 1H), 3.64-3.81 (m, 1H), 3.48-3.59 (m, 1H), 3.34-3.47 (m, 2H), 2.76-2.87 (m, 2H), 2.65-2.76 (m, 2H), 2.41-2.53 (m, 1H), 2.26-2.37 (m, 1H), 1.95-2.09 (m, 3H), 1.85-1.95 (m, 2H), 1.63-1.84 (m, 2H).

[0191] 1 H NMR showed that the salt number of Form A of the compound of formula (II) was 1.

[0192] Example 3: Crystalline Form B of the Compound of Formula (II)

[0193] The compound of formula (II) crystal form A (60.91 mg) was weighed, and a mixture of acetonitrile (1 mL) and water (1 mL) was added, and the solid was dissolved under stirring. The solvent was evaporated to dryness at 40° C., and the solid precipitated. The solid was collected and dried under reduced pressure (55° C., −0.1 MPa) to obtain the compound of formula (II) crystal form B. LCMS (ESI) m / z: 368 (M+1); 1 H NMR (DMSO-d6+D2O, 400MHz): δppm 7.85(s,1H),6.10(s,2H),4.60-4.64(m,1H),3.74-3.86(m,1H),3.38-3.52(m,1H),3.21-3.37(m,2H),3.04(br s,4H),2.37-2.47(m,1H),2.16-2.24(m,1H),2.02-2.12(m,1H),1.91-2.01(m,2H),1.87(br s,4H).

[0194] 1 H NMR showed that the salt number of the crystalline form B of the compound of formula (II) was 1.

[0195] Example 4: Crystalline Form C of the Compound of Formula (II)

[0196] At room temperature, methanol (56 mL) was added to the compound of formula (I) (0.80137 g, 2.18 mmol, 1 eq), and maleic acid (505.29 mg, 4.35 mmol, 2 eq) was added under stirring at 75°C, and stirred at 75°C for 0.5 hours, at 40°C for 1 hour, and then cooled to 25°C and stirred for 2 hours. The solid was collected by filtration, washed with methanol (10 mL*2), and the solid was dried under reduced pressure (55°C, -0.1 MPa) to obtain the crystalline form C of the compound of formula (II). LCMS (ESI) m / z: 368 (M+1); 1 H NMR (DMSO-d6+D2O, 400MHz): δppm 7.92(s,1H),6.09(s,2H),4.62-4.64(m,1H),3.62-3.75(m,1H),3.39-3.54(m,1H),3.23-3.37(m,2H),3.04(br s,4H),2.37-2.48(m,1H),2.15-2.28(m,1H),2.02-2.14(m,1H),1.92-2.01(m,2H),1.87(br s,4H).

[0197] 1 H NMR showed that the salt number of Form C of the compound of formula (II) was 1.

[0198] Example 5: Stability test of the crystalline form A of the compound of formula (II)

[0199] Experimental procedures

[0200] Weigh 50 mg of the crystalline form A of the compound of formula (II) and place it in a dry and clean glass bottle, spread it into a thin layer, and place it under accelerated test conditions (60°C, 25 / 92.5% RH, 40°C / 75% RH and 60°C / 75% RH). The sample is completely exposed and covered with aluminum foil and pierced with small holes. Samples were taken and analyzed at 5 days, 10 days, 1 month, 2 months and 3 months. The results are shown in Table 6.

[0201] Table 6 Stability test results of the crystalline form A of the compound of formula (II)

[0202]

[0203] RRT: relative main peak retention time; TRS: total impurity content; RH: relative humidity; “ / ” means not detected.

[0204] Conclusion: The impurity content of the crystalline form A of the compound of formula (II) remains basically unchanged under high temperature and high humidity conditions, and the stability is good.

[0205] Example 6: Hygroscopicity test of the crystalline form A of the compound of formula (II)

[0206] Test method:

[0207] Place a dry stoppered glass weighing bottle in a desiccator with a saturated ammonium chloride solution at the bottom. Place the weighing bottle open, cover the desiccator lid, and then place the desiccator in a 25°C thermostat overnight. After the weighing bottle has been placed overnight, take it out and weigh it accurately, which is recorded as m1.

[0208] Take an appropriate amount of Form A of the compound of formula (II), spread it evenly in a weighing bottle with a predetermined weight (the sample thickness is about 1 mm), and then accurately weigh the weight, which is recorded as m2.

[0209] Place the weighing bottle with the lid open and put it in a desiccator with a saturated ammonium chloride solution at the bottom. Place the weighing bottle with the lid open and cover the desiccator. Then place the desiccator in a 25°C constant temperature box for 24 hours. After 24 hours, cover the weighing bottle and take it out to accurately weigh the weight, which is recorded as m3.

[0210] Calculation of hygroscopic weight gain: weight gain percentage = 100% x (m3-m2) / (m2-m1)

[0211] Experimental results: The weight gain percentage (average value) of the crystalline form A of the compound of formula (II) is 0.18%.

[0212] Conclusion: The average hygroscopicity of the crystalline form A of the compound of formula (II) is less than 0.2%, so the crystalline form A of the compound of formula (II) has no or almost no hygroscopicity.

[0213] Example 7: Infrared absorption test of the crystalline form A of the compound of formula (II)

[0214] Instrument: Thermo Fisher Nicolet iS5

[0215] Method: ATR attenuated total reflectance method

[0216] Experimental results: The infrared absorption spectrum of the crystal form A of the compound of formula (II) is shown in Table 7. The infrared absorption spectrum of the crystal form A of the compound of formula (II) is shown in Figure 6 .

[0217] Table 7 Infrared absorption spectrum measurement results of the compound of formula (II) Form A

[0218]

[0219]

[0220] Conclusion: The structures of the tested compounds contain -NH, -OH, benzene ring, -CH2, -CH3, CO, -C=O, -C=N and other groups, which are consistent with the structure of Form A of the compound of formula (II).

[0221] Example 8: Ultraviolet absorption test of the crystalline form A of the compound of formula (II)

[0222] Instrument: Agilent G6860A UV-Vis spectrophotometer

[0223] Experimental results: The UV measurement results of the crystal form A of the compound of formula (II) are shown in Table 8. The UV absorption spectrum of the crystal form A of the compound of formula (II) is shown in Fig.11 .

[0224] Table 8 UV measurement results

[0225]

[0226] Analytical conclusion: The tested compound has ultraviolet absorption in water at λ=259.0nm, λ=214.4nm, and λ=322.6nm, which are the B-band and R-band absorption peaks of the aromatic ring, indicating that the structure contains aromatic rings, which is consistent with the structure of the crystalline form A of the compound of formula (II).

[0227] Example 9: AKT enzyme activity test of the crystalline form A of the compound of formula (II)

[0228] CDC7 kinase plays an important role in the initiation of DNA replication and DNA damage response. When CDC7 is abnormally overexpressed, abnormal activation of the CDC7-MCM2 signaling pathway axis will lead to abnormal DNA replication, and the activity of the key checkpoint proteins CHK2 and MK2 for DNA damage repair will also be inhibited, thereby inhibiting apoptosis and promoting proliferation, leading to tumorigenesis.

[0229] Experimental principle:

[0230] ADP-Glo TM The kinase assay is a chemiluminescent kinase assay. When the kinase reacts with the substrate, ATP is required to provide energy, so that ATP is converted into ADP. The amount of ADP produced in the reaction is used to measure the enzyme activity. This assay converts ADP into ATP, which is then reacted with Ultra-Glo TM Luciferase binds and emits light, and the intensity of the light is proportional to the amount of ADP.

[0231] This experiment uses the ADP-GloTM method to study the inhibitory effect of the test compound formula (II) compound crystal form A on CDC7 / DBF4 kinase activity. The highest concentration of the compound tested is 10 μM, 5-fold gradient dilution, 8 concentration points. The compound was retested on different days, and all tested twice.

[0232] Reagents and consumables:

[0233] name Manufacturer 384-well microplate Greiner 384-well compound plate Costar CDC7 / DBF4Active Signal Chem PDKtide Signal Chem Kinase Assay Buffer III Signal Chem DTT(0.1M) Signal Chem ADP-Glo ​​Kinase Assay Promega

[0234] Main instruments:

[0235] name Manufacturer model VICTOR Nivo PerkinElmer VICTOR Nivo 5F

[0236] Experimental methods:

[0237] 1. Preparation of solutions and buffers

[0238] 1) 10mM DTT preparation (prepared immediately before use)

[0239] Take 1 μL of 0.1 M DTT and add 9 μL of deionized water.

[0240] 2) 2x analysis buffer (prepared immediately before use)

[0241] Take 168 μL of Kinase Assay Buffer III, dilute it 2.5 times with 248 μL of deionized water, add 4.2 μL of 10 mM DTT, and place it on ice for later use.

[0242] 3) 1x analysis buffer (prepared immediately before use)

[0243] Take 250 μL of 2x analysis buffer and add 250 μL of deionized water to dilute it into 1x analysis buffer, and place it on ice for later use.

[0244] 4) ADP-Glo ​​Kinase Assay

[0245] The kit contains ADP-GloTM reagent, kinase assay solution and kinase assay matrix and 10mM ATP stock solution.

[0246] Preparation of kinase assay solution: After the kinase assay buffer and kinase assay matrix are equilibrated to room temperature, mix the two and use them after they are completely dissolved. The unused solution needs to be aliquoted and stored at -20°C.

[0247] ADP-GloTM reagent: Equilibrate to room temperature when used for the first time. Unused solution needs to be aliquoted and stored at -20℃.

[0248] 2. Preparation of compound working solution concentration

[0249] 1) Dilute 10 mM test compound to 1 mM with 100% DMSO, take 5 μL of 10 mM compound and add 45 μL of 100% DMSO. The concentration of the compound after dilution is 1 mM, and put it into row A of 384-well compound plate as the highest concentration;

[0250] 2) Add 20 μL of 100% DMSO to rows B to H of a 384-well compound plate;

[0251] 3) Take 5 μL of compound from row A to row B and mix well, then take 5 μL of compound from row B to row C and mix well, repeat the above steps to row H to complete the gradient dilution of the compound;

[0252] 4) Preparation of compound working solution: Take 0.5 μL of compound per well from the compound plate to a new 384-well compound plate, add 9.5 μL of 1x assay buffer; take 0.5 μL of 100% DMSO to the positive control well, add 9.5 μL of 1x assay buffer;

[0253] 5) The blank control wells are wells without CDC7 / DBF4 enzyme added, and the positive control is 1% DMSO.

[0254] 3. Reaction steps

[0255] 1) CDC7 / DBF4 enzyme stock solution, substrate PDKtide, and ATP stock solution were thawed on ice, and the above reagents need to be kept on ice during the experiment. The unused stock solution needs to be stored in aliquots to avoid repeated freezing and thawing;

[0256] 2) Take 1 μL of compound working solution per well and add it to a 384-well microplate. For the positive control, add 1 μL of 1x assay buffer containing 5% DMSO per well. For the blank control, add 1 μL of 1x assay buffer per well.

[0257] 3) After the enzyme is completely dissolved, dilute the enzyme stock solution to 3.125 ng / μL with 1x assay buffer, i.e., take 7 μL of enzyme stock solution (100 ng / μL) and dilute it with 217 μL of 1x assay buffer;

[0258] 4) Take 2 μL of enzyme solution per well and add it to the microplate. Add 2 μL of 1x assay buffer per well to the blank control well. The enzyme amount is 6.25 ng per well. Note: This step needs to be performed on ice.

[0259] 5) Use a mixture of 122 μL 2x assay buffer and 3.2 μL ATP (2 mM) to dilute 125 μL substrate PDKtide stock solution by 2 times, so that the concentration of ATP in the obtained substrate and ATP mixture is 25 μM, and the concentration of PDKtide is 0.5 mg / mL. Place the substrate and ATP mixture on ice for later use;

[0260] 6) Take 2 μL of the mixed solution of substrate and ATP from each well into the microplate. At this time, the concentration of substrate is 0.2 mg / mL, the concentration of ATP is 10 μM, and the concentration of DMSO is 1%;

[0261] 7) Seal the microplate and incubate at 25°C for 60 minutes;

[0262] 8) ADP-GloTM reagent and kinase detection solution need to be equilibrated to room temperature before use;

[0263] 9) After the incubation, take 5 μL of ADP-GloTM reagent per well and add it to the microplate. After sealing the microplate, place it at 25°C and incubate for 40 minutes.

[0264] 10) After the incubation, take 10 μL of kinase detection solution from each well and add it to the microplate. After sealing the microplate, place it at 25°C and incubate for 30 minutes.

[0265] 11) After the incubation, use Nivo to perform luminescence detection and read the luminescence value (RLU);

[0266] 12) Calculation of enzyme activity:

[0267] % enzyme activity = [(RLU(compound)-RLU(blank)) / (RLU(positive)-RLU(blank))] x 100%

[0268] Experimental Results

[0269] The percentage of enzyme activity was calculated from the original reading according to the calculation formula, and then Prism was used to draw statistics and calculate the IC of the compound. 50 , and the results are shown in the following table.

[0270]

[0271] Conclusion: Crystal form A of the compound of formula (II) has a strong inhibitory effect on the enzyme CDC7 / DBF4.

[0272] Example 10: Cellular activity test of the crystalline form A of the compound of formula (II)

[0273] This experiment studies the inhibitory effect of Form A of the compound of formula (II) on the proliferation of colorectal cells COLO205 with high expression of Cdc7.

[0274] Reagents and consumables:

[0275] 1) Compound information

[0276] Crystalline Form A of the compound of formula (II).

[0277] 2) Main instruments

[0278] name Manufacturer model Envision PerkinElmer EXT

[0279] 3) Cell information

[0280] Cell Type Cell Name Cell growth pattern Cell culture medium Cell source Human colorectal cancer cells Colo-205 Adhesion RPMI 1640+10%FBS+1%double antibody Pronose Biotech

[0281] 4) Main reagents and consumables

[0282] 1640 culture medium, Biological Industries; double antibody, Pronocell; 0.25% trypsin, Base Media; fetal bovine serum, Biosera; DMSO, Sinopharm; 96-well cell culture plate, Corning; 96-well compound plate, Shanghai Jingrao Biotechnology Co., Ltd.; Luminescent Cell Viability Assay, Promega; cell culture dishes, Nice.

[0283] Experimental principle:

[0284] Luminescent Cell Viability Assay( The Luminescent Cell Viability Assay Kit is a homogenous rapid assay for detecting viability of living cells in culture by quantitatively measuring ATP. ATP is an indicator of the metabolism of living cells. Cell lysis and the resulting luminescent signal are proportional to the amount of ATP present, which is directly proportional to the number of living cells in culture.

[0285] Experimental methods:

[0286] 1) Cell passaging

[0287] Digest the cells until 80%-90% cell fusion has been achieved with trypsin, terminate the digestion with 1 mL of culture medium, transfer to a new culture dish in appropriate proportion, add 10 mL of fresh cell culture medium to the culture dish, and continue culturing in a 37°C, 5% CO2 incubator.

[0288] 2) Preparation of compound working solution concentration

[0289] The compound was diluted to 2 mM with 100% DMSO, i.e., 5 μL of 10 mM compound stock solution was placed in the first column of a 96-well compound plate, and then 20 μL of 100% DMSO was added and mixed; 20 μL of 100% DMSO was added to columns 2 to 9 of the 96-well compound plate; 10 μL of compound was taken from column 1 to column 2 and mixed, and then 10 μL of compound was taken from column 2 to column 3 and mixed, and the above steps were repeated to column 9 to complete the gradient dilution of the compound; compound working solution was prepared by taking 2 μL of compound per well from the compound plate to a new 96-well compound plate, and adding 78 μL of cell culture medium per well; 2 μL of 100% DMSO was taken from the negative control well, and 78 μL of cell culture medium was added; the compound concentration was the final concentration, of which the blank control well was the culture medium well without cells, and the negative control was 0.5% DMSO.

[0290] 3) Cell inoculation and drug treatment

[0291] Digest COLO205 cells that have reached 80%-90% cell confluence with trypsin, count the cells after digestion; dilute the cell suspension with cell culture medium to the required density after counting. The cell density is shown in the following table. Inoculate 80 μL of cell suspension into each well of a 96-well cell culture plate and incubate overnight at 37°C, 5% CO2 incubator. On the day of the experiment, add 20 μL of compound working solution to each well of the cell plate and continue incubating at 37°C, 5% CO2 incubator. The specific cell plating density and incubation days are shown in the following table.

[0292] Plating density of COLO205 cells and incubation time of cells with compounds

[0293] Cell Name Plating density (cells / well) Compound incubation days (days) COLO205 3000 3

[0294] After the incubation, 100 μL CTG detection reagent was added to each well of the cell plate and incubated at 25°C for 10 minutes. After the incubation, Envision was used to detect the luminescent signal.

[0295] Experimental results:

[0296] The measured signal values ​​of each group were normalized by subtracting the background blank control well signal value. The normalized data were used to calculate the cell viability after compound treatment according to the following formula: % inhibition rate = [1-(RFU 化合物 –RFU 空白对照 ) / (RFU 阳性对照 –RFU 空白对照 )] × 100%. Blank control: treatment with only medium without cells; positive control: cells treated with 0.5% DMSO, and then Prism was used to plot and calculate the IC of the compound 50 value.

[0297] The inhibition rate was calculated from the original reading according to the calculation formula, and then Prism was used to draw statistics and calculate the IC of the compound. 50 .

[0298] Inhibitory effect of the crystal form A of the compound of formula (II) on the proliferation of COLO205 cells IC 50 As shown in the following table.

[0299]

[0300] Conclusion: Crystal form A of the compound of formula (II) has a strong inhibitory effect on COLO205 cells.

[0301] Example 11: Confirmation of the stereo configuration of the compound of formula (I)

[0302] The stereo configuration of the compound of formula (I) was confirmed by X-ray single crystal diffraction.

[0303] Single crystal preparation of compound of formula (I):

[0304] Weigh 20 mg of the compound of formula (I) and dissolve it in 1 mL of methanol / methyl tert-butyl ether (1:1) at room temperature. Place the sample solution in a 4 mL semi-sealed sample bottle and evaporate slowly at room temperature. On the third day, colorless columnar crystals were obtained for X-ray single crystal structure analysis.

[0305] Experimental results: The three-dimensional structure ellipsoid diagram of the compound of formula (I) is shown in Fig.12 , which is S configuration. The crystal structure data and parameters of the compound of formula (I) are shown in Tables 9, 10, 11 and 12.

[0306] Table 9 Crystal structure data and determination parameters of the compound of formula (I)

[0307]

[0308] Table 10 Atomic coordinates of the crystal of compound of formula (I) (×10 4 ) and the equivalent isotropic shift parameter

[0309]

[0310]

[0311] Table 11 Bond lengths of the crystals of the compound of formula (I) and bond angle [deg]

[0312]

[0313] Table 12 Twist angle of the crystal of compound of formula (I) [deg]

[0314]

[0315]

Claims

1. A compound of formula (II) or a crystalline form thereof:

2. The crystalline form of the compound of formula (II) according to claim 1, wherein the X-ray powder diffraction pattern of the crystalline form comprises characteristic peaks at 2θ values ​​of 11.46±0.20°, 12.06±0.20°, 16.96±0.20°, 17.60±0.20°, 18.48±0.20°, 19.55±0.20°, 24.03±0.20° and 25.16±0.20°.

3. The crystalline form of the compound of formula (II) according to claim 2, wherein the X-ray powder diffraction pattern of the crystalline form comprises 2θ values ​​of 6.46°±0.20°, 9.36°±0.20°, 11.46°±0.20°, 12.06°±0.20°, 12.39°±0.20°, 12.89°±0.20°, 13.37°±0.20°, 13.87°±0.2 0°、14.09°±0.20°、16.10°±0.20°、16.96°±0.20°、17.19°±0.20°、17.60°±0.20°、18.48°±0.20°、18.75°±0.20°、19.37°±0.20°、19.55°±0.20°、21.21°±0.20°、21.65°±0.20°、 22.36°±0.20°、23.06°±0.20°、23.42°±0.20°、23.74°±0.20°、24.03°±0.20°、25.16°±0.20°、25.47°±0.20°、26.59°±0.20°、27.32°±0.20°、28.11°±0.20°、28.77°±0.20°、29 The characteristic peaks are at .73°±0.20°, 31.81°±0.20°, 33.09°±0.20°, 33.80°±0.20°, 34.19°±0.20°, 35.49°±0.20°, 35.99°±0.20°, 37.66°±0.20°, 38.14°±0.20°, 38.77°±0.20° and 39.38°±0.20°.

4. The crystalline form of the compound of formula (II) according to claim 2 or 3, wherein the crystalline form has the following X-ray powder diffraction pattern data:

5. The crystalline form of the compound of formula (II) according to claim 2 or 3, whose X-ray powder diffraction pattern is shown in Figure 1.

6. The crystalline form of the compound of formula (II) according to claim 2 or 3, wherein the DSC curve of the crystalline form has a starting point of an endothermic peak at 230.7°C±3°C.

7. The crystal form of the compound of formula (II) according to claim 2 or 3, whose DSC curve spectrum is shown in Figure 4.

8. The crystalline form of the compound of formula (II) according to claim 1, wherein the X-ray powder diffraction pattern of the crystalline form comprises characteristic peaks at 2θ values ​​of 5.86±0.20°, 10.17±0.20°, 11.73±0.20°, 15.83±0.20°, 17.64±0.20°, 23.59±0.20°, 24.89±0.20° and 25.80±0.20°.

9. The crystalline form of the compound of formula (II) according to claim 8, wherein the X-ray powder diffraction pattern of the crystalline form comprises 2θ values ​​of 5.86°±0.20°, 10.17°±0.20°, 11.73°±0.20°, 12.57°±0.20°, 14.15°±0.20°, 14.40°±0.20°, 15.23°±0.20°, 15.83°±0. 20°、16.26°±0.20°、16.71°±0.20°、17.64°±0.20°、18.04°±0.20°、18.73°±0.20°、19.99°±0.20°、20.57°±0.20°、21.08°±0.20°、23.59°±0.20°、24.36°±0.20°、24.89°±0.20° 、25.41°±0.20°、25.80°±0.20°、27.20°±0.20°、27.90°±0.20°、28.90°±0.20°、29.39°±0.20°、29.70°±0.20°、30.52°±0.20°、30.81°±0.20°、31.99°±0.20°、34.29°±0.20°、35 The characteristic peaks are at .83°±0.20°, 39.64°±0.20°, 28.90°±0.20°, 29.39°±0.20°, 29.70°±0.20°, 30.52°±0.20°, 30.81°±0.20°, 31.99°±0.20°, 34.29°±0.20°, 35.83°±0.20° and 39.64°±0.20°.

10. The crystalline form of the compound of formula (II) according to claim 8 or 9, wherein the crystalline form has the following X-ray powder diffraction pattern data:

11. The crystalline form of the compound of formula (II) according to claim 8 or 9, whose X-ray powder diffraction pattern is shown in Figure 2.

12. The crystalline form of the compound of formula (II) according to claim 8 or 9, wherein the DSC curve of the crystalline form has onset points of endothermic peaks at 65.1°C±3°C, 113.7°C±3°C, 208.8°C±3°C and 221.1°C±3°C.

13. The crystal form of the compound of formula (II) according to claim 8 or 9, whose DSC curve is shown in Figure 7.

14. The crystalline form of the compound of formula (II) according to claim 1, wherein the X-ray powder diffraction pattern of the crystalline form comprises characteristic peaks at 2θ values ​​of 7.40±0.20°, 11.21±0.20°, 13.95±0.20°, 15.01±0.20°, 15.72±0.20°, 20.61±0.20°, 22.18±0.20° and 23.82±0.20°.

15. The crystalline form of the compound of formula (II) according to claim 14, wherein the X-ray powder diffraction pattern of the crystalline form comprises 2θ values ​​of 7.40°±0.20°, 10.50°±0.20°, 11.21°±0.20°, 11.81°±0.20°, 13.05°±0.20°, 13.95°±0.20°, 15.01°±0.20°, 15.72°, 16.28°, 17.64°, 18. Characteristic peaks at 35°, 18.73°, 19.53°, 20.14°, 20.61°, 22.18°, 22.51°, 23.82°, 24.37°, 25.49°, 26.36°±0.20°, 27.19°±0.20°, 28.93°±0.20°, 30.70°±0.20°, 31.60°±0.20°, 32.50°±0.20° and 34.33°±0.20°.

16. The crystalline form of the compound of formula (II) according to claim 14 or 15, wherein the crystalline form has the following X-ray powder diffraction pattern data:

17. The crystalline form of the compound of formula (II) according to claim 14 or 15, whose X-ray powder diffraction pattern is shown in Figure 3.

18. Use of the compound of formula (II) or a crystal form thereof according to any one of claims 1 to 17 in the preparation of a Cdc7 inhibitor.

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