Crystals of nitrogen-containing heterocyclic compound having MYT1 inhibitory activity

Abstract

To provide crystals of a compound having inhibitory activity on MYT1, a salt thereof, or a solvate of the compound or salt.　Crystals of a compound represented by formula (1), a salt thereof, or a solvate of the compound or salt.

Description

Crystals of nitrogen-containing heterocyclic compounds having MYT1 inhibitory activity

[0001] The present invention relates to a crystal of a nitrogen-containing heterocyclic compound or a salt thereof, or a solvate thereof, which has MYT1 inhibitory activity, and also to a pharmaceutical agent containing the compound or salt as an active ingredient, which is useful for treating and preventing cancer.

[0002] Cells are exposed to exogenous or endogenous stress factors daily, resulting in DNA damage. Typically, cells with DNA damage repair the DNA damage by activating the DDR (DNA Damage Response) pathway. It is known that many cancers have abnormalities in genes and proteins involved in the DDR pathway, and treatments are being investigated (Non-Patent Documents 1-2). One method for treating cancers with abnormalities in genes and proteins involved in the DDR pathway is the use of PARP inhibitors, which are used to treat cancers with BRCA1 mutations or BRCA2 mutations (Non-Patent Document 3). Currently, genes and proteins involved in the DDR pathway, such as ATM, ATR, CHK1, DNA-PK, and WEE1, are being identified as potential targets, and active research is being conducted on these targets (Non-Patent Document 4).

[0003] MYT1 is an enzyme of the WEE1 family and is known to be a negative regulator of CDK1. MYT1 phosphorylates CDK1, inactivating it and playing an important role in regulating the cell cycle. In recent years, new MYT1 inhibitors have been reported, which have been shown to induce synthetic lethality in certain cancers (Non-Patent Document 5). However, no approved drugs have yet been developed, and new therapeutic agents are desired.

[0004] Cancers (Basel), 2020 Apr 23;12(4):1050. Front Pharmacol. , 2021 Feb 8; 11:629266. N. Engl. J. Med. , 2018 Dec 27;379(26):2495-2505. Nat. Rev. Cancer, 2023 23, 78-94. Nature, 2022 Apr; 604 (7907): 749-756. PLoS Compute Biol. 2019 May 20;15(5):e1006752.

[0005] An object of the present invention is to provide a compound having MYT1 inhibitory activity, or a salt thereof, or a crystal of a solvate thereof, and to provide a pharmaceutical agent containing the compound or salt as an active ingredient, which is useful for the treatment and prevention of cancer.

[0006] The present inventors have conducted extensive research to solve the above problems and have found that a compound represented by the following formula (1), or a salt thereof, or a solvate thereof, has excellent MYT1 inhibitory activity, and have also found crystals of a compound represented by the following formula (1), or a salt thereof, or a solvate thereof, thereby completing the present invention.

[0007] That is, in one aspect of the present invention, the following invention is provided: [A1] A compound represented by the following formula (1): A crystal of a compound represented by formula (1), or a salt thereof, or a solvate thereof. [A2] The crystal according to [A1], wherein the compound represented by formula (1), or a salt thereof, or a solvate thereof is the compound represented by formula (1). [A3] The crystal according to [A1], wherein the compound represented by formula (1), or a salt thereof, or a solvate thereof is a salt of the compound represented by formula (1). [A4] The crystal according to [A1], wherein the compound represented by formula (1), or a salt thereof, or a solvate thereof is a solvate of the compound represented by formula (1) or a solvate of a salt of the compound. [A5] The crystal according to [A1] or [A4], wherein the compound represented by formula (1), or a salt thereof, or a solvate thereof is a solvate of a salt of the compound represented by formula (1). [A6] The crystal according to [A1] or [A3], wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof, is at least one selected from the group consisting of hydrochloride, hydrobromide, hydroiodide, sulfate, mesylate, esylate, tosylate, besylate, phosphate, nitrate, acetate, oxalate, maleate, fumarate, citrate, malate, malonate, gluconate, mandelate, salicylate, fluoroacetate, trifluoroacetate, camsylate, tartrate, propionate, glutarate, lithium salt, sodium salt, potassium salt, cesium salt, rubidium salt, magnesium salt, calcium salt, strontium salt, barium salt, alkylammonium salt, dialkylammonium salt, trialkylammonium salt, and tetraalkylammonium salt of the compound represented by formula (1). [A7] The crystal according to [A1] or [A3], wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is at least one selected from the group consisting of hydrochloride, hydrobromide, hydroiodide, sulfate, mesylate, esylate, tosylate, besylate, phosphate, nitrate, acetate, oxalate, maleate, fumarate, citrate, malate, malonate, gluconate, mandelate, salicylate, fluoroacetate, trifluoroacetate, camsylate, tartrate, propionate, and glutarate salts of the compound represented by formula (1).[A8] The crystal according to [A1] or [A3], wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is at least one selected from the group consisting of hydrochloride, hydrobromide, sulfate, mesylate, tosylate, and besylate of the compound represented by formula (1). [A9] The crystal according to [A1] or [A3], wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is at least one selected from the group consisting of hydrochloride, sulfate, and mesylate of the compound represented by formula (1). [A10] The crystal according to [A1] or [A3], wherein the salt of the compound represented by formula (1) is the hydrochloride of the compound. [A11] The crystal according to [A1], [A4], or [A5], wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof, is at least one solvate selected from the group consisting of the compound represented by formula (1) and hydrochloride, hydrobromide, hydroiodide, sulfate, mesylate, esylate, tosylate, besylate, phosphate, nitrate, acetate, oxalate, maleate, fumarate, citrate, malate, malonate, gluconate, mandelate, salicylate, fluoroacetate, trifluoroacetate, camsylate, tartrate, propionate, glutarate, lithium salt, sodium salt, potassium salt, cesium salt, rubidium salt, magnesium salt, calcium salt, strontium salt, barium salt, alkylammonium salt, dialkylammonium salt, trialkylammonium salt, and tetraalkylammonium salt of the compound. [A12] The crystal according to [A1], [A4] or [A5], wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is at least one solvate selected from the group consisting of the compound represented by formula (1) and hydrochloride, hydrobromide, hydroiodide, sulfate, mesylate, esylate, tosylate, besylate, phosphate, nitrate, acetate, oxalate, maleate, fumarate, citrate, malate, malonate, gluconate, mandelate, salicylate, fluoroacetate, trifluoroacetate, camsylate, tartrate, propionate, and glutarate salts of the compound.[A13] The crystal according to [A1], [A4] or [A5], wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is the compound represented by formula (1) and at least one solvate selected from the group consisting of hydrochloride, hydrobromide, sulfate, mesylate, tosylate, and besylate of the compound. [A14] The crystal according to [A1], [A4] or [A5], wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is the compound represented by formula (1) and at least one solvate selected from the group consisting of hydrochloride, sulfate, and mesylate of the compound. [A15] The crystal according to [A1], [A4] or [A5], wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is a solvate of the hydrochloride of the compound represented by formula (1). [A16] The crystal according to [A1], [A4], or [A5], wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is a solvate of the compound represented by formula (1) or a salt of the compound with a solvent selected from the group consisting of water, methanol, ethanol, 1-propanol, 2-propanol, 1-butanol, 2-butanol, benzyl alcohol, acetonitrile, ethyl acetate, n-propyl acetate, n-butyl acetate, tetrahydrofuran, 1,4-dioxane, t-butyl methyl ether, dichloromethane, dimethyl sulfoxide, acetic acid, formic acid, acetone, 2-butanone, methyl isobutyl ketone, anisole, toluene, chlorobenzene, formamide, dimethylformamide, and dimethylacetamide. [A17] The crystal according to [A1], [A4], or [A5], wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is a solvate of the compound represented by formula (1) or a salt of the compound with a solvent selected from the group consisting of water, ethanol, 1-propanol, 2-propanol, 2-butanol, benzyl alcohol, tetrahydrofuran, dimethyl sulfoxide, acetic acid, formic acid, acetone, formamide, dimethylformamide, and dimethylacetamide.[A18] The crystal according to [A1], [A4], or [A5], wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is a solvate of the compound represented by formula (1) or a salt of the compound with a solvent selected from the group consisting of water, ethanol, 2-propanol, 2-butanol, dimethyl sulfoxide, acetic acid, and formic acid. [A19] The crystal according to [A1], [A4], or [A5], wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is a solvate of the compound represented by formula (1) or a salt of the compound with a solvent selected from the group consisting of water and ethanol. [A20] The crystal according to [A1], [A4], or [A5], wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is a hydrate of the compound represented by formula (1) or a hydrate of a salt of the compound. [A21] The crystal according to [A1], [A4], or [A5], wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is a solvate of the compound represented by formula (1) or a salt of the compound with ethanol. [A22] The crystal according to [A1], [A4], or [A5], wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is a solvate of at least one selected from the group consisting of the compound represented by formula (1) and hydrochloride, hydrobromide, sulfate, mesylate, tosylate, and besylate of the compound with a solvent selected from the group consisting of water, ethanol, 2-propanol, 2-butanol, dimethyl sulfoxide, acetic acid, and formic acid. [A23] The crystal according to [A1], [A4], or [A5], wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is a hydrate of the hydrochloride of the compound represented by formula (1). [A24] The crystal according to [A1], [A4] or [A5], wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is a monohydrate of the hydrochloride of the compound represented by formula (1). [A25] The crystal according to [A1] or [A4], wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is a hydrate of the compound represented by formula (1).[A26] The crystal according to [A1] or [A3], wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is the hydrochloride of the compound represented by formula (1). [A27] The crystal according to [A1], [A4], or [A5], wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is a dihydrate of the 0.5 hydrochloride of the compound represented by formula (1). [A28] The crystal according to [A1], [A4], or [A5], wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is a hydrate of the 1.5 hydrochloride of the compound represented by formula (1). [A29] The crystal according to [A1], [A4], or [A5], wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is a hydrate of the sulfate of the compound represented by formula (1). [A30] The crystal according to [A1], [A4] or [A5], wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is a hydrate of 0.5 mesylate of the compound represented by formula (1). [A31] The crystal according to [A1], [A4] or [A5], wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is a hydrate of the mesylate of the compound represented by formula (1). [A32] The crystal according to [A1], [A4] or [A5], wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is a hydrate of the tosylate of the compound represented by formula (1). [A33] The crystal according to any one of [A1] to [A32], wherein the crystal comprises at least one, three, five, seven, or nine peaks selected from the group consisting of 6.82°, 8.98°, 10.35°, 10.89°, 11.88°, 12.09°, 12.36°, 12.72°, 13.46°, and 15.27° (±0.2°) as diffraction angles (2θ values) in powder X-ray diffraction at 23°C. [A34] The crystal according to any one of [A1] to [A32], wherein the crystal has peaks at diffraction angles (2θ values) of 6.82°, 8.98°, 10.35°, 10.89°, 11.88°, 12.09°, 12.36°, 12.72°, 13.46°, and 15.27° (±0.2°) as determined by powder X-ray diffraction at 23°C.[A35] The crystal according to any one of [A1] to [A32], wherein the crystal comprises at least one, three, five, seven, or nine peaks selected from the group consisting of 5.75°, 6.30°, 7.93°, 10.14°, 10.62°, 11.52°, 12.63°, 15.80°, 18.55°, and 23.72° (±0.2°) as diffraction angles (2θ values) in powder X-ray diffraction at 23°C. [A36] The crystal according to any one of [A1] to [A32], wherein the crystal has peaks at diffraction angles (2θ values) of 5.75°, 6.30°, 7.93°, 10.14°, 10.62°, 11.52°, 12.63°, 15.80°, 18.55°, and 23.72° (±0.2°) as determined by powder X-ray diffraction at 23°C. [A37] The crystal according to any one of [A1] to [A32], wherein the crystal comprises at least one, three, five, seven, or nine peaks selected from the group consisting of 7.01°, 8.74°, 9.14°, 10.17°, 10.82°, 11.78°, 13.31°, 13.93°, 17.00°, and 22.77° (±0.2°) as diffraction angles (2θ values) in powder X-ray diffraction at 23°C. [A38] The crystal according to any one of [A1] to [A32], wherein the crystal has peaks at diffraction angles (2θ values) of 7.01°, 8.74°, 9.14°, 10.17°, 10.82°, 11.78°, 13.31°, 13.93°, 17.00°, and 22.77° (±0.2°) as determined by powder X-ray diffraction at 23°C. [A39] The crystal according to any one of [A1] to [A32], wherein the crystal comprises at least one, three, five, seven, or nine peaks selected from the group consisting of 8.94°, 11.49°, 12.65°, 12.94°, 16.03°, 16.34°, 17.61°, 19.11°, 19.68°, and 20.64° (±0.2°) as diffraction angles (2θ values) in powder X-ray diffraction at 100°C.[A40] The crystal according to any one of [A1] to [A32], wherein the crystal has peaks at diffraction angles (2θ values) of 8.94°, 11.49°, 12.65°, 12.94°, 16.03°, 16.34°, 17.61°, 19.11°, 19.68°, and 20.64° (±0.2°) as determined by powder X-ray diffraction at 100°C. [A41] The crystal according to any one of [A1] to [A32], wherein the crystal comprises at least one, three, five, seven, or nine peaks selected from the group consisting of 9.02°, 11.47°, 12.71°, 12.98°, 16.50°, 17.83°, 19.24°, 19.68°, 20.80°, and 22.29° (±0.2) as diffraction angles (2θ values) in powder X-ray diffraction at 35°C. [A42] The crystal according to any one of [A1] to [A32], wherein the crystal has peaks at diffraction angles (2θ values) of 9.02°, 11.47°, 12.71°, 12.98°, 16.50°, 17.83°, 19.24°, 19.68°, 20.80°, and 22.29° (±0.2) as determined by powder X-ray diffraction at 35°C. [A43] The crystal according to any one of [A1] to [A32], wherein the crystal comprises at least one, three, five, seven, or nine peaks selected from the group consisting of 6.59°, 9.19°, 9.70°, 12.70°, 13.15°, 13.29°, 13.80°, 14.93°, 15.80°, and 16.45° (±0.2) as diffraction angles (2θ values) in powder X-ray diffraction at 23°C. [A44] The crystal according to any one of [A1] to [A32], wherein the crystal has peaks at diffraction angles (2θ values) of 6.59°, 9.19°, 9.70°, 12.70°, 13.15°, 13.29°, 13.80°, 14.93°, 15.80°, and 16.45° (±0.2) as determined by powder X-ray diffraction at 23°C.[A45] The crystal according to any one of [A1] to [A32], wherein the crystal comprises at least one, three, five, seven, or nine peaks selected from the group consisting of 7.07°, 9.24°, 12.06°, 14.37°, 15.24°, 15.84°, 17.86°, 18.30°, 18.52°, and 19.31° (±0.2) as diffraction angles (2θ values) in powder X-ray diffraction at 23°C. [A46] The crystal according to any one of [A1] to [A32], wherein the crystal has peaks at diffraction angles (2θ values) of 7.07°, 9.24°, 12.06°, 14.37°, 15.24°, 15.84°, 17.86°, 18.30°, 18.52°, and 19.31° (±0.2) as determined by powder X-ray diffraction at 23°C. [A47] The crystal according to any one of [A1] to [A32], wherein the crystal comprises at least one, three, five, seven, or nine peaks selected from the group consisting of 10.24°, 10.60°, 10.85°, 12.75°, 12.91°, 13.12°, 13.96°, 14.76°, 15.93°, and 16.84° (±0.2) as diffraction angles (2θ values) in powder X-ray diffraction at 23°C. [A48] The crystal according to any one of [A1] to [A32], wherein the crystal has peaks at diffraction angles (2θ values) of 10.24°, 10.60°, 10.85°, 12.75°, 12.91°, 13.12°, 13.96°, 14.76°, 15.93°, and 16.84° (±0.2) as determined by powder X-ray diffraction at 23°C. [A49] The crystal according to any one of [A1] to [A32], wherein the crystal comprises at least one, three, five, seven, or nine peaks selected from the group consisting of 6.95°, 7.50°, 9.05°, 11.28°, 14.32°, 15.13°, 15.72°, 17.89°, 19.86°, and 20.69° (±0.2°) as diffraction angles (2θ values) in powder X-ray diffraction at 23°C.[A50] The crystal according to any one of [A1] to [A32], wherein the crystal has peaks at diffraction angles (2θ values) of 6.95°, 7.50°, 9.05°, 11.28°, 14.32°, 15.13°, 15.72°, 17.89°, 19.86°, and 20.69° (±0.2°) as determined by powder X-ray diffraction at 23°C. [A51] The crystal according to any one of [A1] to [A32], wherein the crystal comprises at least one, three, five, seven, or nine peaks selected from the group consisting of 5.56°, 7.53°, 10.30°, 10.52°, 11.16°, 11.61°, 12.96°, 15.70°, 16.88°, and 17.58° (±0.2°) as diffraction angles (2θ values) in powder X-ray diffraction at 23°C. [A52] The crystal according to any one of [A1] to [A32], wherein the crystal has peaks at diffraction angles (2θ values) of 5.56°, 7.53°, 10.30°, 10.52°, 11.16°, 11.61°, 12.96°, 15.70°, 16.88°, and 17.58° (±0.2°) as determined by powder X-ray diffraction at 23°C. [A53] The crystal according to any one of [A1] to [A32], wherein the crystal comprises at least one, three, five, seven, or nine peaks selected from the group consisting of 5.75°, 8.37°, 9.38°, 10.89°, 13.14°, 16.81°, 17.36°, 17.79°, 18.37°, and 18.78° (±0.2°) as diffraction angles (2θ values) in powder X-ray diffraction at 23°C. [A54] The crystal according to any one of [A1] to [A32], wherein the crystal has peaks at diffraction angles (2θ values) of 5.75°, 8.37°, 9.38°, 10.89°, 13.14°, 16.81°, 17.36°, 17.79°, 18.37°, and 18.78° (±0.2°) as determined by powder X-ray diffraction at 23°C.[A55] The crystal according to any one of [A1] to [A32], wherein the crystal comprises at least one, three, five, seven, or nine peaks selected from the group consisting of 6.00°, 8.06°, 8.61°, 10.06°, 12.11°, 12.78°, 15.01°, 16.46°, 17.64°, and 18.09° (±0.2°) as diffraction angles (2θ values) in powder X-ray diffraction at 23°C. [A56] The crystal according to any one of [A1] to [A32], wherein the crystal comprises peaks at diffraction angles (2θ values) of 6.00°, 8.06°, 8.61°, 10.06°, 12.11°, 12.78°, 15.01°, 16.46°, 17.64°, and 18.09° (±0.2°) by powder X-ray diffraction at 23°C. [B1] A pharmaceutical composition comprising a crystal of the compound or a salt thereof or a solvate thereof according to any one of [A1] to [A56]. [B2] The pharmaceutical composition according to [B1] for the prevention and / or treatment of cancer. [B3] A method for the prevention and / or treatment of cancer, comprising administering to a subject an effective amount of the compound or a salt thereof or a solvate thereof according to any one of [A1] to [A56]. [B4] A crystal of the compound described in any one of [A1] to [A56], or a salt thereof, or a solvate thereof, for use in the prevention and / or treatment of cancer. [B5] Use of a crystal of the compound described in any one of [A1] to [A56], or a salt thereof, or a solvate thereof, for the manufacture of a pharmaceutical composition for the prevention and / or treatment of cancer. [B6] A pharmaceutical composition comprising a crystal of the compound described in any one of [A1] to [A56], or a salt thereof, or a solvate thereof, mixed with a pharmaceutically acceptable carrier or vehicle. [B7] A pharmaceutical composition comprising a crystal of the compound described in any one of [A1] to [A56], or a salt thereof, or a solvate thereof, dissolved in a pharmaceutically acceptable carrier or vehicle. [C1] A compound represented by the following formula (1): A method for producing a pharmaceutical composition containing, as an active ingredient, a compound represented by formula (1), or a salt thereof, or a solvate of the compound or salt described in any of [A1] to [A56], or a crystal of a solvate of the compound or salt, with a pharmaceutically acceptable carrier or medium. [C2] The method according to [C1], wherein the crystal of the compound represented by formula (1), or a salt thereof, or a solvate of the compound or salt is a crystal of a solvate of the hydrochloride of the compound represented by formula (1). [C3] The method according to [C2], wherein the crystal of the compound represented by formula (1), or a salt thereof, or a solvate of the compound or salt is a crystal of a hydrate of the hydrochloride of the compound represented by formula (1). [C4] The method according to [C3], wherein the crystal of the compound represented by formula (1), or a salt thereof, or a solvate of the compound or salt is a crystal of a monohydrate of the hydrochloride of the compound represented by formula (1).

[0008] According to the present invention, it is possible to provide a compound having MYT1 inhibitory activity, a salt thereof, or a crystal of a solvate thereof, and also to provide a pharmaceutical useful for treating and preventing cancer, which contains the compound or salt as an active ingredient.

[0009] The results of powder X-ray diffraction measurement of the crystals obtained in Example α1 are shown. The vertical axis is the diffraction intensity, and the horizontal axis is the diffraction angle 2θ (°). The results of simultaneous thermogravimetric and differential thermal analysis of the crystals obtained in Example α1 are shown. The horizontal axis is the temperature (°C), and the right vertical axis is the weight change (%) of the sample in the thermogravimetric analysis. The left vertical axis represents the heat flow observed in the differential thermal analysis. １

[0033] Figure 1 shows the results of H-NMR measurement. The vertical axis represents signal intensity, and the horizontal axis represents chemical shift δ (ppm).

[0034] Figure 1 shows the results of powder X-ray diffraction measurement of the crystals obtained in Example α2. The vertical axis represents diffraction intensity, and the horizontal axis represents diffraction angle 2θ (°).

[0035] Figure 1 shows the results of simultaneous thermogravimetry and differential thermal analysis of the crystals obtained in Example α2. The horizontal axis represents temperature (°C), and the right vertical axis represents the weight change (%) of the sample in thermogravimetry. The left vertical axis represents the heat flow observed in differential thermal analysis.

[0036] Figure 1 shows the results of powder X-ray diffraction measurement of the crystals obtained in Example α3. The vertical axis represents diffraction intensity, and the horizontal axis represents diffraction angle 2θ (°).

[0037] Figure 1 shows the crystal structure of the crystals obtained in Example α3. Compound 1 is depicted using the Capped Stick model, and the others are depicted using the Ball and Stick model.

[0038] Figure 1 shows the results of powder X-ray diffraction measurement of the crystals obtained in Example α4. The vertical axis represents the diffraction intensity, and the horizontal axis represents the diffraction angle 2θ (°). １ The results of H-NMR measurement are shown. The vertical axis is signal intensity, and the horizontal axis is chemical shift δ (ppm). The results of powder X-ray diffraction measurement of the crystals obtained in Example α5 are shown. The vertical axis is diffraction intensity, and the horizontal axis is diffraction angle 2θ (°). The results of powder X-ray diffraction measurement of the crystals obtained in Example α6 are shown. The vertical axis is diffraction intensity, and the horizontal axis is diffraction angle 2θ (°). The results of simultaneous thermogravimetric and differential thermal analysis of the crystals obtained in Example α6 are shown. The horizontal axis is temperature (°C), and the right vertical axis is the weight change (%) of the sample in thermogravimetric analysis. The left vertical axis represents the heat flow observed in differential thermal analysis. The results of powder X-ray diffraction measurement of the crystals obtained in Example α6 are shown. The results of simultaneous thermogravimetric and differential thermal analysis of the crystals obtained in Example α6 are shown. The horizontal axis is temperature (°C), and the right vertical axis is the weight change (%) of the sample in thermogravimetric analysis. The left vertical axis represents the heat flow observed in differential thermal analysis. １The results of H-NMR measurement are shown. The vertical axis represents signal intensity, and the horizontal axis represents chemical shift δ (ppm). The results of powder X-ray diffraction measurement of the crystals obtained in Example α7 are shown. The vertical axis represents diffraction intensity, and the horizontal axis represents diffraction angle 2θ (°). The results of simultaneous thermogravimetry, differential thermal analysis, and mass spectrometry measurement of the crystals obtained in Example α7 are shown. The horizontal axis represents temperature (°C), and the left vertical axis represents the weight change (%) of the sample in thermogravimetry (these results are for reference only, as the sample was directly measured after filtration and drying through a mesh). The right vertical axis represents the heat flow observed in differential thermal analysis (left) and the peak intensity observed in mass spectrometry (right). The results of powder X-ray diffraction measurement of the crystals obtained in Example α8 are shown. The vertical axis represents diffraction intensity, and the horizontal axis represents diffraction angle 2θ (°). The results of simultaneous thermogravimetry, differential thermal analysis, and mass spectrometry measurement of the crystals obtained in Example α8 are shown. The horizontal axis is temperature (°C), and the left vertical axis is the weight change (%) of the sample in thermogravimetric analysis (these results are for reference only, as the sample was directly measured after filtration and drying through a mesh). The right vertical axis represents the heat flow (left) observed in differential thermal analysis and the peak intensity (right) observed in mass spectrometry. The results of powder X-ray diffraction measurement of the crystals obtained in Example α9 are shown. The vertical axis represents the diffraction intensity, and the horizontal axis represents the diffraction angle 2θ (°). The results of simultaneous thermogravimetric, differential thermal, and mass spectrometry measurements of the crystals obtained in Example α9 are shown. The horizontal axis is temperature (°C), and the left vertical axis represents the weight change (%) of the sample in thermogravimetric analysis (these results are for reference only, as the sample was directly measured after filtration and drying through a mesh). The right vertical axis represents the heat flow (left) observed in differential thermal analysis and the peak intensity (right) observed in mass spectrometry. The results of powder X-ray diffraction measurement of the crystals obtained in Example α10 are shown. The vertical axis represents the diffraction intensity, and the horizontal axis represents the diffraction angle 2θ (°). The results of simultaneous thermogravimetric, differential thermal, and mass spectrometry measurements of the crystals obtained in Example α10 are shown. The horizontal axis is temperature (°C), and the left vertical axis is the weight change (%) of the sample in thermogravimetric analysis. The right vertical axis represents the heat flow observed in differential thermal analysis (left) and the peak intensity observed in mass spectrometry (right). The results of powder X-ray diffraction measurements of the crystals obtained in Example α11 are shown. The vertical axis represents the diffraction intensity, and the horizontal axis represents the diffraction angle 2θ (°).The results of simultaneous thermogravimetry, differential thermal analysis, and mass spectrometry of the crystals obtained in Example α11 are shown. The horizontal axis is temperature (°C), and the left vertical axis is the weight change (%) of the sample in thermogravimetry. The right vertical axis represents the heat flow observed in differential thermal analysis (left) and the peak intensity observed in mass spectrometry (right). The results of powder X-ray diffraction measurement of the crystals obtained in Example α12 at 100°C are shown. The vertical axis is diffraction intensity, and the horizontal axis is diffraction angle 2θ (°). The peak near 6.65° in the figure is a peak derived from the measuring instrument. The results of powder X-ray diffraction measurement of the crystals obtained in Example α12 at 35°C are shown. The vertical axis is diffraction intensity, and the horizontal axis is diffraction angle 2θ (°). The peak near 6.65° in the figure is a peak derived from the measuring instrument. The results of powder X-ray diffraction measurement of the crystals obtained in Example α13 are shown. The vertical axis is diffraction intensity, and the horizontal axis is diffraction angle 2θ (°).

[0033] Figure 1 shows the results of simultaneous thermogravimetry, differential thermal analysis, and mass spectrometry of the crystals obtained in Example α13. The horizontal axis is temperature (°C), and the left vertical axis is the weight change (%) of the sample in thermogravimetry. The right vertical axis represents the heat flow observed in differential thermal analysis (left) and the peak intensity observed in mass spectrometry (right). The crystal structure of the crystals obtained in Example α14 is shown. Compound 1 is depicted using the Capped Stick model, and the others are depicted using the Ball and Stick model.

[0034] Figure 1 shows the results of powder X-ray diffraction measurement of the crystals obtained in Example α14 (A) and the powder X-ray diffraction pattern calculated from the crystal structure obtained in Example α14 (B). The vertical axis represents the diffraction intensity, and the horizontal axis represents the diffraction angle 2θ (°).

[0035] Figure 1 shows the results of powder X-ray diffraction measurement of the crystals obtained in Example α15. The vertical axis represents the diffraction intensity, and the horizontal axis represents the diffraction angle 2θ (°). The results of powder X-ray diffraction measurement of the crystals obtained in Example α16 are shown. The vertical axis represents the diffraction intensity, and the horizontal axis represents the diffraction angle 2θ (°). The results of simultaneous measurements of thermogravimetry, differential thermal analysis, and mass spectrometry for the crystals obtained in Example α16 are shown. The horizontal axis represents temperature (°C), and the left vertical axis represents the weight change (%) of the sample in thermogravimetry. The right vertical axis represents the heat flow observed in differential thermal analysis (left) and the peak intensity observed in mass spectrometry (right). The results of powder X-ray diffraction measurements for the crystals obtained in Example α17 are shown. The vertical axis represents the diffraction intensity, and the horizontal axis represents the diffraction angle 2θ (°).

[0033] Figure 1 shows the results of simultaneous thermogravimetry and differential thermal analysis of the crystals obtained in Example α17. The horizontal axis is temperature (°C), and the right vertical axis is the weight change (%) of the sample in the thermogravimetric analysis. The left vertical axis represents the heat flow observed in the differential thermal analysis.

[0034] Figure 1 shows the results of powder X-ray diffraction analysis of the crystals obtained in Example α19. The vertical axis is diffraction intensity, and the horizontal axis is diffraction angle 2θ (°).

[0035] Figure 1 shows the results of simultaneous thermogravimetry and differential thermal analysis of the crystals obtained in Example α19. The horizontal axis is temperature (°C), and the right vertical axis is the weight change (%) of the sample in the thermogravimetric analysis. The left vertical axis represents the heat flow observed in the differential thermal analysis.

[0036] Figure 1 shows the results of powder X-ray diffraction analysis of the (A) crystal (Form G) and (B) crystal (Form A) obtained in Example α19. The vertical axis is diffraction intensity, and the horizontal axis is diffraction angle 2θ (°).

[0037] Figure 1 shows the results of powder X-ray diffraction analysis of the crystals obtained in Example α20. The vertical axis represents the diffraction intensity, and the horizontal axis represents the diffraction angle 2θ (°). The results of powder X-ray diffraction measurement of the crystals obtained in Example α22 are shown. The vertical axis represents the diffraction intensity, and the horizontal axis represents the diffraction angle 2θ (°). The results of simultaneous thermogravimetric and differential thermal analysis of the crystals obtained in Example α22 are shown. The horizontal axis represents the temperature (°C), and the right vertical axis represents the weight change (%) of the sample in the thermogravimetric analysis. The left vertical axis represents the heat flow observed in the differential thermal analysis. The crystals obtained in Example α22. １ The results of H-NMR measurement are shown. The vertical axis is signal intensity, and the horizontal axis is chemical shift δ (ppm). The results of powder X-ray diffraction measurement of the crystals obtained in Example α24 are shown. The vertical axis is diffraction intensity, and the horizontal axis is diffraction angle 2θ (°). The results of powder X-ray diffraction measurement of the crystals obtained in Example α25 are shown. The vertical axis is diffraction intensity, and the horizontal axis is diffraction angle 2θ (°). The results of powder X-ray diffraction measurement of the crystals obtained in Example α28 are shown. The vertical axis is diffraction intensity, and the horizontal axis is diffraction angle 2θ (°). The results of simultaneous thermogravimetric and differential thermal analysis of the crystals obtained in Example α28 are shown. The horizontal axis is temperature (°C), and the right vertical axis is the weight change (%) of the sample in thermogravimetric analysis. The left vertical axis represents the heat flow observed in differential thermal analysis. The results of powder X-ray diffraction measurement of the crystals obtained in Example α28 are shown. １The results of H-NMR measurement are shown. The vertical axis is signal intensity, and the horizontal axis is chemical shift δ (ppm). The results of powder X-ray diffraction measurement of the crystal obtained in Example α29 are shown. The vertical axis is diffraction intensity, and the horizontal axis is diffraction angle 2θ (°). The results of powder X-ray diffraction measurement of the crystal (Form F) obtained in Example α30 are shown. The vertical axis is diffraction intensity, and the horizontal axis is diffraction angle 2θ (°). The results of simultaneous thermogravimetric and differential thermal analysis of the crystal (Form F) obtained in Example α30 are shown. The horizontal axis is temperature (°C), and the right vertical axis is the weight change (%) of the sample in thermogravimetric analysis. The left vertical axis represents the heat flow observed in differential thermal analysis. The results of the crystal (Form F) obtained in Example α30 １

[0033] Figure 1 shows the results of H-NMR measurement. The vertical axis represents signal intensity, and the horizontal axis represents chemical shift δ (ppm).

[0034] Figure 1 shows the results of powder X-ray diffraction measurement of the crystals (Form H) obtained in Example α30. The vertical axis represents diffraction intensity, and the horizontal axis represents diffraction angle 2θ (°).

[0035] Figure 1 shows the results of powder X-ray diffraction measurement of the crystals obtained in Example α31. The vertical axis represents diffraction intensity, and the horizontal axis represents diffraction angle 2θ (°).

[0036] Figure 1 shows the results of powder X-ray diffraction measurement of the crystals obtained in Example α32. The vertical axis represents diffraction intensity, and the horizontal axis represents diffraction angle 2θ (°).

[0037] Figure 1 shows the results of powder X-ray diffraction measurement of the crystals obtained in Example α33 at 100°C. The vertical axis represents diffraction intensity, and the horizontal axis represents diffraction angle 2θ (°).

[0038] Figure 1 shows the results of powder X-ray diffraction measurement of the crystals obtained in Example α35. The vertical axis represents diffraction intensity, and the horizontal axis represents diffraction angle 2θ (°). The results of simultaneous thermogravimetric and differential thermal analysis of the crystals obtained in Example α35 are shown. The horizontal axis represents temperature (°C), and the right vertical axis represents the weight change (%) of the sample in the thermogravimetric analysis. The left vertical axis represents the heat flow observed in the differential thermal analysis. １ The results of H-NMR measurement are shown. The vertical axis is signal intensity, and the horizontal axis is chemical shift δ (ppm). The results of powder X-ray diffraction measurement of the crystals obtained in Example α39 are shown. The vertical axis is diffraction intensity, and the horizontal axis is diffraction angle 2θ (°). The results of simultaneous thermogravimetric and differential thermal analysis of the crystals obtained in Example α39 are shown. The horizontal axis is temperature (°C), and the right vertical axis is the weight change (%) of the sample in thermogravimetric analysis. The left vertical axis represents the heat flow observed in differential thermal analysis. The results of simultaneous thermogravimetric and differential thermal analysis of the crystals obtained in Example α39 are shown. １

[0033] Figure 1 shows the results of H-NMR measurement. The vertical axis represents signal intensity, and the horizontal axis represents chemical shift δ (ppm). The results of powder X-ray diffraction measurement of the crystals obtained in Example α40, when heated to 150°C and then cooled to 25°C, are shown. The vertical axis represents diffraction intensity, and the horizontal axis represents diffraction angle 2θ (°). The peak near 6.65° in the figure is a peak derived from the measuring instrument. The results of powder X-ray diffraction measurement of the crystals (Form B) obtained in Example α41 are shown. The vertical axis represents diffraction intensity, and the horizontal axis represents diffraction angle 2θ (°). The results of powder X-ray diffraction measurement of the crystals (Form H) obtained in Example α41 are shown. The vertical axis represents diffraction intensity, and the horizontal axis represents diffraction angle 2θ (°). The results of simultaneous thermogravimetry and differential thermal analysis of the crystals (Form B) obtained in Example α41 are shown. The horizontal axis represents temperature (°C), and the right vertical axis represents the weight change (%) of the sample in the thermogravimetric analysis. The left vertical axis represents the heat flow observed in differential thermal analysis. １

[0033] Figure 1 shows the results of H-NMR measurement. The vertical axis represents signal intensity, and the horizontal axis represents chemical shift δ (ppm). The crystal structure of the crystal obtained in Example α42 is shown. The diagram is drawn using a capped stick model.

[0034] Figure 1 shows the results of powder X-ray diffraction measurement of the crystal obtained in Example α42 (A) and the powder X-ray diffraction pattern calculated from the crystal structure obtained in Example α42 (B). The vertical axis represents diffraction intensity, and the horizontal axis represents diffraction angle 2θ (°). The results of powder X-ray diffraction measurement of the crystal obtained in Example α43 when heated to 150°C and then cooled to 30°C are shown. The vertical axis represents diffraction intensity, and the horizontal axis represents diffraction angle 2θ (°). The peak near 6.65° in the figure is a peak derived from the measuring instrument. The results of powder X-ray diffraction measurement of the crystal obtained in Example α44 are shown. The vertical axis represents diffraction intensity, and the horizontal axis represents diffraction angle 2θ (°). The results of simultaneous thermogravimetry, differential thermal analysis, and mass analysis of the crystals obtained in Example α44 are shown. The horizontal axis is temperature (°C), and the left vertical axis is the weight change (%) of the sample in the thermogravimetry analysis (these results are for reference only, as the sample was directly measured after filtration and drying through a mesh). The right vertical axis represents the heat flow (left) observed in the differential thermal analysis and the peak intensity (right) observed in the mass analysis. The results of powder X-ray diffraction measurement of the crystals obtained in Example α46 are shown. The vertical axis represents the diffraction intensity, and the horizontal axis represents the diffraction angle 2θ (°). The results of simultaneous thermogravimetry and differential thermal analysis of the crystals obtained in Example α46 are shown. The horizontal axis is temperature (°C), and the right vertical axis represents the weight change (%) of the sample in the thermogravimetry analysis. The left vertical axis represents the heat flow observed in the differential thermal analysis. The results of powder X-ray diffraction measurement of the crystals obtained in Example α46 are shown. The vertical axis represents the diffraction intensity, and the horizontal axis represents the diffraction angle 2θ (°). The results of simultaneous thermogravimetry and differential thermal analysis of the crystals obtained in Example α46 are shown. The horizontal axis represents temperature (°C), and the right vertical axis represents the weight change (%) of the sample in the thermogravimetry analysis. The left vertical axis represents the heat flow observed in the differential thermal analysis. １ The results of H-NMR measurement are shown. The vertical axis represents signal intensity, and the horizontal axis represents chemical shift δ (ppm). The results of powder X-ray diffraction measurement of the crystals obtained in Example α48 are shown. The vertical axis represents diffraction intensity, and the horizontal axis represents diffraction angle 2θ (°). The results of simultaneous thermogravimetric, differential thermal, and mass spectrometry measurements of the crystals obtained in Example α48 are shown. The horizontal axis represents temperature (°C), and the left vertical axis represents the weight change (%) of the sample in thermogravimetric analysis. The right vertical axis represents the heat flow (left) observed in differential thermal analysis and the peak intensity (right) observed in mass spectrometry. ... １1 shows the results of H-NMR measurement. The vertical axis represents signal intensity, and the horizontal axis represents chemical shift δ (ppm). The results of the dissolution test obtained in Example α53 are shown. The vertical axis represents the solution concentration of Compound 1 (μg / mL), and the horizontal axis represents the time (minutes) elapsed from the start of the dissolution test. This is a graph showing the cell viability calculated in a pemetrexed-induced cell damage test when Compound 1 was added.

[0010] The present invention will be described in detail below by showing definitions of symbols, terms, etc. used in this specification, and embodiments of the present invention.

[0011] In one embodiment, the present invention relates to a crystal of a compound represented by formula (1) or a salt thereof, or a solvate thereof. Specific examples of the crystal of this compound include a nonsolvate crystal of this compound (hereinafter also simply referred to as a "crystal of the compound"), a solvate crystal, a nonsolvate crystal of a salt of this compound (hereinafter also simply referred to as a "crystal of a salt of the compound"), or a solvate crystal. Preferred examples of the solvate crystal include hydrate crystals. For convenience, herein, "a compound represented by formula (1) or a salt thereof, or a solvate crystal of these" may be collectively referred to as "the API of the present invention."

[0012] The compounds described herein may be salts or solvates thereof. In addition, as used herein, the term "compounds or salts thereof, or solvates thereof" includes compounds (hereinafter also referred to as "free forms of compounds"), salts of compounds, solvates of compounds, and solvates of salts of compounds.

[0013] The compounds described herein, their salts, or solvates thereof include all stereoisomers thereof (e.g., enantiomers, diastereomers (including cis and trans geometric isomers)), racemates of such isomers, and other mixtures thereof. For example, the compounds of the present invention may have axial chirality, and the present invention includes each stereoisomer of such compounds and mixtures thereof.

[0014] sulfonates such as methanesulfonate (mesylate), ethanesulfonate (esylate), p-toluenesulfonate (tosylate), and benzenesulfonate (besylate); nitrate; carboxylates such as acetate, oxalate, maleate, fumarate, citrate, malate, malonate, gluconate, mandelate, salicylate, fluoroacetate, trifluoroacetate, camsylate, tartrate, propionate, and glutarate; or alkali metal salts such as lithium salt, sodium salt, potassium salt, cesium salt, and rubidium salt; alkaline earth metal salts such as magnesium salt, calcium salt, strontium salt, and barium salt; and ammonium salts such as ammonium salt, alkylammonium salt, dialkylammonium salt, trialkylammonium salt, and tetraalkylammonium salt. These salts are produced, for example, by contacting the compound represented by formula (1) with an acid or a base.

[0015] The salt of the compound represented by formula (1) is preferably hydrochloride, hydrobromide, hydroiodide, sulfate, mesylate, esylate, tosylate, besylate, phosphate, nitrate, acetate, oxalate, maleate, fumarate, citrate, malate, malonate, gluconate, mandelate, salicylate, fluoroacetate, trifluoroacetate, camsylate, tartrate, propionate, or glutarate, more preferably hydrochloride, hydrobromide, sulfate, mesylate, tosylate, or besylate, even more preferably hydrochloride, sulfate, or mesylate, and particularly preferably hydrochloride.

[0016] As used herein, the term "solvate" refers to a compound that forms a single molecular group together with a solvent, and is not particularly limited as long as it is a solvate formed with a solvent that is acceptable for ingestion accompanying pharmaceutical administration. Examples of solvates of the compound represented by formula (1) or a salt thereof include solvates with a single solvent such as water; alcohols (methanol, ethanol, 1-propanol, 2-propanol, 1-butanol, 2-butanol, benzyl alcohol, etc.), acetonitrile, esters (ethyl acetate, n-propyl acetate, n-butyl acetate, etc.), ethers (tetrahydrofuran, 1,4-dioxane, t-butyl methyl ether, etc.), ketones (acetone, 2-butanone, methyl isobutyl ketone, etc.), halogenated solvents (dichloromethane, etc.), aromatic hydrocarbon solvents (anisole, toluene, chlorobenzene, etc.), amide solvents (formamide, dimethylformamide, dimethylacetamide, etc.), and dimethyl sulfoxide, as well as solvates formed with multiple solvents per molecule of the compound, or solvates formed with multiple types of solvents per molecule of the compound. If the solvent is water, it is called a hydrate.

[0017] The solvate of the compound represented by formula (1) or a salt thereof is preferably a solvate of the compound represented by formula (1) or a salt of the compound with a solvent selected from the group consisting of water, methanol, ethanol, 1-propanol, 2-propanol, 1-butanol, 2-butanol, benzyl alcohol, acetonitrile, ethyl acetate, n-propyl acetate, n-butyl acetate, tetrahydrofuran, 1,4-dioxane, t-butyl methyl ether, dichloromethane, dimethyl sulfoxide, acetic acid, formic acid, acetone, 2-butanone, methyl isobutyl ketone, anisole, toluene, chlorobenzene, formamide, dimethylformamide, and dimethylacetamide, and more preferably a solvate of the compound represented by formula (1) or a salt of the compound with water, ethanol, and a solvate of the compound represented by formula (1) or a salt of the compound with a solvent selected from the group consisting of water, ethanol, 2-propanol, 2-butanol, benzyl alcohol, tetrahydrofuran, dimethyl sulfoxide, acetic acid, formic acid, acetone, formamide, dimethylformamide, and dimethylacetamide, more preferably a solvate of the compound represented by formula (1) or a salt of the compound with a solvent selected from the group consisting of water, ethanol, 2-propanol, 2-butanol, dimethyl sulfoxide, acetic acid, and formic acid, even more preferably a solvate of the compound represented by formula (1) or a salt of the compound with a solvent selected from the group consisting of water and ethanol, and particularly preferably a hydrate of the compound represented by formula (1) or a hydrate of a salt of the compound.

[0018] In one embodiment, the crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal of a solvate of at least one selected from the group consisting of the compound represented by formula (1) and hydrochloride, hydrobromide, sulfate, mesylate, tosylate, and besylate of the compound, and a solvent selected from the group consisting of water, ethanol, 2-propanol, 2-butanol, dimethyl sulfoxide, acetic acid, and formic acid.

[0019] In one embodiment, the crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal of a hydrate of the hydrochloride salt of the compound represented by formula (1).

[0020] In one embodiment, the crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal of the monohydrate of the hydrochloride salt of the compound represented by formula (1).

[0021] In one embodiment, the crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal of a hydrate of the compound represented by formula (1).

[0022] In one embodiment, the crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal of the hydrochloride salt of the compound represented by formula (1).

[0023] In one embodiment, the crystals of the compound represented by formula (1) or a salt thereof, or a solvate thereof are crystals of a dihydrate of the 0.5 hydrochloride salt of the compound represented by formula (1).

[0024] In one embodiment, the crystals of the compound represented by formula (1) or a salt thereof, or a solvate thereof are crystals of a hydrate of the sesquihydrochloride salt of the compound represented by formula (1).

[0025] In one embodiment, the crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal of a hydrate of the sulfate salt of the compound represented by formula (1).

[0026] In one embodiment, the crystals of the compound represented by formula (1) or a salt thereof, or a solvate thereof are crystals of a hydrate of the 0.5 mesylate salt of the compound represented by formula (1).

[0027] In one embodiment, the crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal of a hydrate of the mesylate salt of the compound represented by formula (1).

[0028] In one embodiment, the crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal of a hydrate of the tosylate salt of the compound represented by formula (1).

[0029] The diffraction angle 2θ in powder X-ray diffraction is the diffraction peak measured using CuKα or CuKα1 radiation. In this specification, "powder X-ray diffraction" refers to a numerical value that can be determined using X-ray diffraction phenomena and is used for the identification and structural analysis of crystalline substances, and is a value specific to any crystal. This value is usually expressed as one or more 2θ values. Those skilled in the art can measure this using a commercially available powder X-ray diffraction measurement instrument and following its instruction manual. More specifically, the 2θ value can be measured by irradiating the sample to be measured with CuKα or CuKα1 X-rays and measuring the diffracted X-rays relative to the incident X-rays. For example, this can be performed according to a conventional method such as the "powder X-ray diffraction measurement method" described in the Japanese Pharmacopoeia (18th edition).

[0030] Peak values ​​(2θ values) in powder X-ray diffraction spectra may have some error depending on the measuring instrument or measurement conditions, such as peak reading conditions. In this specification, peak values ​​may have a measurement error of about ±0.2°. The Japanese Pharmacopoeia (18th edition) states that for the same crystalline form, the diffraction angle 2θ typically coincides within a range of ±0.2°. Therefore, the present invention includes not only crystals in which the diffraction angles of peaks in powder X-ray diffraction are completely identical, but also crystals in which the diffraction angles of peaks coincide with an error of about ±0.2°. When the diffraction angle 2θ is expressed, if "(±0.2°)" is added at the end of the listed diffraction angle 2θ, this means that a range of ±0.2° is allowed for all listed diffraction angles 2θ.

[0031] In one embodiment, a crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal comprising at least one, three, five, seven, or nine peaks selected from the group consisting of 6.82°, 8.98°, 10.35°, 10.89°, 11.88°, 12.09°, 12.36°, 12.72°, 13.46°, and 15.27° (±0.2°) as diffraction angles (2θ values) in powder X-ray diffraction at 23° C. The crystal comprising the above peaks is preferably a crystal of the compound represented by formula (1).

[0032] In one embodiment, a crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal having peaks at diffraction angles (2θ values) of 6.82°, 8.98°, 10.35°, 10.89°, 11.88°, 12.09°, 12.36°, 12.72°, 13.46°, and 15.27° (±0.2°) in powder X-ray diffraction at 23° C. The crystal having the above peaks is preferably a crystal of the compound represented by formula (1).

[0033] In one embodiment, a crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal comprising at least one, three, five, seven, or nine peaks selected from the group consisting of 5.75°, 6.30°, 7.93°, 10.14°, 10.62°, 11.52°, 12.63°, 15.80°, 18.55°, and 23.72° (±0.2°) as diffraction angles (2θ values) in powder X-ray diffraction at 23° C. The crystal comprising the above peaks is preferably a crystal of a hydrate of the compound represented by formula (1).

[0034] In one embodiment, a crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal having peaks at diffraction angles (2θ values) of 5.75°, 6.30°, 7.93°, 10.14°, 10.62°, 11.52°, 12.63°, 15.80°, 18.55°, and 23.72° (±0.2°) in powder X-ray diffraction at 23° C. The crystal having the above peaks is preferably a crystal of a hydrate of the compound represented by formula (1).

[0035] In one embodiment, a crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal comprising at least one, three, five, seven, or nine peaks selected from the group consisting of 7.01°, 8.74°, 9.14°, 10.17°, 10.82°, 11.78°, 13.31°, 13.93°, 17.00°, and 22.77° (±0.2°) as diffraction angles (2θ values) in powder X-ray diffraction at 23° C. The crystal comprising the above peaks is preferably a crystal of a hydrate of the compound represented by formula (1).

[0036] In one embodiment, a crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal having peaks at diffraction angles (2θ values) of 7.01°, 8.74°, 9.14°, 10.17°, 10.82°, 11.78°, 13.31°, 13.93°, 17.00°, and 22.77° (±0.2°) in powder X-ray diffraction at 23° C. The crystal having the above peaks is preferably a crystal of a hydrate of the compound represented by formula (1).

[0037] In one embodiment, a crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal comprising at least one, three, five, seven, or nine peaks selected from the group consisting of 8.94°, 11.49°, 12.65°, 12.94°, 16.03°, 16.34°, 17.61°, 19.11°, 19.68°, and 20.64° (±0.2°) as diffraction angles (2θ values) in powder X-ray diffraction at 100° C. The crystal comprising the above peaks is preferably a crystal of the hydrochloride salt of the compound represented by formula (1).

[0038] In one embodiment, a crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal having peaks at diffraction angles (2θ values) of 8.94°, 11.49°, 12.65°, 12.94°, 16.03°, 16.34°, 17.61°, 19.11°, 19.68°, and 20.64° (±0.2°) in powder X-ray diffraction at 100° C. The crystal having the above peaks is preferably a crystal of the hydrochloride salt of the compound represented by formula (1).

[0039] In one embodiment, a crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal comprising at least one, three, five, seven, or nine peaks selected from the group consisting of 9.02°, 11.47°, 12.71°, 12.98°, 16.50°, 17.83°, 19.24°, 19.68°, 20.80°, and 22.29° (±0.2) as diffraction angles (2θ values) in powder X-ray diffraction at 35° C. The crystal comprising the above peaks is preferably a crystal of the hydrochloride salt of the compound represented by formula (1).

[0040] In one embodiment, a crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal having peaks at diffraction angles (2θ values) of 9.02°, 11.47°, 12.71°, 12.98°, 16.50°, 17.83°, 19.24°, 19.68°, 20.80°, and 22.29° (±0.2) in powder X-ray diffraction at 35° C. The crystal having the above peaks is preferably a crystal of the hydrochloride salt of the compound represented by formula (1).

[0041] In one embodiment, a crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof, is a crystal comprising at least one, three, five, seven, or nine peaks selected from the group consisting of 6.59°, 9.19°, 9.70°, 12.70°, 13.15°, 13.29°, 13.80°, 14.93°, 15.80°, and 16.45° (±0.2) as diffraction angles (2θ values) in powder X-ray diffraction at 23° C. The crystal comprising the above peaks is preferably a crystal of a hydrate of the hydrochloride of the compound represented by formula (1), more preferably a crystal of a monohydrate of the hydrochloride of the compound represented by formula (1).

[0042] In one embodiment, a crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal having peaks at diffraction angles (2θ values) of 6.59°, 9.19°, 9.70°, 12.70°, 13.15°, 13.29°, 13.80°, 14.93°, 15.80°, and 16.45° (±0.2) in powder X-ray diffraction at 23° C. The crystal having the above peaks is preferably a crystal of a hydrate of the hydrochloride of the compound represented by formula (1), more preferably a crystal of a monohydrate of the hydrochloride of the compound represented by formula (1).

[0043] In one embodiment, a crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof, is a crystal containing at least one, three, five, seven, or nine peaks selected from the group consisting of 7.07°, 9.24°, 12.06°, 14.37°, 15.24°, 15.84°, 17.86°, 18.30°, 18.52°, and 19.31° (±0.2) as diffraction angles (2θ values) in powder X-ray diffraction at 23° C. The crystal containing the above peaks is preferably a crystal of a hydrate of the hydrochloride salt of the compound represented by formula (1), more preferably a crystal of a dihydrate of the 0.5 hydrochloride salt of the compound represented by formula (1).

[0044] In one embodiment, a crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal having peaks at diffraction angles (2θ values) of 7.07°, 9.24°, 12.06°, 14.37°, 15.24°, 15.84°, 17.86°, 18.30°, 18.52°, and 19.31° (±0.2) in powder X-ray diffraction at 23° C. The crystal having the above peaks is preferably a crystal of a hydrate of the hydrochloride salt of the compound represented by formula (1), more preferably a crystal of a dihydrate of the 0.5 hydrochloride salt of the compound represented by formula (1).

[0045] In one embodiment, a crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof, is a crystal comprising at least one, three, five, seven, or nine peaks selected from the group consisting of 10.24°, 10.60°, 10.85°, 12.75°, 12.91°, 13.12°, 13.96°, 14.76°, 15.93°, and 16.84° (±0.2) as diffraction angles (2θ values) in powder X-ray diffraction at 23° C. The crystal comprising the above peaks is preferably a crystal of a hydrate of the hydrochloride salt of the compound represented by formula (1), more preferably a crystal of a hydrate of the sesquihydrochloride salt of the compound represented by formula (1).

[0046] In one embodiment, a crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal having peaks at diffraction angles (2θ values) of 10.24°, 10.60°, 10.85°, 12.75°, 12.91°, 13.12°, 13.96°, 14.76°, 15.93°, and 16.84° (±0.2) in powder X-ray diffraction at 23° C. The crystal having the above peaks is preferably a crystal of a hydrate of the hydrochloride salt of the compound represented by formula (1), more preferably a crystal of a hydrate of the sesquihydrochloride salt of the compound represented by formula (1).

[0047] In one embodiment, a crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal comprising at least one, three, five, seven, or nine peaks selected from the group consisting of 6.95°, 7.50°, 9.05°, 11.28°, 14.32°, 15.13°, 15.72°, 17.89°, 19.86°, and 20.69° (±0.2°) as diffraction angles (2θ values) in powder X-ray diffraction at 23° C. The crystal comprising the above peaks is preferably a crystal of a hydrate of the sulfate salt of the compound represented by formula (1).

[0048] In one embodiment, a crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal having peaks at diffraction angles (2θ values) of 6.95°, 7.50°, 9.05°, 11.28°, 14.32°, 15.13°, 15.72°, 17.89°, 19.86°, and 20.69° (±0.2°) in powder X-ray diffraction at 23° C. The crystal having the above peaks is preferably a crystal of a hydrate of the sulfate salt of the compound represented by formula (1).

[0049] In one embodiment, a crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal comprising at least one, three, five, seven, or nine peaks selected from the group consisting of 5.56°, 7.53°, 10.30°, 10.52°, 11.16°, 11.61°, 12.96°, 15.70°, 16.88°, and 17.58° (±0.2°) as diffraction angles (2θ values) in powder X-ray diffraction at 23° C. The crystal comprising the above peaks is preferably a crystal of a hydrate of the mesylate of the compound represented by formula (1), more preferably a crystal of a hydrate of the 0.5 mesylate of the compound represented by formula (1).

[0050] In one embodiment, a crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal having peaks at diffraction angles (2θ values) of 5.56°, 7.53°, 10.30°, 10.52°, 11.16°, 11.61°, 12.96°, 15.70°, 16.88°, and 17.58° (±0.2°) in powder X-ray diffraction at 23° C. The crystal having the above peaks is preferably a crystal of a hydrate of the mesylate of the compound represented by formula (1), more preferably a crystal of a hydrate of the 0.5 mesylate of the compound represented by formula (1).

[0051] In one embodiment, a crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal comprising at least one, three, five, seven, or nine peaks selected from the group consisting of 5.75°, 8.37°, 9.38°, 10.89°, 13.14°, 16.81°, 17.36°, 17.79°, 18.37°, and 18.78° (±0.2°) as diffraction angles (2θ values) in powder X-ray diffraction at 23° C. The crystal comprising the above peaks is preferably a crystal of a hydrate of the mesylate of the compound represented by formula (1).

[0052] In one embodiment, a crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal having peaks at diffraction angles (2θ values) of 5.75°, 8.37°, 9.38°, 10.89°, 13.14°, 16.81°, 17.36°, 17.79°, 18.37°, and 18.78° (±0.2°) in powder X-ray diffraction at 23° C. The crystal having the above peaks is preferably a crystal of a hydrate of the mesylate of the compound represented by formula (1).

[0053] In one embodiment, a crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal comprising at least one, three, five, seven, or nine peaks selected from the group consisting of 6.00°, 8.06°, 8.61°, 10.06°, 12.11°, 12.78°, 15.01°, 16.46°, 17.64°, and 18.09° (±0.2°) as diffraction angles (2θ values) in powder X-ray diffraction at 23° C. The crystal comprising the above peaks is preferably a crystal of a hydrate of the tosylate of the compound represented by formula (1).

[0054] In one embodiment, a crystal of the compound represented by formula (1) or a salt thereof, or a solvate thereof is a crystal having peaks at diffraction angles (2θ values) of 6.00°, 8.06°, 8.61°, 10.06°, 12.11°, 12.78°, 15.01°, 16.46°, 17.64°, and 18.09° (±0.2°) in powder X-ray diffraction at 23° C. The crystal having the above peaks is preferably a crystal of a hydrate of the tosylate of the compound represented by formula (1).

[0055] In this specification, the term "to" indicating a range of values ​​includes both ends of the range. For example, "A to B" means a range of values ​​equal to or greater than A and equal to or less than B.

[0056] As used herein, the term "about" when used in conjunction with a numerical value means a range of values ​​of plus and minus 10% of that numerical value.

[0057] As used herein, the term "and / or" includes any combination of "and" and "or." Specifically, for example, "A, B, and / or C" includes the following seven variations: (i) A, (ii) B, (iii) C, (iv) A and B, (v) A and C, (vi) B and C, and (vii) A, B, and C.

[0058] The compound represented by formula (1) can be synthesized by various methods, and may be synthesized by a method known to those skilled in the art. The compound represented by formula (1) can be synthesized, for example, by the method described in Example 1 below.

[0059] <Production Method> Crystals of the compound represented by formula (1) or a salt thereof, or a solvate thereof can be produced, for example, by the method shown below. Crystals of the compound represented by formula (1) or a salt thereof, or a solvate thereof can be obtained by adding a solvent suitable for crystallization to the compound, optionally adding seed crystals, and stirring as necessary. The solvent added during crystallization is not particularly limited as long as it is a solvent that allows the compound to form crystals, but a solvent that allows an operation to reduce the solubility of the compound in a solution in which the compound is dissolved is preferred. For example, when crystallization is possible by reducing the solubility of the compound by adding a poor solvent or cooling the solution, a solvent that allows such an operation is exemplified. Furthermore, when crystals of the compound can be obtained by maintaining crude crystals of the compound in a suspension state for a desired period of time, a solvent that allows such an operation can be used for crystallization. Specific examples of the solvent added during crystallization include hydrochloric acid, formic acid, acetic acid, acetone, water, dimethyl sulfoxide (DMSO), dimethylformamide (DMF), acetonitrile, tetrahydrofuran, methyl ethyl ketone, methyl isobutyl ketone, ethyl acetate, heptane, polyethylene glycol (PEG), 2-propanol, methanol, ethanol, and mixed solvents thereof.

[0060] <Pharmaceutical Composition> One aspect of the present invention provides a pharmaceutical composition containing a crystalline compound represented by formula (1) or a salt thereof, or a solvate thereof. In this specification, a pharmaceutical composition containing a crystalline compound represented by formula (1) or a salt thereof, or a solvate thereof, may be referred to as the "composition of the present invention." The pharmaceutical composition of this embodiment can be formulated by known methods by incorporating a pharmaceutically acceptable carrier in addition to the crystalline compound represented by formula (1) or a salt thereof, or a solvate thereof. For formulation, commonly used excipients, binders, lubricants, colorants, flavorings, and, if necessary, stabilizers, emulsifiers, absorption enhancers, surfactants, pH adjusters, preservatives, antioxidants, etc. may be used. The composition is formulated by conventional methods by blending ingredients commonly used as raw materials for pharmaceutical formulations. For formulation, the active ingredients used in pharmaceuticals can be processed by known methods into the optimal shape or properties, i.e., dosage form, suited to the method and purpose of use. Examples of commonly used dosage forms include liquid pharmaceutical preparations (liquid preparations) such as injections, suspensions, emulsions, and eye drops, and solid pharmaceutical preparations (solid preparations) such as tablets, powders, fine granules, granules, coated tablets, capsules, dry syrups, lozenges, and suppositories, but are not limited to these.

[0061] For example, to prepare a liquid formulation, a pharmaceutically acceptable carrier or vehicle, specifically, pharmaceutically acceptable additives commonly used in the field of pharmaceutical formulations, such as sterile water, physiological saline, vegetable oil, emulsifier, suspending agent, surfactant, stabilizer, flavoring agent, excipient, vehicle, preservative, binder, etc., are added in appropriate combination to a crystal of the compound represented by formula (1) or its salt, or a solvate thereof, and then the mixture is mixed to form a unit dose required for generally accepted pharmaceutical practice. Alternatively, a solid formulation prepared for liquid formulation can be dissolved as needed by adding an appropriate solvent, such as sterile water or physiological saline, before administration, and then used for administration.

[0062] Such liquid preparations can also be used parenterally, for example, in the form of injections of sterile solutions or suspensions in water or other pharmaceutically acceptable liquids. For example, they can be formulated by appropriately combining them with pharmaceutically acceptable carriers or vehicles, specifically, sterile water, physiological saline, vegetable oils, emulsifiers, suspending agents, surfactants, stabilizers, flavoring agents, excipients, vehicles, preservatives, binders, etc., and blending them in unit dosage forms required for generally accepted pharmaceutical practice. Specific examples of carriers include lactose, starch, carmellose calcium, carmellose sodium, hydroxypropyl cellulose, polyvinyl acetal diethylaminoacetate, polyvinylpyrrolidone, gelatin, medium-chain fatty acid triglycerides, polyoxyethylene hydrogenated castor oil 60, sucrose, carboxymethylcellulose, cornstarch, inorganic salts, etc. The amount of active ingredient in these preparations is such that an appropriate volume within the specified range can be obtained. Sterile compositions for injection can be formulated according to conventional pharmaceutical practice using a vehicle such as distilled water for injection.

[0063] Examples of aqueous solutions for injection include isotonic solutions containing physiological saline or other adjuvants, such as D-sorbitol, D-mannose, and sodium chloride, and may be used in combination with an appropriate solubilizing agent, such as alcohol, specifically ethanol, polyalcohols such as propylene glycol and polyethylene glycol, or nonionic surfactants such as Polysorbate 80 (registered trademark) and HCO-50.

[0064] Examples of oily liquids include sesame oil and soybean oil, and they may be used in combination with solubilizing agents such as benzyl benzoate and benzyl alcohol. They may also contain buffers such as phosphate buffer and sodium acetate buffer, soothing agents such as procaine hydrochloride, stabilizers such as benzyl alcohol, phenol, and antioxidants. The prepared injection solution is usually filled into suitable ampoules.

[0065] For example, to produce a solid preparation, an excipient and, if necessary, pharmaceutically acceptable additives commonly used in the field of pharmaceutical preparations such as binders, disintegrants, lubricants, colorants, and flavoring agents are added in appropriate combination to crystals of the compound represented by formula (1) or a salt thereof, or a solvate thereof, and then the mixture is formed into tablets, powders, fine granules, granules, coated tablets, capsules, dry syrups, troches, suppositories, etc. by conventional methods.

[0066] Examples of pharmaceutically acceptable additives used in such solid preparations include animal and vegetable oils such as soybean oil, beef tallow, and synthetic glycerides; hydrocarbons such as liquid paraffin, squalane, and solid paraffin; ester oils such as octyldodecyl myristate and isopropyl myristate; higher alcohols such as cetostearyl alcohol and behenyl alcohol; silicone resins; silicone oils; surfactants such as polyoxyethylene fatty acid esters, sorbitan fatty acid esters, glycerin fatty acid esters, polyoxyethylene sorbitan fatty acid esters, polyoxyethylene hydrogenated castor oil, and polyoxyethylene polyoxypropylene block copolymers; water-soluble polymers such as hydroxyethyl cellulose, polyacrylic acid, carboxyvinyl polymers, polyethylene glycol, polyvinylpyrrolidone, and methylcellulose; lower alcohols such as ethanol and isopropanol; polyhydric alcohols such as glycerin, propylene glycol, dipropylene glycol, and sorbitol; sugars such as lactose, lactose hydrate, fructose, and sucrose; inorganic powders such as silicic anhydride, aluminum magnesium silicate, and aluminum silicate; and purified water.

[0067] Examples of excipients include sugars (e.g., lactose, lactose hydrate, fructose, sucrose, etc.), starches (corn starch, potato starch, wheat starch, rice starch, partially pregelatinized starch, pregelatinized starch, etc.), inorganic salts (e.g., calcium silicate, anhydrous calcium hydrogen phosphate, precipitated calcium carbonate, etc.), and the like.

[0068] Examples of binders include polyvinyl alcohol, polyvinyl ether, methyl cellulose, ethyl cellulose, gum arabic, tragacanth, gelatin, shellac, hydroxypropyl cellulose, polyvinylpyrrolidone, polypropylene glycol-polyoxyethylene block polymers, and the like.

[0069] Examples of disintegrants include carmellose sodium, hydroxypropyl cellulose, carmellose, carmellose calcium, methylcellulose, povidone, and polysorbate.

[0070] Examples of lubricants include calcium stearate, talc, sucrose fatty acid esters, sodium stearyl fumarate, and hardened oils.

[0071] Coloring agents that are permitted to be added to pharmaceuticals are used, and flavoring agents include cocoa powder, peppermint, aromatic powder, peppermint oil, borneol, cinnamon powder, etc.

[0072] The pharmaceutical composition according to this embodiment can be administered orally or parenterally to a subject, and the administration method can be selected appropriately depending on the age and symptoms of the patient. Specific examples of parenteral administration include injections, intranasal administrations, pulmonary administrations, and transdermal administrations. Examples of injections include intravenous injections, intramuscular injections, intraperitoneal injections, and subcutaneous injections, which can be administered systemically or locally.

[0073] The dosage of the pharmaceutical composition according to this embodiment is preferably 0.001 to 50 mg per kg of body weight of the subject per day (0.001 to 50 mg / kg / day), more preferably 0.001 to 20 mg / kg / day, and even more preferably 0.002 to 10 mg / kg / day. The dosage and administration method vary depending on the body weight, age, symptoms, etc. of the patient, but can be appropriately selected by those skilled in the art.

[0074] In some embodiments, the compounds of the present invention, or salts thereof, or crystals of solvates thereof, can be used to inhibit MYT1.

[0075] [Treatment or Prevention of Cancer] One embodiment of the present invention is a method for preventing and / or treating cancer, which comprises administering to a subject a crystalline form of a compound represented by formula (1) or a salt thereof, or a solvate thereof.

[0076] In certain embodiments, the method comprises administering a composition of the present invention in combination with a chemotherapeutic agent (concomitant administration) to treat or prevent cancer in a patient who has been detected as having a positive RB1 gene mutation or reduced expression of the RB1 gene or protein.

[0077] In certain embodiments, the method comprises administering a composition of the present invention in combination with a chemotherapeutic agent (concomitant administration) to treat or prevent cancer in a cancer patient who is positive for an RB1 gene mutation or has reduced expression of the RB1 gene or protein.

[0078] "Cancer treatment" in the present invention means a reduction in the number of cancer cells in an individual, inhibition of cancer cell proliferation, reduction in tumor volume, reduction in tumor weight, inhibition of cancer cell metastasis, or amelioration of various symptoms caused by cancer, or a combination thereof. Furthermore, "cancer prevention" in the present invention means preventing the development of new cancer cells, preventing an increase in the number of cancer cells due to the re-proliferation of reduced cancer cells, preventing the regrowth of cancer cells whose proliferation has been inhibited, preventing a re-increase in the volume or weight of reduced tumors, or a combination thereof.

[0079] RB1 (retinoblastoma gene, also known as Rb or RB) is a gene encoding the RB1 protein, a representative cell cycle regulator, and is involved in the G1 / S checkpoint. The RB1 protein (also known as RB1 or pRb) forms a complex with E2F, a transcription factor responsible for inducing the expression of genes involved in the transition from the G1 phase to the S phase of the cell cycle, thereby suppressing the activity of E2F. When E2F activity is suppressed, the transition from the G1 phase to the S phase is inhibited.

[0080] As used herein, "positive for RB1 gene mutation" means that, when the nucleotide sequence corresponding to the RB1 gene of interest is analyzed, some mutation (e.g., a mutation resulting in the insertion, substitution, deletion, and / or addition of at least one amino acid residue relative to the wild-type RB1 protein) is found in the nucleotide sequence compared to the nucleotide sequence of the wild-type RB1 gene, or, if a mutation in the nucleotide sequence of the RB1 gene is reflected in a base change in a transcription product or an amino acid change in a translation product, the change is detected in the transcription product or translation product. In certain embodiments, a positive RB1 gene mutation is detected in a biological sample (e.g., cancer cells) derived from a cancer patient. As used herein, "detecting a mutation" generally means detecting a mutation in genomic DNA, but also includes detecting the change in the transcription product or translation product (i.e., indirect detection) when the mutation in the genomic DNA is reflected in a base change in a transcription product or an amino acid change in a translation product. A preferred embodiment of the method herein is a method of detecting a mutation by directly determining the nucleotide sequence of the RB1 gene region in a cancer cell. There are no particular limitations on the method for detecting a positive RB1 gene mutation, and for example, confirmation and determination can be performed by NGS (next generation sequencer).

[0081] In the present invention, the term "RB1 gene region" refers to a certain region on genomic DNA that contains the RB1 gene. This region independently includes, in addition to the translated region, untranslated regions such as expression control regions of each gene (e.g., promoter regions and enhancer regions) and the 3'-terminal untranslated region of each gene. In this method, a DNA sample is first prepared from a biological sample. Examples of DNA samples include genomic DNA samples and cDNA samples prepared by reverse transcription from RNA.

[0082] In this embodiment, DNA containing the RB1 gene region is then isolated, and the nucleotide sequence of the isolated DNA is determined. The DNA can be isolated, for example, by PCR using genomic DNA or RNA as a template, using a pair of oligonucleotide primers designed to flank all or part of the RB1 gene region. The nucleotide sequence of the isolated DNA can be determined by methods known to those skilled in the art, such as the Maxam-Gilbert method or the Sanger method, and a next-generation sequencer or the like capable of rapid and comprehensive analysis of gene nucleotide sequences can also be used.

[0083] By comparing the determined DNA or cDNA base sequence with a control (for example, if the biological sample is derived from a cancer patient, with the DNA or cDNA base sequence derived from non-cancerous tissue of the same patient or with a publicly known database), it is possible to determine whether or not there is a mutation in the RB1 gene region in the cancer cells of the biological sample.

[0084] Mutations in the RB1 gene region can be detected by various methods that allow detection of mutations, in addition to direct determination of the base sequence of DNA or cDNA.

[0085] If the mutation involves an amino acid change in the RB1 protein, the sample prepared from the biological sample may be a protein. In such cases, the mutation can be detected by a method using a molecule (e.g., an antibody) that specifically binds to the site where the amino acid change occurs due to the mutation, peptide mass fingerprinting (PMF), protein sequencer (Edman degradation), or the like.

[0086] As used herein, "decreased expression of the RB1 gene or protein" means that, when the RB1 gene or protein of a subject is analyzed, the expression level of the RB1 gene or protein is lower than that of a control (e.g., expression level in a healthy subject or in non-cancerous tissue of the same patient). In a specific embodiment, the decreased expression level of the RB1 gene or protein is detected in a biological sample (e.g., cancer cells) derived from a cancer patient.

[0087] The method for detecting decreased expression of the RB1 gene is not particularly limited, and examples include a method in which the expression level of RB1 is detected at the transcription level or translation level and compared with the control. In a method for detecting the expression level of the RB1 gene at the transcription level, RNA or cDNA is first prepared from a biological sample. The method for extracting RNA from a biological sample and the method for preparing cDNA from the extracted RNA are not particularly limited, and known methods can be appropriately selected and used. Examples include extraction methods using phenol and chaotropic salts (more specifically, extraction methods using commercially available kits such as Trizol (Invitrogen) and Isogen (Wako Pure Chemical Industries)), and methods using other commercially available kits (RNAPrep Total RNA Extraction Kit (Beckman Coulter), RNeasy Mini (QIAGEN), RNA Extraction Kit (Pharmacia Biotech), etc.). Furthermore, the reverse transcriptase used to prepare cDNA from the extracted RNA is not particularly limited, and examples thereof include reverse transcriptases derived from retroviruses such as RAV (Rous associated virus) and AMV (Avian myeloblastosis virus), and reverse transcriptases derived from mouse retroviruses such as MMLV (Moloney murine leukemia virus).

[0088] The oligonucleotide primers or oligonucleotide probes are then used in an amplification reaction or hybridization reaction, respectively, to detect the amplified product or hybrid product. Examples of such methods include RT-PCR, Northern blotting, dot blotting, DNA arrays, in situ hybridization, RNase protection assays, and mRNA-seq. Those skilled in the art can routinely design oligonucleotide primers or oligonucleotide probes suitable for each method based on the nucleotide sequence of RB1 cDNA.

[0089] It is known in the art that promoter hypermethylation is one of the factors that contribute to decreased gene expression. Therefore, in detecting whether or not RB1 function is inhibited, it is conceivable to use methylation of the RB1 gene promoter as an indicator. Promoter methylation can be detected by known methods, such as a method in which a change in the base sequence after bisulfite treatment, which has the activity of converting methylated cytosine to uracil, is directly detected by base sequencing, or a method in which a restriction endonuclease that can recognize (cleave) the base sequence before bisulfite treatment but cannot recognize (cleave) the base sequence after bisulfite treatment.

[0090] The method for detecting decreased expression of RB1 protein is not particularly limited, and can be confirmed and determined, for example, by IHC (immunohistochemical staining) using an antibody specific to RB1 protein. In antibody-based protein detection methods, a protein sample is first prepared from a biological sample. Then, RB1 protein is detected by an antigen-antibody reaction using an antibody specific to RB1 protein. When the antibody specific to RB1 protein is labeled, RB1 protein can be detected directly. However, when the antibody is unlabeled, RB1 protein can be detected indirectly by further reacting it with a labeled molecule that recognizes the antibody (e.g., a secondary antibody or protein A), utilizing the label of the molecule. Examples of such methods include immunohistochemistry (immunostaining), Western blotting, ELISA, flow cytometry, imaging cytometry, radioimmunoassay, immunoprecipitation, and analysis using an antibody array. This method also has the advantage of simultaneously obtaining additional information, such as the morphology and distribution of cancer cells in tissue, through immunohistochemistry.

[0091] As used herein, the amplification of the copy number of the CCNE1 gene can be determined in a diagnostic or prognostic assay by assessing the copy number of the CCNE1 gene using a biological sample derived from a cancer patient (e.g., by next-generation sequencing, digital PCR, array CGH, or FISH).

[0092] As used herein, a "patient" may be a mouse, rat, guinea pig, monkey, dog, sheep, horse, or human. A "patient" is preferably a human. In the present invention, a "cancer patient" refers not only to a person currently suffering from cancer, but also to a person suspected of suffering from cancer. The composition of the present invention can be suitably used in humans.

[0093] As used herein, the term "biological sample derived from a cancer patient" is not particularly limited as long as it is a biological sample capable of detecting the presence or absence of an RB1 gene mutation or reduced expression of the RB1 gene or protein, but is preferably a sample such as a cancer biopsy specimen, blood, urine, body cavity fluid, or tumor cell-derived circulating tumor DNA (ctDNA). Protein extracts or nucleic acid extracts (e.g., mRNA extracts, cDNA preparations or cRNA preparations prepared from mRNA extracts) obtained from the specimens may also be used. As used herein, "biological sample" includes samples derived from cancer patients and samples derived from cancer cell cultures.

[0094] The RB1 gene mutation may include a mutation that results in the insertion, deletion, or addition of at least one amino acid residue relative to the wild-type RB1 protein, or a mutation that results in the substitution of an existing amino acid residue. The RB1 gene mutation may also be a nonsense mutation, a frameshift mutation, a splice site mutation, a heterozygous deletion, or a homozygous deletion. Preferably, the RB1 gene mutation is a mutation that reduces the function of RB1. "Mutations that reduce the function of RB1" can be confirmed, for example, via the Internet at: https: / / www.oncokb.org / gene / RB1 [searched February 20, 2023].

[0095] A typical DNA (cDNA) nucleotide sequence of a human wild-type RB1 gene is shown in SEQ ID NO: 1 (NCBI Reference No.: NM_000321.3), and a typical amino acid sequence of a human wild-type RB1 protein is shown in SEQ ID NO: 2 (NCBI Reference No.: NP_000312.2). In the case of a human RB1 gene, an RB1 gene mutation refers to a nucleotide sequence that is different from the genomic sequence of human RB1 described at positions 48,303,751 to 48,481,890 in NCBI Reference No. NC_13.11, a nucleotide sequence that is different from the nucleotide sequence of human RB1 described at positions 4921 to 5161 in NCBI Reference No. NG_9009.1, or a mutation that results in an amino acid sequence that is different from the amino acid sequence of the human RB1 protein described as SEQ ID NO: 2, and may be a mutation that results in at least one of the following (1) to (5): Even in RB1 without mutation, individual differences in the sequence may occur due to polymorphism, etc. (1) The codon corresponding to the serine residue (S) at position 82 in the amino acid sequence of SEQ ID NO: 2 is replaced with a stop codon, (2) The codon corresponding to the arginine residue (R) at position 467 in the amino acid sequence of SEQ ID NO: 2 is replaced with a stop codon, (3) At least one base is inserted or deleted in the codon corresponding to the amino acid residue at position 182 in the amino acid sequence of SEQ ID NO: 2, forming a new reading frame starting with an isoleucine residue (I), and the third reading frame therefrom is a stop codon, (4) The glutamic acid residue (E) at position 837 in the amino acid sequence of SEQ ID NO: 2 is replaced with a lysine residue (K), and a portion of the RB1 gene is homozygously deleted, or (5) The glycine residue (G) at position 449 in the amino acid sequence of SEQ ID NO: 2 is replaced with a glutamic acid residue (E), and a portion of the RB1 gene is homozygously deleted.

[0096] As used herein, decreased expression of the RB1 gene or protein includes decreased gene expression due to methylation of the RB1 gene or via microRNA.

[0097] The composition of the present invention can be used to treat or prevent cancer, and is particularly suitable for treating or preventing cancer in cancer patients who have been detected as positive for RB1 gene mutation or in whom reduced expression of the RB1 gene or protein has been detected, or in cancer patients who have been detected as positive for RB1 gene mutation or in whom reduced expression of the RB1 gene or protein has occurred. The composition of the present invention can be used alone or in combination with a chemotherapeutic agent.

[0098] In a particular embodiment, the cancer is lung cancer. The compositions of the present invention are particularly suitable for the treatment or prevention of cancer, and particularly for the treatment or prevention of lung cancer.

[0099] The compositions of the present invention may be used in combination with chemotherapeutic agents. In one embodiment, the compositions of the present invention are administered simultaneously with or separately from the chemotherapeutic agents. In another embodiment, the compositions of the present invention are administered as a combination drug with the chemotherapeutic agents.

[0100] The term "combination" refers to the use of two or more components in combination. For example, the combination of an API of the present invention (hereinafter sometimes referred to as the "first component") and a chemotherapeutic agent (hereinafter sometimes referred to as the "second component") includes "administration as a single formulation containing the first component and the second component" (i.e., administration of the first component and the second component as a combined drug) and "administration of the first component and the second component as separate formulations, either simultaneously or separately." In the latter embodiment, the formulation containing the first component may be administered first, or the formulation containing the second component may be administered first. The latter embodiment may be any of "an embodiment in which the first component and the second component are formulated separately and administered simultaneously via the same administration route," "an embodiment in which the first component and the second component are formulated separately and administered separately via the same administration route at different times," "an embodiment in which the first component and the second component are formulated separately and administered simultaneously via different administration routes (administered from different sites in the same patient)," and "an embodiment in which the first component and the second component are formulated separately and administered separately via different administration routes at different times." In the case of "an embodiment in which the first component and the second component are formulated separately and administered simultaneously via the same administration route," the two formulations may be mixed immediately before administration. "Separately" means that one formulation is administered before or after the other formulation.

[0101] In other words, "combined use" can also be said to be a method of use in which one component is present in the patient's body while the other component is present in the patient's body. That is, a preferred embodiment is one in which the first component and the second component are administered so that they are simultaneously present in the patient's body, for example, in the blood, and a preferred embodiment is one in which one formulation is administered to the patient simultaneously, or one formulation is administered to the patient within 48 hours of the other formulation being administered.

[0102] A chemotherapeutic agent is a substance that has anti-cancer activity (also called anti-tumor activity). A chemotherapeutic agent can be, for example, an antimetabolite.

[0103] An antimetabolite is a substance that has a chemical structure similar to that of a metabolic substance (e.g., folic acid) and antagonizes or inhibits the metabolic mechanism of an organism. Examples of an antimetabolite include antifolates.

[0104] Antifolates are substances that inhibit DNA biosynthesis by suppressing or inhibiting the activity of enzymes that reduce folic acid to active folic acid, which is essential for nucleic acid synthesis. Examples of antifolates include methotrexate and pemetrexed. A more preferred antifolate is pemetrexed.

[0105] In this embodiment, the preferred chemotherapeutic agent to combine with the compositions of the present invention is pemetrexed.

[0106] The dosage of the API of the present invention is preferably 0.001 to 50 mg per kg of body weight of the subject per day (0.001 to 50 mg / kg / day), more preferably 0.001 to 20 mg / kg / day, and even more preferably 0.002 to 10 mg / kg / day. When the dosage of the API of the present invention is within these ranges, the cancer treatment or prevention effect is further enhanced. The frequency of administration of the API of the present invention can be, for example, once or more per week, twice per week, once per day, or twice per day.

[0107] The dosage of the chemotherapeutic agent is preferably 0.005 to 300 mg per kg of body weight of the subject per day (0.005 to 300 mg / kg / day), more preferably 0.01 to 250 mg / kg / day, and even more preferably 0.02 to 200 mg / kg / day. When the dosage of the chemotherapeutic agent is within these ranges, the cancer treatment or prevention effect is further enhanced. The frequency of administration of the chemotherapeutic agent can be, for example, once or more per week, twice per week, once per day, or twice per day.

[0108] In the present invention, administration methods include oral, rectal, parenteral (intravenous, intramuscular, subcutaneous, transdermal), intracisternal, intravaginal, intraperitoneal, intravesical, or topical (injection, infusion, powder, ointment, gel, or cream) administration, and inhalation (buccal or nasal spray). Dosage forms include, for example, tablets, capsules, granules, powders, pills, aqueous and non-aqueous oral solutions and suspensions, and parenteral solutions packaged in containers adapted for individual dosages. Dosage forms can also be adapted for various administration methods, including controlled-release formulations such as subcutaneous implants.

[0109] The first component can be, for example, administered by any of the methods described above, and the second component can be, for example, administered by the same or different method as the first component.

[0110] Another aspect of this embodiment is the use of a composition of the invention for the manufacture of a medicament to be administered in combination with a chemotherapeutic agent for the treatment or prevention of cancer in a patient in whom RB1 gene mutation or reduced expression of the RB1 gene or protein has been detected. Another aspect of this embodiment is the use of a composition of the invention for the manufacture of a medicament to be administered in combination with a chemotherapeutic agent for the treatment or prevention of cancer in a patient in whom RB1 gene mutation or reduced expression of the RB1 gene or protein has occurred.

[0111] [Second Embodiment Relating to Cancer Treatment or Prevention] A second embodiment of the present invention relating to cancer treatment or prevention is a method for treating or preventing cancer in a patient in whom a positive RB1 gene mutation or reduced expression of the RB1 gene or protein has been detected, the method comprising administering to the cancer patient a combination of a chemotherapeutic agent and a composition of the present invention. One aspect of this embodiment is a method for treating or preventing cancer, the method comprising detecting, or having a third party detect, a positive RB1 gene mutation or reduced expression of the RB1 gene or protein in a biological sample derived from the cancer patient, and administering to the cancer patient a combination of a chemotherapeutic agent and a composition of the present invention.

[0112] When administering a chemotherapeutic agent in combination with the composition of the present invention, both the composition of the present invention and the chemotherapeutic agent may be administered simultaneously, or may be administered separately at a fixed interval. The administration routes of the composition of the present invention and the chemotherapeutic agent may be the same or different. The composition of the present invention may also be administered in the form of a combination drug containing the chemotherapeutic agent. That is, the pharmaceutical composition may contain both the composition of the present invention and the chemotherapeutic agent.

[0113] The present invention provides a pharmaceutical composition comprising a crystalline compound of the present invention, or a salt thereof, or a solvate thereof, mixed with a pharmaceutically acceptable carrier or vehicle. The present invention also provides a method for producing a pharmaceutical composition comprising a crystalline compound of the present invention, or a salt thereof, or a solvate thereof, as an active ingredient, the method comprising mixing the crystalline compound with a pharmaceutically acceptable carrier or vehicle.

[0114] The present invention provides a method for producing a pharmaceutical composition containing the compound of the present invention, or a salt thereof, or a solvate of the compound or the salt as an active ingredient, the method comprising the step of mixing the compound, the salt thereof, or the solvate of the compound or the salt with a pharmaceutically acceptable carrier or vehicle.

[0115] As used herein, "mixing a crystal of a compound or a salt thereof, or a solvate thereof with a pharmaceutically acceptable carrier or vehicle" includes both of the following: (a) adding another component sequentially to either (1) a crystal of a compound or a salt thereof, or a solvate thereof, and (2) a pharmaceutically acceptable carrier or vehicle; and (b) adding (1) and (2) simultaneously. Furthermore, as used herein, "mixing a compound or a salt thereof, or a solvate thereof with a pharmaceutically acceptable carrier or vehicle" includes both of the following: (a') adding another component sequentially to either (1') a compound or a salt thereof, or a solvate thereof, and (2') a pharmaceutically acceptable carrier or vehicle; and (b') adding (1') and (2') simultaneously. Here, "mixing" does not necessarily require that a homogeneous mixture be obtained when mixing (1) and (2), or (1') and (2'). For example, "mixing" includes "dissolving," "suspending," and "emulsifying."

[0116] The present disclosure will be described in more detail below based on examples, but the present disclosure is not limited to the following examples.

[0117] Examples of abbreviations used herein are listed below along with their meanings: DCM: dichloromethane DIPEA: N,N-diisopropyl-N-ethylamine DMA: N,N-dimethylacetamide DMSO: dimethyl sulfoxide TFA: trifluoroacetic acid THF: tetrahydrofuran T3P: 2,4,6-tripropyl-1,3,5,2,4,6-trioxatriphosphorinane-2,4,6-trioxide TBME: tert-butyl methyl ether TFE: 2,2,2-trifluoroethanol FA: formic acid CH ３ CN: acetonitrile

[0118] Mass spectral data were obtained using a Shimadzu Corporation ultra-high performance liquid chromatograph (product name: Nexera UC) equipped with a single quadrupole mass spectrometer (product name: LCMS-2020) or a Waters Corporation Acquity ultra-high performance liquid chromatograph (UPLC or UPLC I-Class) equipped with a single quadrupole mass spectrometer (SQD or SQD2).

[0119] The high performance liquid chromatography conditions were either those shown in Table 1 or Table 2 below.

[0120]

[0121]

[0122] Commercially available reagents were used without further purification. All non-aqueous reactions were carried out using commercially available anhydrous solvents. Concentration under reduced pressure or solvent evaporation was carried out using a rotary evaporator.

[0123] As used herein, "room temperature" means a temperature of about 20°C to about 25°C.

[0124] Example 1: Preparation of compound 1 3-amino-4-[6,7-difluoro-1-(oxan-2-yl)indazol-4-yl]-6-methyl-1H-1,7-phenanthrolin-2-one (compound 1) and the monohydrate of the hydrochloride of compound 1 were prepared according to the following method.

[0125] 6,7-difluoro-1H-indazole-4-carboxylic acid

[0126] 4-Bromo-6,7-difluoro-1H-indazole (10 g, 42.92 mmol), oxalic acid dihydrate (8.12 g, 64.37 mmol), palladium acetate (0.48 g, 2.15 mmol), and 4,5-bis(diphenylphosphino)-9,9-dimethylxanthene (1.24 g, 2.15 mmol) were added to a reaction vessel, and the reaction vessel was degassed under reduced pressure and purged with nitrogen. To this mixture, acetic anhydride (6.09 mL, 64.37 mmol) and DMF (90 mL) were added, and the reaction vessel was purged with nitrogen. The reaction mixture was heated to 80°C, and DIPEA (11.99 mL, 68.66 mmol) was added dropwise over 1 hour. The reaction mixture was stirred at 80°C for 1 hour, after which oxalic acid dihydrate (5.41 g, 42.92 mmol) and acetic anhydride (4.06 mL, 42.92 mmol) were added, and DIPEA (7.5 mL, 42.9 mmol) was added dropwise over 1 hour. The reaction mixture was stirred at 80°C for 1 hour. After cooling the reaction mixture to 0°C, 5 M aqueous potassium hydroxide solution (103 mL, 0.51 mol) was added dropwise. The reaction mixture was stirred at 0°C for 30 minutes and then at room temperature for 1 hour. Isopropyl acetate (100 mL) was added to the reaction mixture, and the insoluble matter was filtered off. The organic layer was discarded, and the aqueous layer was washed with isopropyl acetate (100 mL). Concentrated hydrochloric acid was added to the aqueous layer to adjust the pH to 3-4. The resulting solid was collected by filtration and washed twice with water (20 mL). The resulting solid was dried under reduced pressure at 50° C. to give the title compound (8.47 g, yield 96%) as a pale brown solid. LCMS: m / z 199 [M+H] ＋ HPLC retention time: 2.03 minutes (Analysis conditions LC-A)

[0127] 6,7-difluoro-1-(oxan-2-yl)-1H-indazole-4-carboxylic acid

[0128] 6,7-Difluoro-1H-indazole-4-carboxylic acid (3.00 g, 15 mmol) was added to acetonitrile (60 mL) in a reaction vessel and stirred at 40°C. Methanesulfonic acid (72.8 mg, 0.76 mmol) and 3,4-dihydro-2H-pyran (1.91 g, 22.7 mmol) were added sequentially, and the mixture was stirred at 40°C for 2 hours. The reaction mixture was cooled to 25°C and stirred for 1 hour. The precipitate was collected by filtration and washed twice with acetonitrile (15 mL). The solid obtained was dried under reduced pressure to give the title compound (3.81 g, 89% yield) as a white solid. LCMS: m / z 283.08 [M+H] ＋ HPLC retention time: 3.47 minutes (measurement conditions LC-A)

[0129] 6,7-difluoro-N-methoxy-N-methyl-1-(oxan-2-yl)-1H-indazole-4-carboxamide

[0130] DIPEA (15.22 g, 117.78 mmol) and N,O-dimethylhydroxylamine hydrochloride (3.73 g, 38.28 mmol) were added to a suspension of 6,7-difluoro-1-(oxan-2-yl)-1H-indazole-4-carboxylic acid (8.31 g, 29.45 mmol) and acetonitrile (58 mL) in a reaction vessel cooled to 10°C, and the mixture was stirred at 0°C. A 50% T3P ethyl acetate solution (37.48 g, 58.89 mmol) was added over 60 minutes, and after the addition, the mixture was heated to 25°C and stirred for 1 hour. The mixture was concentrated under reduced pressure to 42 mL. Toluene (42 mL) was added, and the mixture was concentrated under reduced pressure to 42 mL. After cooling to 10°C, the mixture was washed twice with 10% aqueous potassium carbonate and then with 10% brine. The obtained organic layer was concentrated under reduced pressure to 21 mL. Toluene (42 ml) was added to the concentrated solution, and the mixture was concentrated under reduced pressure to 21 ml. Toluene (25 ml) was added to the concentrated solution, and the mixture was concentrated under reduced pressure to 21 ml to obtain the title compound (21.22 g, content 31.7%, yield 70%) as a toluene solution. LCMS: m / z 326.12 [M+H] ＋ HPLC retention time: 3.61 minutes (measurement conditions LC-A)

[0131] 6-iodo-8-methylquinolin-5-amine

[0132] A reaction vessel containing 8-methylquinolin-5-amine (15.0 g, 95 mmol), acetonitrile (75.0 mL), and water (75.0 mL) was cooled on ice. TFA (21.62 g, 190 mmol) and N-iodosuccinimide (20.48 g, 91 mmol) were added, and the mixture was stirred under ice-cooling for 3 hours. A 5% aqueous solution of L-ascorbic acid (150 g) was added to the reaction mixture, and the mixture was stirred at room temperature. The resulting aqueous solution was divided equally into three containers, and the following procedure was carried out on one of the containers: 2 mol / L aqueous sodium hydroxide solution (45.0 mL) was added and stirred. The solid was collected by filtration and washed with a mixture of acetonitrile (2.5 mL), water (22.5 mL), and a mixture of ethanol (2.5 mL), and heptane (22.5 mL), to give the title compound (7.56 g, 84% yield) as a brown solid. LCMS: m / z 285 [M+H] ＋ HPLC retention time: 2.45 minutes (Analysis conditions LC-A)

[0133] Seed crystals of (E)-N'-(6-iodo-8-methylquinolin-5-yl)-N,N-dimethylmethanimidamide N,N-Dimethylformamide dimethyl acetal (3.77 mL, 28.2 mmol) was added to a suspension of 6-iodo-8-methylquinolin-5-amine (4.00 g, 14.08 mmol) in ethanol (20.0 mL) in a reaction vessel, and the mixture was stirred at 90°C for 2 hours. The reaction mixture was concentrated under reduced pressure, and 10% aqueous sodium chloride solution (30 mL) was added, followed by extraction with isopropyl acetate (30 mL). The resulting organic layer was concentrated under reduced pressure, and TBME and n-heptane were added to the concentrated residue, followed by stirring. The solid was collected by filtration and washed with n-heptane to obtain the title compound (3.8 g, yield 80%) as a pale brown solid. LCMS: m / z 340 [M+H] ＋ HPLC retention time: 0.77 minutes (Analysis conditions Y)

[0134] (E)-N'-(6-iodo-8-methylquinolin-5-yl)-N,N-dimethylmethanimidamide

[0135] N,N-Dimethylformamide dimethyl acetal (4.68 mL, 35.2 mmol) was added to a suspension of 6-iodo-8-methylquinolin-5-amine (compound c-270, 5.00 g, 17.6 mmol) in ethanol (20.0 mL) in a reaction vessel, and the mixture was stirred at 80°C for 2 hours. The reaction mixture was cooled to 25°C, and water (10 mL) was added. Seed crystals of (E)-N'-(6-iodo-8-methylquinolin-5-yl)-N,N-dimethylmethanimidamide (5.0 mg) were added, and the mixture was stirred for 1 hour. Water (36.5 mL) was added over 1 hour, and the mixture was stirred. The solid was collected by filtration and washed twice with water (20.0 mL) to give the title compound (5.3 g, 88% yield) as a pale brown solid. LCMS: m / z 340 [M+H] ＋ HPLC retention time: 2.20 minutes (Analysis conditions LC-A)

[0136] Seed crystals of (E)-N'-[6-[6,7-difluoro-1-(oxan-2-yl)-1H-indazole-4-carbonyl]-8-methylquinolin-5-yl]-N,N-dimethylmethanimidamide A toluene solution (1.5 mL) of (E)-N'-(6-iodo-8-methylquinolin-5-yl)-N,N-dimethylmethanimidamide (146 mg, 0.430 mmol) in a reaction vessel was cooled to -10°C, and a 1.3 M solution of isopropylmagnesium chloride lithium chloride complex in THF (0.355 mL, 0.461 mmol) was added, followed by stirring at -10°C for 1 hour. To the reaction mixture, a toluene solution (0.25 mL) of 6,7-difluoro-N-methoxy-N-methyl-1-(oxan-2-yl)indazole-4-carboxamide (100 mg, 0.307 mmol) was added dropwise. The reaction mixture was stirred at -10°C for 10 minutes and then at 0°C for 30 minutes. A 17% aqueous ammonium chloride solution was added to the reaction mixture, which was extracted with ethyl acetate and concentrated under reduced pressure. Acetonitrile (1.0 mL) was added to the residue, and the mixture was dissolved at 70° C. and then cooled to 35° C. The solid was collected by filtration and washed with acetonitrile (2 mL) to give the title compound (36 mg, yield 24%) as a yellow solid. LCMS: m / z 478 [M+H] ＋ HPLC retention time: 1.84 minutes (Analysis conditions Y)

[0137] (E)-N'-[6-[6,7-difluoro-1-(oxan-2-yl)-1H-indazole-4-carbonyl]-8-methylquinolin-5-yl]-N,N-dimethylmethaneimidamide

[0138] Toluene (14 mL) was added to (E)-N'-(6-iodo-8-methylquinolin-5-yl)-N,N-dimethylmethanimidamide (2.92 g, 8.61 mmol) in a reaction vessel and cooled to -15°C. A THF solution of isopropyl magnesium chloride-lithium chloride complex (6.6 mL, 8.61 mmol) was added, and the mixture was stirred at the same temperature for 1 hour. A toluene solution of 6,7-difluoro-N-methoxy-N-methyl-1-(oxan-2-yl)-1H-indazole-4-carboxamide (9.35 g, 6.15 mmol) in a separate reaction vessel was transferred to the previously prepared reaction solution. After the transfer was complete, the reaction solution was warmed to 0°C and stirred for 4 hours. 20% aqueous ammonium chloride solution (22 mL) and THF (14 mL) were added, and the mixture was separated. 15% brine (20 mL) was added to the resulting organic layer, and the mixture was separated. The obtained organic layer was concentrated under reduced pressure to 6 ml. Toluene (2 ml) and isopropanol (2 ml) were added to the concentrated solution, and the mixture was heated to 70°C to form a homogeneous solution. The temperature was lowered to 40°C, isopropanol (32 ml) was added, and the mixture was cooled to 0°C. The precipitate was collected by filtration and washed with a mixture of toluene (1.1 ml) and isopropanol (4.9 ml) cooled to 5°C, and methanol (6 ml). The obtained solid was dried under reduced pressure to give the title compound (2.28 g, yield 78%) as a yellow solid. LCMS: m / z 478.20 [M+H] ＋ HPLC retention time: 3.20 minutes (Analysis conditions LC-A)

[0139] (5-amino-8-methylquinolin-6-yl)[6,7-difluoro-1-(oxan-2-yl)-1H-indazol-4-yl]methanone DMSO solvate

[0140] DMSO (60 mL), water (9.0 mL), and 50% aqueous potassium hydroxide solution (1.1 g, 9.4 mmol) were added to a reaction vessel and cooled to 7° C. THF (15 mL) was added to N'-{6-[6,7-difluoro-1-(oxan-2-yl)-1H-indazole-4-carbonyl]-8-methylquinolin-5-yl}-N,N-dimethylmethanimidamide (1.5 g, 3.1 mmol) in a separately prepared reaction vessel, and the resulting solution was made homogeneous. This solution was then transferred to the reaction solution prepared above and stirred at 7° C. for 4 hours. Acetic acid (1.5 g, 9.4 mmol) and seed crystals of the DMSO solvate of (5-amino-8-methylquinolin-6-yl)[6,7-difluoro-1-(oxan-2-yl)-1H-indazol-4-yl]methanone were added, and the mixture was heated to 40°C. After concentration under reduced pressure, the THF was removed by distillation. After cooling to 20°C, water (6.0 mL) was added and the mixture was stirred for 2 hours. The precipitate was collected by filtration and washed twice with water (7.5 mL). The solid obtained was dried under reduced pressure to give the DMSO solvate of the title compound (0.92 g, yield 70.8%) as a yellow solid. LCMS: m / z 423.16 [M+H] ＋ HPLC retention time: 3.48 minutes (Analysis conditions LC-A)

[0141] (5-amino-8-methylquinolin-6-yl)[6,7-difluoro-1-(oxan-2-yl)-1H-indazol-4-yl]methanone

[0142] THF (22.0 mL) was added to a DMSO solvate of (5-amino-8-methylquinolin-6-yl)[6,7-difluoro-1-(oxan-2-yl)-1H-indazol-4-yl]methanone (4.82 g, containing 4.00 g of (5-amino-8-methylquinolin-6-yl)[6,7-difluoro-1-(oxan-2-yl)-1H-indazol-4-yl]methanone, 9.47 mmol) in a reaction vessel, and the mixture was stirred at 60°C to form a homogeneous solution. A mixture of ethanol (2.40 mL) and water (7.20 mL) was added to the solution over 5 minutes, and the temperature was then lowered to 40°C over 30 minutes. To the solution was added (5-amino-8-methylquinolin-6-yl)[6,7-difluoro-1-(oxan-2-yl)-1H-indazol-4-yl]methanone (4.3 mg, 10.18 μmol), and the mixture was stirred for 30 minutes. A mixture of ethanol (8.00 mL) and water (24.0 mL) was added over 1 hour, and the mixture was stirred for another 1 hour. The temperature was lowered to 5°C over 1 hour, and the mixture was stirred for 3 hours. The precipitate was filtered and washed once with ethanol (16.0 mL) and twice with n-heptane (16.0 mL). The residue was dried under reduced pressure to give the title compound (3.82 g, 95% yield) as a yellow solid. LCMS: m / z = 423.21 [M+H] + HPLC retention time: 3.55 minutes (Analysis conditions LC-A)

[0143] (5-amino-8-methylquinolin-6-yl)[6,7-difluoro-1-(oxan-2-yl)-1H-indazol-4-yl]methanone

[0144] The title compound can also be produced by the following method. N'-{6-[6,7-difluoro-1-(oxan-2-yl)-1H-indazole-4-carbonyl]-8-methylquinolin-5-yl}-N,N-dimethylmethanimidamide (1.0 g, 2.1 mmol) and THF (15 mL) were added to a reaction vessel, and the mixture was cooled to -10°C to form a homogeneous solution. 40% tetrabutylammonium hydroxide aqueous solution (1.6 g, 2.5 mmol) was added, and the mixture was stirred for 8 hours. Acetic acid (0.28 g, 4.6 mmol) was added, and the mixture was heated to 0°C and stirred for 10 minutes. 15% brine (10 mL) was added, and the mixture was separated. The obtained organic layer was concentrated under reduced pressure to 10 mL. Ethanol (0.6 mL) / water (1.8 mL) was added, and the mixture was heated to 60°C to form a homogeneous solution, and the temperature was then lowered to 40°C over 30 minutes. To the solution was added (5-amino-8-methylquinolin-6-yl)[6,7-difluoro-1-(oxan-2-yl)-1H-indazol-4-yl]methanone (1.0 mg) and stirred for 30 minutes. A mixture of ethanol (4.0 mL) and water (12.0 mL) was added over 2 hours, and the mixture was stirred for an additional 1 hour. The temperature was lowered to 5°C over 3.5 hours, and the mixture was stirred for 14 hours. The precipitate was filtered and washed once with ethanol (4.0 mL) and twice with n-heptane (4.0 mL). The residue was dried under reduced pressure to give the title compound (0.66 g, 75.2% yield) as a yellow solid. LCMS: m / z 423.16 [M+H] ＋ HPLC retention time: 3.48 minutes (Analysis conditions LC-A)

[0145] 3-amino-4-[6,7-difluoro-1-(oxan-2-yl)indazol-4-yl]-6-methyl-1H-1,7-phenanthrolin-2-one

[0146] To a solution of (5-amino-8-methylquinolin-6-yl)[6,7-difluoro-1-(oxan-2-yl)-1H-indazol-4-yl]methanone (2.5 g, 5.92 mmol) in DMA (32.5 mL) in a reaction vessel, chloroacetyl chloride (706 μL, 8.88 mmol) was added and stirred at 7°C for 30 minutes. Pyridine (10.0 mL, 0.12 mmol) was added to the reaction mixture, and the mixture was stirred at 70°C for 4 hours. 80% aqueous hydrazine solution (1.80 mL, 29.6 mmol) was added to the reaction mixture over 1 hour, and the mixture was stirred at 63°C for 1 hour. A suspension of 3-amino-4-[6,7-difluoro-1-(oxan-2-yl)-1H-indazol-4-yl]-6-methyl-1,7-phenanthrolin-2(1H)-one (50.0 mg, 108 μmol) in DMA (250 μL, 2.7 mmol) / water (250 μL, 14 mmol) was added to the reaction mixture and stirred for 3 hours. Water (15.0 mL, 0.83 mmol) was then added over 1 hour. After stirring for 30 minutes, water (15.0 mL, 0.83 mmol) was added over 1 hour and the mixture was cooled to 25°C. The precipitate was filtered and washed twice with water (12.5 mL) and once with ethanol (12.5 mL). The residue was dried under reduced pressure to give the title compound (2.67 g, 98% yield) as a tan solid. LCMS: m / z = 462.25 [M+H] + HPLC retention time: 3.23 minutes, 3.29 minutes (Analysis conditions LC-A)

[0147] 3-amino-4-(6,7-difluoro-1H-indazol-4-yl)-6-methyl-1H-1,7-phenanthrolin-2-one (Compound 1)

[0148] A suspension of 3-amino-4-[6,7-difluoro-1-(oxan-2-yl)indazol-4-yl]-6-methyl-1H-1,7-phenanthrolin-2-one (2.0 g, 4.33 mmol) in 2,2,2-trifluoroethanol (20.0 mL) was placed in a reaction vessel and stirred at 30°C. 6M aqueous hydrochloric acid solution (1.59 mL, 9.53 mmol) was added and stirred at 30°C for 14 hours. Ethanol (20.0 mL) was added to the reaction mixture and stirred at 30°C. Triethylamine (1.63 mL, 11.7 mmol) was added over 1 hour and stirred. The solid was collected by filtration and washed three times with ethanol (10 mL). The solid was dried at 40°C to obtain the title compound (1.5 g, 92.9%) as a brown solid. LCMS: m / z 378.06 [M+H] ＋ HPLC retention time: 4.96 minutes (Analysis conditions: LC-C)

[0149] Seed crystals of the monohydrate of the hydrochloride salt of 3-amino-4-(6,7-difluoro-1H-indazol-4-yl)-6-methyl-1H-1,7-phenanthrolin-2-one

[0150] A 1M aqueous solution of hydrochloric acid (25.0 mL) was added to a reaction vessel and the temperature was raised to 60°C. A solution of 3-amino-4-(6,7-difluoro-2H-indazol-4-yl)-6-methyl-1,7-phenanthrolin-2(1H)-one (2.50 g, 6.63 mmol) in THF / water (9:1, 62.5 mL) was added dropwise over 1 hour and stirred for 1 hour. The temperature was lowered to 30°C and stirred for 1 hour. The temperature was lowered to 10°C and stirred for 1 hour. The solid was collected by filtration and washed twice with ethanol (12.5 mL). The solid was dried at 40°C to obtain the title compound (2.67 g, 92.4%) as a yellow solid. LCMS: m / z 378.15 [M+H] ＋ HPLC retention time: 2.47 minutes (Analysis conditions: LC-A)

[0151] 3-amino-4-(6,7-difluoro-1H-indazol-4-yl)-6-methyl-1H-1,7-phenanthrolin-2-one hydrochloride monohydrate

[0152] A 1M aqueous solution of hydrochloric acid (8.0 mL) was added to a reaction vessel, and the temperature was raised to 55°C. Seed crystals of the title compound (1.2 mg) were then added. A solution of 3-amino-4-(6,7-difluoro-2H-indazol-4-yl)-6-methyl-1,7-phenanthrolin-2(1H)-one (800 mg, 2.12 mmol) in THF / water (9:1, 22.4 mL) was added dropwise over 4 hours, followed by stirring for 4 hours. The temperature was lowered to 25°C over 1 hour, and the mixture was stirred for 1 hour. The solid was collected by filtration and washed three times with ethanol (4.0 mL). The mixture was dried at 40°C to obtain the title compound (879.0 mg, 96%) as a yellow solid. LCMS: m / z 378.06 [M+H] ＋ HPLC retention time: 4.96 minutes (Analysis conditions: LC-C)

[0153] Example α1 (1) Preparation of Free Form Crystals of Compound 1 (Form Z) 10.988 mg and 6.859 mg (total 17.8 mg) of free form solvate crystals of Compound 1 (Form B, described below) were weighed into open aluminum pans and heated using a simultaneous thermogravimetry and differential thermal analysis apparatus (Example β11) to obtain a solid of the title compound. Powder X-ray diffraction measurement of the obtained crystals (Example β1) confirmed the following main peaks: 6.82°, 8.98°, 10.35°, 10.89°, 11.88°, 12.09°, 12.36°, 12.72°, 13.46°, and 15.27° (±0.2°). The measurement results are shown in Figure 1. Furthermore, simultaneous thermogravimetry and differential thermal analysis of the obtained crystals (Example β10), and １ H-NMR measurement (Example β18) was carried out. The results of simultaneous thermogravimetry and differential thermal analysis are shown in Figure 2. １ The results of H-NMR measurement are shown in Figure 3. In Figure 2, no weight loss was observed up to the decomposition point around 330°C. １ In the H-NMR measurement, peaks other than those corresponding to the measurement solvent DMSO, water, and Compound 1 were not observed, and it was confirmed that the product was a free-form crystalline form of Compound 1 (Form Z).

[0154] Example α2 (1) Preparation of Free Solvate Crystals of Compound 1 (Form A) DMSO (0.40 mL) was added to the free solvate crystals of Compound 1 (Form B, 100.2 mg) and stirred at 60°C to dissolve. Ethanol (1.00 mL) was added to this solution and stirred at room temperature for 16 hours, after which the solid was collected by filtration and washed with 2-propanol (0.50 mL). The resulting solid was dried in vacuo for 1 day to obtain a solid of the title compound. Powder X-ray diffraction measurement (Example β1) of the resulting crystals confirmed the following major peaks: 5.75°, 6.30°, 7.93°, 10.14°, 10.62°, 11.52°, 12.63°, 15.80°, 18.55°, and 23.72° (±0.2°). The measurement results are shown in FIG. 4. The results of simultaneous thermogravimetry and differential thermal analysis (Example β9) are shown in Figure 5. Since a weight loss due to the solvent was observed, the obtained solid was confirmed to be a free solvate crystal of Compound 1.

[0155] Example α3 (4) Preparation of Compound 1 Free Solvate Crystals (Form A) and Single Crystal Measurement Compound 1 free solvate crystals (Form B, 122.6 mg) were dissolved in DMSO (0.613 mL), and this solution (0.015 mL) was freeze-dried at -20°C for 3 days. Tetrahydrofuran (0.015 mL) was added to the resulting freeze-dried product, and the mixture was shaken and stirred at room temperature for 7 days to obtain solid Compound 1. The results of powder X-ray diffraction measurement of the obtained crystals (Example β3) are shown in Figure 6. The obtained crystals were confirmed to be Compound 1 free hydrate crystals (Form A) by single crystal X-ray structural analysis (Example β14). The structure is shown in Figure 7.

[0156] Example α4 Preparation of a Free Solvate of Compound 1 (Ethanol Solvate Crystal, Form H) A suspension of 3-amino-4-[6,7-difluoro-1-(oxan-2-yl)indazol-4-yl]-6-methyl-1H-1,7-phenanthrolin-2-one (1.0 g, 2.2 mmol) in 2,2,2-trifluoroethanol (10.0 mL) was added to a reaction vessel at 25°C with a stirrer and 6 M aqueous hydrochloric acid (0.79 mL, 4.77 mmol). The mixture was stirred at 25°C. Triethylamine (0.82 mL, 5.85 mmol) was added over 1 hour and the mixture was stirred. The solid was collected by filtration and washed three times with ethanol (10 mL). Powder X-ray diffraction analysis (Example β4) of the obtained crystals was performed. The results are shown in Figure 8. １ The results of H-NMR measurement are shown in Figure 9. It was confirmed that the compound was an ethanol solvate.

[0157] Example α5 Measurement of free solvate (Form A) of Compound 1 Powder X-ray diffraction measurement (Example β4) was carried out on 3-amino-4-(6,7-difluoro-1H-indazol-4-yl)-6-methyl-1H-1,7-phenanthrolin-2-one obtained in Example 1. The results are shown in FIG.

[0158] Example α6 (1) Preparation of Free Solvate Crystals of Compound 1 (Form B) 3-amino-4-[6,7-difluoro-1-(oxan-2-yl)indazol-4-yl]-6-methyl-1H-1,7-phenanthrolin-2-one (9.61 g) obtained in Example 1 was added to a 2,2,2-trifluoroethanol-water mixture (10:1 (v / v), 210 mL) and stirred at room temperature to dissolve. Triisopropylsilane (34.2 mL) was added to this solution, followed by trimethylsilyl chloride (13.1 mL) and stirring at room temperature for 3 hours. 600 mL of sodium bicarbonate water was added to the reaction solution, and the solid was collected by filtration. It was then washed with water (700 mL) and an acetonitrile-water mixture (600 mL), and air-dried. 80 mL of acetonitrile was added to this solid, and the mixture was stirred at room temperature, followed by filtration. The solid was washed with acetonitrile (40 mL × 2). DMSO (50 mL) was added to this solid, and the temperature was raised to 40°C, followed by filtration of the solution. Water (300 mL) was added to this solution, and the resulting precipitate was collected by filtration and washed with water. This solid was air-dried overnight and then vacuum-dried overnight to obtain a solid of the title compound (7.07 g). Powder X-ray diffraction measurement (Example β1) of the obtained crystals confirmed the following main peaks: 7.01°, 8.74°, 9.14°, 10.17°, 10.82°, 11.78°, 13.31°, 13.93°, 17.00°, and 22.77° (±0.2°). The measurement results are shown in Figure 11. The results of simultaneous thermogravimetry and differential thermal analysis (Example β9) are shown in Figure 12. １ The results of H-NMR measurement (Example β19) are shown in Figure 13. While weight loss due to the solvent was observed in Figure 12, peaks other than those due to the measurement solvent DMSO, water, and the internal standard substances tetramethylsilane (TMS) and 1,3,5-trimethoxybenzene were not observed in Figure 13, confirming that the obtained solid was a free-form hydrate crystal of Compound 1 (Form B).

[0159] Example α7 (1) Preparation of Compound 1 Free Solvate Crystals (Form C) Compound 1 free solvate crystals (Form B, 122.6 mg) were dissolved in DMSO (0.613 mL), and this solution (0.015 mL) was freeze-dried at −20° C. for 3 days. 1-Propanol (0.015 mL) was added to the resulting freeze-dried product, and the mixture was shaken and stirred at room temperature for 7 days to obtain a solid of Compound 1. The results of powder X-ray diffraction measurement of the resulting solid (Example β3) are shown in FIG. 14. The solid was confirmed to be crystalline. The results of simultaneous thermogravimetry, differential thermal analysis, and mass spectrometry (Example β12) are shown in FIG. 15. Ion peaks corresponding to the m / z values ​​of 1-propanol and DMSO were detected near 195° C. and 212° C., respectively, confirming that the solid was a 1-propanol-DMSO solvate crystal of Compound 1 free form (Form C).

[0160] Example α8 (1) Preparation of Compound 1 Free Solvate Crystals (Form D) Compound 1 free solvate crystals (Form B, 122.6 mg) were dissolved in DMSO (0.613 mL), and the resulting solution (0.015 mL) was freeze-dried at −20° C. for 3 days. 2-Butanol (0.015 mL) was added to the resulting freeze-dried product, and the mixture was shaken and stirred at room temperature for 7 days to obtain a solid of Compound 1. The results of powder X-ray diffraction measurement of the resulting solid (Example β3) are shown in FIG. 16 . This confirmed that the solid was a crystalline solid. The results of simultaneous thermogravimetry, differential thermal analysis, and mass spectrometry (Example β12) are shown in FIG. 17 . Ion peaks corresponding to the m / z values ​​of 2-butanol and DMSO were detected near 178° C. and 184° C., respectively, confirming that the solid was a 2-butanol-DMSO solvate crystal of Compound 1 free form (Form D).

[0161] Example α9 (1) Preparation of Compound 1 Free Solvate Crystals (Form E) Compound 1 free solvate crystals (Form B, 122.6 mg) were dissolved in DMSO (0.613 mL), and the resulting solution (0.015 mL) was lyophilized at −20° C. for 3 days. Formamide (0.015 mL) was added to the resulting lyophilized product, and the mixture was shaken and stirred at room temperature for 7 days to obtain a solid of Compound 1. The results of powder X-ray diffraction measurement of the resulting solid (Example β3) are shown in FIG. 18. The solid was confirmed to be crystalline. The results of simultaneous thermogravimetry, differential thermal analysis, and mass spectrometry (Example β12) are shown in FIG. 19. An ion peak corresponding to the m / z of formamide was detected around 233° C., confirming that the solid was a free formamide solvate crystal of Compound 1 (Form E).

[0162] Example α10 (1) Preparation of Free Solvate Crystals of Compound 1 (Form F) The free solvate crystals of Compound 1 (Form B, 3.4 mg), the free solvate crystals of Compound 1 (Form A, 3.4 mg), and the free solvate crystals of Compound 1 (Form Z, 4.4 mg) were weighed into a reaction vessel, and one glass bead was added. Formic acid (0.015 mL) was added, and the mixture was stirred with shaking at 37°C for 1 week to obtain a solid of Compound 1 (Sample A). Formic acid (0.142 mL) was added to the free solvate crystals of Compound 1 (Form B, 56.6 mg) in the reaction vessel, and the mixture was stirred at 60°C for 10 minutes. After that, the aforementioned Sample A (approximately 0.1 mg) was added, and the mixture was stirred at room temperature for an additional 16 hours. The resulting solid was collected by filtration, washed with 2-propanol (0.57 mL), and then vacuum-dried overnight. The results of powder X-ray diffraction measurement of the obtained solid (Example β3) are shown in Figure 20. It was confirmed to be a crystal. The results of simultaneous thermogravimetry, differential thermal analysis, and mass spectrometry (Example β13) are shown in Figure 21. An ion peak corresponding to the m / z of the solvent formic acid was detected around 120°C, and the weight loss value corresponds to the amount of 0.5 molecules of formic acid per molecule of Compound 1. From the above, it was confirmed that it was a free form formic acid solvate crystal of Compound 1 (Form F).

[0163] Example α11 (1) Preparation of Compound 1 Free Solvate Crystals (Form G) Acetic acid (0.265 mL) was added to Compound 1 free solvate crystals (Form B, 53.0 mg) in a reaction vessel, and the mixture was stirred at 60°C for 10 minutes, followed by stirring at room temperature for an additional 16 hours. The resulting solid was collected by filtration, washed with 2-propanol (0.53 mL), and then vacuum-dried overnight. The results of powder X-ray diffraction measurement of the resulting solid (Example β3) are shown in Figure 22. The solid was confirmed to be crystalline. The results of simultaneous thermogravimetry, differential thermal analysis, and mass spectrometry (Example β13) are shown in Figure 23. An ion peak corresponding to the m / z of the solvent acetic acid was detected around 120°C, and the weight loss value corresponds to the amount of two acetic acid molecules per one molecule of Compound 1. From the above, the solid was confirmed to be Compound 1 free acetic acid solvate crystals (Form G).

[0164] Example α12 (1) Preparation of Compound 1 Hydrochloride Crystal (Form P) To confirm the change in crystal form due to temperature change in the Compound 1 hydrochloride monohydrate crystal (Form A, described below) prepared in Example α13 under a nitrogen atmosphere, temperature-controlled powder X-ray diffraction measurement (Example β5) was performed. The results obtained by heating to 100°C are shown in Figure 24, and the results obtained by cooling to 35°C while maintaining the nitrogen atmosphere are shown in Figure 25. It was confirmed that a new crystal form was obtained. When this crystal was removed from the laboratory, it rapidly transformed into Compound 1 hydrochloride monohydrate crystal (Form A). As shown in Figure 27, simultaneous thermogravimetric, differential thermal, and mass spectrometry measurements of Compound 1 hydrochloride monohydrate crystal (Form A) revealed a dehydration peak near 85°C, and the weight loss value corresponded to the amount of one water molecule in Compound 1 hydrochloride monohydrate crystal (Form A). This confirmed that the product was a hydrochloride crystal (Form P) of Compound 1. The hydrochloride crystal (Form P) of Compound 1 has major peaks at 100°C: 8.94°, 11.49°, 12.65°, 12.94°, 16.03°, 16.34°, 17.61°, 19.11°, 19.68°, and 20.64° (±0.2°). The major peaks at 35°C are 9.02°, 11.47°, 12.71°, 12.98°, 16.50°, 17.83°, 19.24°, 19.68°, 20.80°, and 22.29° (±0.2°).

[0165] Example α13 (1) Preparation of Compound 1 Hydrochloride Monohydrate (Form A) DMSO (1.7 mL) was added to the free solvate crystals of Compound 1 (Form B, 342.5 mg) in a reaction vessel, and the mixture was dissolved at room temperature. 2 M hydrochloric acid (2.57 mL) and water (0.86 mL) were added to this solution, and the mixture was stirred at room temperature for 6 minutes. A wet solid of the hydrochloride salt of Compound 1 (Sample B described below) was added to the reaction vessel, and the mixture was stirred at room temperature for 23 hours. The solid was then filtered and washed with 2-propanol (4.3 mL). The resulting solid was dried in vacuo for 4 days to obtain a solid of the title compound. Powder X-ray diffraction measurement of the obtained crystals (Example β2) confirmed the following major peaks: 6.59°, 9.19°, 9.70°, 12.70°, 13.15°, 13.29°, 13.80°, 14.93°, 15.80°, and 16.45° (±0.2°). The measurement results are shown in Figure 26. The results of simultaneous thermogravimetry, differential thermal analysis, and mass spectrometry measurement of the obtained solid (Example β13) are shown in Figure 27. Ion peaks corresponding to the m / z values ​​of water and HCl were detected around 100°C and 230°C, respectively, and the weight loss value corresponds to the amount of one molecule of water and one molecule of HCl per molecule of Compound 1. In addition, chloride ion content analysis (Example β16) was performed using ultra-high performance liquid chromatography, and the quantitative value was 7.87±0.19%, corresponding to hydrochloride monohydrate. From the above, it was confirmed that the obtained solid was Compound 1 hydrochloride monohydrate crystal (Form A). (2) Preparation of Sample B (Seed Crystal): 2M hydrochloric acid (0.10 mL) and DMSO (0.30 mL) were added to Compound 1 free form solvate crystal (Form A, 71.7 mg) in a reaction vessel, and the mixture was stirred at room temperature for approximately 1 minute. DMSO (0.40 mL) was then added, and the mixture was stirred at room temperature for 1 minute. DMSO (0.40 mL) was then added, and the mixture was stirred at room temperature for 1 minute. The resulting suspension (0.045 mL) was lyophilized at -20°C for 3 days. Water (0.015 mL) was added to the resulting lyophilized product, and the mixture was shaken and stirred at room temperature for 7 days to obtain a solid (Sample B-1). DMSO (0.15 mL) was added to Compound 1 free form solvate crystal (Form B, 30.0 mg) in a reaction vessel, and the mixture was dissolved at room temperature. To this solution, 2M hydrochloric acid (0.23 mL) and water (0.075 mL) were added, and the mixture was stirred at room temperature for 3 minutes.The above-mentioned sample B-1 was added to a reaction vessel, and after stirring at room temperature for 23 hours, the solid was collected by filtration and washed with 2-propanol (0.15 mL). The obtained wet solid was dried in vacuum overnight to obtain a solid. Using the obtained solid, the same operation was repeated twice to obtain sample B.

[0166] Example α14 (1) Preparation of Compound 1 Hydrochloride Monohydrate Crystal (Form A) and Single Crystal Measurement DMSO (0.60 mL) was added to Compound 1 free solvate crystal (Form A, approximately 10 mg) in a reaction vessel, and the mixture was stirred at 50°C to dissolve. 2 M hydrochloric acid (0.10 mL) was added to this solution. Dichloromethane (approximately 0.2 mL) was slowly dispersed in 0.10 mL of this solution over two days to obtain a single crystal. The obtained crystal was confirmed to be Compound 1 hydrochloride monohydrate crystal (Form A) by single crystal X-ray structural analysis (Example β15). The crystal structure is shown in Figure 28. The powder X-ray diffraction pattern calculated from the single crystal X-ray structure matched the measured pattern (Figure 29), confirming that the crystal was Compound 1 hydrochloride monohydrate crystal (Form A).

[0167] Example α15 Preparation and Measurement of Single Crystal of Compound 1 Hydrochloride Monohydrate (Form A) Powder X-ray diffraction measurement (Example β4) was carried out on the monohydrate of the monohydrochloride of 3-amino-4-(6,7-difluoro-2H-indazol-4-yl)-6-methyl-1,7-phenanthrolin-2(1H)-one obtained in Example 1. The results are shown in Figure 30.

[0168] Example α16 (1) Preparation of Compound 1 0.5 Hydrochloride Dihydrate Crystals (Form C) Water (0.066 mL) was added to Compound 1 free solvate crystals (Form B, 10.3 mg) in a reaction vessel. A 2M hydrochloric acid-water mixture (1.4:8.6 (v / v), 0.103 mL) was then added, and the mixture was stirred at room temperature for 17 hours to obtain Compound 1 wet solid (Sample B2). Water (0.35 mL) was added to Compound 1 free solvate crystals (Form B, 69.0 mg) in a reaction vessel, and the mixture was suspended at room temperature. A 2M hydrochloric acid-water mixture (1.4:8.6 (v / v), 0.69 mL) was then added to the suspension. Approximately 0.5 mg of Compound 1 wet solid (Sample B2) was then added, and the mixture was stirred at room temperature for 3 hours. Further water (0.35 mL) was added, and the mixture was stirred at room temperature for 22 hours. The solid was then filtered and washed with 2-propanol (0.35 mL). The resulting solid was dried in vacuo for 5 days to give a solid of the title compound. Powder X-ray diffraction measurement (Example β1) of the resulting crystals confirmed the following major peaks: 7.07°, 9.24°, 12.06°, 14.37°, 15.24°, 15.84°, 17.86°, 18.30°, 18.52°, and 19.31° (±0.2°). The measurement results are shown in Figure 31. The results of simultaneous thermogravimetry, differential thermal analysis, and mass spectrometry (Example β13) of the resulting solid are shown in Figure 32. Ion peaks corresponding to the m / z values ​​of water and HCl were detected around 100-120°C and around 210°C, respectively, and the weight loss value corresponded to two molecules of water and 0.5 molecules of HCl per molecule of Compound 1. When the water content was analyzed by Karl Fischer water titration (Example β22), the quantitative value was 7.86%, which coincides with the content of Compound 1 0.5 hydrochloride dihydrate. From the above, it was confirmed that the obtained solid was Compound 1 0.5 hydrochloride dihydrate crystals (Form C).

[0169] Example α17 (1) Preparation of Compound 1 1.5 Hydrochloride Hydrate (Form D) 2,2,2-Trifluoroethanol (30 mL) was added to 3-amino4-[6,7-difluoro-1(oxan-2-yl)-indazol-4-yl]-6-methyl-1H-1,7-phenanthrolin-2-one (3.00 g) and the mixture was suspended at 25°C. 6 M hydrochloric acid (2.4 mL) was added and the mixture was stirred at 25°C for 17 hours. The solid was collected by filtration and washed twice with ethanol (30 mL). The obtained solid was dried under reduced pressure at 40°C for 4 hours to obtain the title compound as a solid. Powder X-ray diffraction measurement of the obtained crystals (Example β1) confirmed the following major peaks: 10.24°, 10.60°, 10.85°, 12.75°, 12.91°, 13.12°, 13.96°, 14.76°, 15.93°, and 16.84° (±0.2°). The measurement results are shown in Figure 33. The results of simultaneous thermogravimetry and differential thermal analysis of the obtained solid (Example β9) are shown in Figure 34. Weight loss was observed around 80°C and 230°C, respectively. In addition, chloride ion content analysis by ultra-high performance liquid chromatography revealed a quantitative value of 13.2±1.1%, corresponding to the 1.5 hydrochloride salt. Water content analysis by Karl Fischer moisture meter measurement (Example β22) revealed a quantitative value of 6.1%, corresponding to the 1.5 hydrate. From the above, it was confirmed that the obtained solid was Compound 1 1.5 hydrochloride hydrate (Form D).

[0170] Example α19 (1) Preparation of Sulfate Hydrate Crystals of Compound 1 (Form A and Form G) 2M aqueous sulfuric acid solution (0.22 mL) and water (2.0 mL) were added to the free solvate crystals of Compound 1 (Form A, 198.9 mg) in a reaction vessel, and the mixture was suspended at room temperature for 6 days. The resulting solid was collected by filtration and air-dried for 5 days to obtain a solid of the title compound. Powder X-ray diffraction measurement (Example β1) of the resulting crystals confirmed the following major peaks: 6.95°, 7.50°, 9.05°, 11.28°, 14.32°, 15.13°, 15.72°, 17.89°, 19.86°, and 20.69° (±0.2°). The measurement results are shown in Figure 35. The results of simultaneous thermogravimetry and differential thermal analysis (Example β9) of the resulting solid are shown in Figure 36. Weight loss was observed around 80°C, and the weight loss value corresponds to the amount of water molecules relative to the 0.67% sulfate monohydrate of Compound 1. Furthermore, when sulfate ion content analysis (Example β17) was performed using ultra-high performance liquid chromatography, the quantitative value was 14.8%, which corresponds to the 0.67% sulfate monohydrate of Compound 1. From the above, it was confirmed that the obtained solid was a sulfate hydrate crystal of Compound 1 (Form A). Furthermore, a comparison of the wet powder (Form G) before filtration and the solid (Form A) after air-drying in the above operation is shown in Figure 37. A transition from Form G to Form A was confirmed.

[0171] Example α20 (1) Preparation of Sulfate Solvate Crystals of Compound 1 (Form B) Free solvate crystals of Compound 1 (Form B, 66.3 mg) were dissolved in DMSO (0.331 mL), and 2 M aqueous sulfuric acid solution (46.6 μL) was added. This solution (0.015 mL) was freeze-dried at −20° C. for 3 days. To the resulting freeze-dried product, an ethanol-water mixture (1:3 (v / v), 0.015 mL) was added, and the mixture was shaken and stirred at room temperature for 7 days to obtain a solid. The results of powder X-ray diffraction measurement of the obtained solid (Example β3) are shown in FIG. 38. It was confirmed to be a crystalline solid.

[0172] Example α22 (1) Preparation of Sulfate Solvate Crystals of Compound 1 (Form D) DMSO (0.15 mL) and 2 M aqueous sulfuric acid solution (0.045 mL) were added to the free solvate crystals of Compound 1 (Form B, 29.1 mg) in a reaction vessel, and the mixture was suspended at room temperature for 16 hours. The resulting solid was collected by filtration, washed with 2-propanol, and air-dried for 3 days to obtain a solid of the title compound. The results of powder X-ray diffraction measurement of the resulting solid (Example β3) are shown in Figure 39. The results of simultaneous thermogravimetry and differential thermal analysis of the resulting solid (Example β9) are shown in Figure 40. A weight loss was observed around 100°C, and the weight loss value corresponds to one molecule of DMSO per molecule of Compound 1. In addition, when sulfate ion content was analyzed by ultra-high performance liquid chromatography (Example β17), the quantitative value was 18.5%, which corresponds to the sulfate monohydrate of Compound 1 in DMSO. １ The results of H-NMR (Example β20) are shown in Figure 41. A peak corresponding to 1 equivalent of DMSO relative to Compound 1 was observed. From the above, it was confirmed that the obtained solid was Compound 1 sulfate mono-DMSO solvate crystal (Form D).

[0173] Example α24 (1) Preparation of Sulfate Solvate Crystals of Compound 1 (Form F) Water (0.225 mL) and 2 M aqueous sulfuric acid solution (0.225 mL) were added to the free solvate crystals of Compound 1 (Form B, 32.0 mg) in a reaction vessel, and the mixture was suspended at room temperature for 16 hours. The resulting solid was filtered to obtain a wet solid of the title compound. The results of powder X-ray diffraction analysis (Example β3) of the resulting solid are shown in Figure 42. The solid was confirmed to be crystalline.

[0174] Example α25 (1) Preparation of Compound 1 Mesylate Solvate Crystals (Form A) Compound 1 free solvate crystals (Form B, 60.9 mg) were dissolved in DMSO (0.301 mL), and 2 M mesylic acid aqueous solution (84.7 μL) was added. This solution (0.015 mL) was lyophilized at −20°C for 3 days. 2-Propanol (0.015 mL) was added to the resulting lyophilized product, and the mixture was shaken and stirred at room temperature for 7 days to obtain a wet solid (Sample C). To Compound 1 free solvate crystals (Form B, 15.6 mg) in a reaction vessel, 2 M mesylate 2-propanol solution (0.050 mL) was added. 2-Propanol (0.106 mL) and the aforementioned solid from Sample C (approximately 0.1 mg) were then added, and the mixture was stirred at room temperature for 1 day. The resulting solid was collected by filtration, washed with 2-propanol, and then air-dried for 13 days to obtain a solid of the title compound. The powder X-ray diffraction measurement (Example β3) of the resulting solid is shown in Figure 43. It was confirmed to be a crystalline solid.

[0175] Example α28 (1) Preparation of Compound 1 0.5 Mesylate Hydrate Crystals (Form D) Compound 1 free solvate crystals (Form B, 60.9 mg) were dissolved in DMSO (0.301 mL), and 2 M aqueous mesylic acid solution (84.7 μL) was added. This solution (0.015 mL) was lyophilized at −20° C. for 3 days. To the resulting lyophilized product was added an ethanol-water mixture (1:3 (v / v), 0.015 mL), and the mixture was shaken and stirred at room temperature for 7 days to obtain a wet solid of Compound 1 (Sample C1). DMSO (0.041 mL) and 2 M aqueous mesylic acid solution (0.019 mL) were added to the free solvate crystals of Compound 1 (Form B, 13.6 mg) in a reaction vessel. Further, an ethanol-water mixture (1:3 (v / v), 0.14 mL) and a wet solid of Compound 1 (Sample C1, approximately 0.1 mg) were added, and the mixture was shaken and stirred at room temperature for 1 hour. Further, an ethanol-water mixture (1:3 (v / v), 0.14 mL) was added, and the mixture was shaken and stirred at room temperature for 1 hour. Further, an ethanol-water mixture (1:3 (v / v), 0.14 mL) was added, and the mixture was shaken and stirred at room temperature for 16 hours. The resulting solid was collected by filtration and dried in vacuo for 3 days to obtain a solid of the title compound. When the obtained solid was subjected to powder X-ray diffraction measurement (Example β1), major peaks were confirmed at 5.56°, 7.53°, 10.30°, 10.52°, 11.16°, 11.61°, 12.96°, 15.70°, 16.88°, and 17.58° (±0.2°). The measurement results are shown in FIG. 44. The results of simultaneous thermogravimetric and differential thermal measurements (Example β9) of the obtained solid are shown in FIG. １ The results of H-NMR (Example β18) are shown in Figure 46. In Figure 45, weight loss due to the solvent was observed around 70°C. In Figure 46, a peak corresponding to mesylic acid equivalent to 0.5 equivalents of Compound 1 was observed, while no peaks other than those corresponding to the measurement solvents DMSO, water, and the internal standard TMS were observed. From the above, it was confirmed that the obtained solid was 0.5 mesylate hydrate crystals of Compound 1 (Form D).

[0176] Example α29 (1) Preparation of Compound 1 Mesylate Solvate Crystals (Form E) Compound 1 free solvate crystals (Form B, 60.9 mg) were dissolved in DMSO (0.301 mL), and 2 M aqueous mesylic acid solution (84.7 μL) was added. This solution (0.015 mL) was lyophilized at −20° C. for 3 days. The resulting solid was subjected to powder X-ray diffraction measurement (Example β3). The measurement results are shown in FIG. 47. It was confirmed to be a crystalline solid.

[0177] Example α30 (1) Preparation of Compound 1 Mesylate Hydrate Crystals (Form F and Form H) DMSO (0.071 mL) and 2 M aqueous mesylic acid solution (0.107 mL) were added to Compound 1 free solvate crystals (Form B, 14.2 mg) in a reaction vessel, and approximately 0.1 mg of Sample C2 (described below) was added. Water (0.036 mL) was added, and the mixture was shaken at room temperature for 3 hours. 2-Propanol (0.14 mL) was then added, and the mixture was shaken at room temperature for 2 hours. 2-Propanol (0.42 mL) was then added, and the mixture was shaken at room temperature for 18 hours. The resulting solid was collected by filtration, washed with 2-propanol, and then vacuum-dried for 3 days to obtain the title compound as a solid. Powder X-ray diffraction measurement (Example β1) of the obtained solid confirmed the following major peaks: 5.75°, 8.37°, 9.38°, 10.89°, 13.14°, 16.81°, 17.36°, 17.79°, 18.37°, and 18.78° (±0.2°). The measurement results are shown in Figure 48. The results of simultaneous thermogravimetry and differential thermal analysis (Example β9) of the obtained solid are shown in Figure 49. A weight loss thought to be due to the solvent was observed around 70°C. １ The results of H-NMR (Example β18) are shown in Figure 50. While a peak corresponding to 1 equivalent of mesylate relative to Compound 1 was observed, no peaks derived from the crystallization solvent were observed other than those of the measurement solvent DMSO, water, and the internal standard TMS. From the above, it was confirmed that the obtained solid was a mesylate hydrate crystal of Compound 1 (Form F). Furthermore, powder X-ray diffraction measurement of the wet solid immediately after filtration (Example β3) is shown in Figure 51. It was confirmed that this solid transitioned from a mesylate solvate crystal of Compound 1 (Form H) to a mesylate hydrate crystal of Compound 1 (Form F) upon vacuum drying.

[0178] Example α31 (2) Preparation of Sample C2 (Seed Crystal) DMSO (0.024 mL), 2 M mesylic acid aqueous solution (0.036 mL), and water (0.012 mL) were added to the free solvate crystals of Compound 1 (Form B, 4.8 mg) in a reaction vessel, and the mixture was stirred at room temperature for 30 minutes. Approximately 0.1 mg of Sample C3 (described below) was added, and the mixture was stirred at room temperature for 17 hours. Powder X-ray diffraction measurement (Example β3) of the obtained solid was performed. The measurement results are shown in Figure 52. It was confirmed that the solid was crystalline.

[0179] Example α32 (3) Preparation of Sample C3 (Seed Crystal) The free solvate crystal of Compound 1 (Form B, 60.9 mg) was dissolved in DMSO (0.301 mL), and 2 M aqueous mesylic acid solution (84.7 μL) was added. This solution (0.015 mL) was lyophilized at −20° C. for 3 days. Water (0.015 mL) was added to the resulting lyophilized product, and the mixture was shaken and stirred at room temperature for 7 days to obtain a solid of the title compound. The results of powder X-ray diffraction measurement of the resulting solid (Example β3) are shown in FIG. 53. It was confirmed to be a crystalline solid.

[0180] Example α33 (1) Preparation of Compound 1 Mesylate Crystal (Form G) To confirm the change in crystal form due to temperature change of Compound 1 mesylate hydrate crystal (Form F) under a nitrogen atmosphere, temperature-controlled powder X-ray diffraction measurement (Example β6) was performed. The results obtained by heating to 100°C are shown in Figure 54. It was confirmed that a new crystal form was obtained. When cooled to 30°C, this crystal transitioned to Compound 1 mesylate hydrate crystal (Form F). As shown in Figure 49, simultaneous thermogravimetry and differential thermal analysis of Compound 1 mesylate hydrate crystal (Form F) showed a dehydration peak near 85°C, and the weight loss corresponded to 1-2 water molecules of Compound 1 mesylate hydrochloride hydrate. From this, it was confirmed that the compound was Compound 1 mesylate crystal (Form G).

[0181] Example α35 (1) Preparation of Compound 1 Tosylate Hydrate Crystals (Form A) Compound 1 free solvate crystals (Form B, 38.7 mg) were dissolved in DMSO (0.271 mL), and 2M aqueous p-toluenesulfonic acid solution (0.108 mL) was added, followed by DMSO (0.379 mL). This solution (0.030 mL) was lyophilized at −20° C. for 3 days. A water-ethanol mixture (3:1 (v / v), 0.015 mL) was added to the resulting lyophilized product, and the mixture was shaken and stirred at room temperature for 7 days to obtain Compound 1 solid (Sample D). p-Toluenesulfonic acid monohydrate (70.7 mg) was dissolved in a water-ethanol mixture (3:1 (v / v), 1.0 mL). To the free solvate crystals of Compound 1 (Form A, 64.2 mg) in a reaction vessel was added the aforementioned water-ethanol mixture of p-toluenesulfonic acid (3:1 (v / v), 0.64 mL), followed by the aforementioned Sample D (approximately 0.1 mg), and the mixture was stirred at room temperature for 7 hours. The resulting solid was collected by filtration, washed with acetone, and then vacuum-dried for 3 days to obtain a solid of the title compound. Powder X-ray diffraction measurement (Example β1) of the resulting solid confirmed the following major peaks: 6.00°, 8.06°, 8.61°, 10.06°, 12.11°, 12.78°, 15.01°, 16.46°, 17.64°, and 18.09° (±0.2°). The measurement results are shown in Figure 55. The results of simultaneous thermogravimetry and differential thermal analysis (Example β9) of the resulting solid are shown in Figure 56. At around 100°C, a weight loss equivalent to that of the solvent was observed. １ The results of H-NMR (Example β18) are shown in Figure 57. A peak corresponding to p-toluenesulfonic acid equivalent to 1 equivalent of Compound 1 was observed, while peaks other than those corresponding to the measurement solvent water, DMSO, the internal standard TMS, and ethanol corresponding to the residual solvent were not observed. From the above, it was confirmed that the obtained solid was the tosylate hydrate crystal of Compound 1 (Form A).

[0182] Example α39 (1) Preparation of Dibesylate Hydrate Crystals of Compound 1 (Form A) Benzenesulfonic acid monohydrate (92.8 mg) was dissolved in ethyl acetate (2.4 mL). The above-described benzenesulfonic acid ethyl acetate solution (2.4 mL) was added to the free solvate crystals of Compound 1 (Form A, 79.9 mg) in a reaction vessel, and the mixture was stirred at 60°C for 30 minutes. The reaction solution was returned to room temperature and stirred for an additional 20 hours. The resulting solid was collected by filtration, washed with ethyl acetate, and then vacuum-dried for one day to obtain a solid of the title compound. Powder X-ray diffraction measurement (Example β3) of the resulting solid was performed. The measurement results are shown in Figure 58. The solid was confirmed to be a crystalline solid. The results of simultaneous thermogravimetry and differential thermal analysis (Example β9) of the resulting solid are shown in Figure 59. A weight loss due to the solvent was detected around 70°C, which corresponds to one water molecule in the dibesylate hydrate of Compound 1. １ The results of H-NMR (Example β18) are shown in Figure 60. A peak corresponding to benzenesulfonic acid equivalent to 2 equivalents of Compound 1 was observed, while no solvent peaks other than those of the measurement solvents DMSO and water and the internal standard TMS were observed. From the above, it was confirmed that the obtained solid was the dibesylate hydrate crystal of Compound 1 (Form A).

[0183] Example α40 (2) Preparation of Compound 1 Dibesylate Crystal (Form G) To confirm the change in crystal form due to temperature change of Compound 1 dibesylate hydrate crystal (Form A) under a nitrogen atmosphere, temperature-controlled powder X-ray diffraction measurement (Example β7) was performed. After heating to 150°C, the temperature was lowered to 25°C while maintaining the nitrogen atmosphere. The results are shown in Figure 61. It was confirmed that a new crystal form was obtained. When this crystal was removed from the laboratory environment, it rapidly transformed into Compound 1 dibesylate hydrate crystal (Form A). As shown in Figure 59, simultaneous thermogravimetry and differential thermal analysis of Compound 1 dibesylate hydrate crystal (Form A) revealed a dehydration peak near 60°C, the weight of which corresponded to one water molecule of Compound 1 dibesylate monohydrate. This confirmed that the compound was Compound 1 dibesylate crystal (Form G).

[0184] Example α41 (1) Preparation of Besylate Hydrate Crystals of Compound 1 (Form B and Form H) The free solvate crystals of Compound 1 (Form A, 41.2 mg) were dissolved in DMSO (0.66 mL), and 2 M aqueous benzenesulfonic acid solution (0.115 mL) was added. This solution (0.030 mL) was lyophilized at −20° C. for 3 days. Benzyl alcohol (0.015 mL) was added to the resulting lyophilized product, and the mixture was shaken and stirred at room temperature for 7 days to obtain a solid of Compound 1 (Sample E). Benzenesulfonic acid monohydrate (35.9 mg) was dissolved in a water-ethanol mixture (3:1 (v / v), 1.0 mL). The above-mentioned water-ethanol mixture of benzenesulfonic acid (3:1 (v / v), 1.0 mL) was added to the free solvate crystals of Compound 1 (Form A, 62.6 mg) in a reaction vessel, and the mixture was stirred at room temperature for 10 minutes. The above-mentioned wet powder of Sample E (approximately 0.1 mg) was added to the reaction solution, and the mixture was stirred at room temperature for 6 hours. The resulting wet solid was collected by filtration and dried in vacuo for 1 day to obtain a solid of the title compound. The results of powder X-ray diffraction measurement of the resulting wet solid (Example β3) are shown in Figure 62, and the results of measurement of the solid immediately after vacuum drying are shown in Figure 63. It was confirmed that the besylate dihydrate crystals of Compound 1 (Form B) transformed into the besylate hydrate crystals of Compound 1 (Form H) upon drying. The results of simultaneous thermogravimetry and differential thermal analysis (Example β9) of the resulting solid (Form B) are shown in Figure 64. Weight loss due to the solvent was observed around 70°C. The resulting solid (Form B) １ The results of H-NMR (Example β19) are shown in Figure 65. A peak corresponding to benzenesulfonic acid equivalent to one equivalent of the compound was observed, while no solvent peaks other than those of the measurement solvents DMSO and water and the internal standard TMS were observed. From the above, it was confirmed that the obtained solid was the besylate hydrate crystal of Compound 1 (Form B).

[0185] Example α42 (2) Preparation and Single Crystal Structure Analysis of Compound 1 Besylate Dihydrate Crystals (Form B) Compound 1 free solvate crystals (Form A, 41.2 mg) were dissolved in DMSO (0.66 mL), and 2 M benzenesulfonic acid aqueous solution (0.115 mL) was added. This solution (0.030 mL) was lyophilized at −20°C for 3 days. A water-ethanol mixture (3:1 (v / v), 0.015 mL) was added to the resulting lyophilized product, and the mixture was shaken and stirred at room temperature for 7 days to obtain single crystals of Compound 1. The resulting crystals were confirmed to be Compound 1 besylate dihydrate crystals (Form B) by single crystal X-ray structural analysis (Example β14). The structure is shown in Figure 66. A comparison of the powder X-ray diffraction pattern calculated from the crystal structure and the powder X-ray diffraction results of Compound 1 besylate hydrate crystals (Form B) is shown in Figure 67.

[0186] Example α43 (4) Preparation of Compound 1 Besylate Crystal (Form J) To confirm the change in crystal form due to temperature change of Compound 1 besylate hydrate crystal (Form B, described above) under a nitrogen atmosphere, temperature-controlled powder X-ray diffraction measurement (Example β8) was performed. After heating to 150°C, the temperature was lowered to 30°C while maintaining the nitrogen atmosphere. The results are shown in Figure 68. It was confirmed that a new crystal form was obtained. When this crystal was removed from the laboratory environment, it rapidly transformed into Compound 1 besylate hydrate crystal (Form B). As shown in Figure 64, simultaneous thermogravimetry and differential thermal analysis of Compound 1 besylate hydrate crystal (Form B) revealed a dehydration peak near 70°C, and the weight loss value corresponded to the water molecules of Compound 1 besylate hydrate. This confirmed that the compound was Compound 1 besylate crystal (Form J).

[0187] Example α44 (1) Preparation of Besylate Solvate of Compound 1 (Form C) The free solvate crystals of Compound 1 (Form A, 52.8 mg) were dissolved in DMSO (0.307 mL), and 2 M aqueous benzenesulfonic acid solution (0.073 mL) was added. This solution (0.015 mL) was lyophilized at −20° C. for 3 days. THF (0.015 mL) was added to the resulting lyophilized product, and the mixture was shaken and stirred at room temperature for 7 days to obtain a solid of Compound 1. The results of powder X-ray diffraction measurement of the resulting solid (Example β3) are shown in FIG. 69. It was confirmed to be a crystalline solid. The results of simultaneous thermogravimetry, differential thermal analysis, and mass spectrometry (Example β12) of the resulting solid are shown in FIG. 70. It was confirmed to be a besylate solvate crystal of Compound 1 (Form C).

[0188] Example α46 (1) Preparation of Compound 1 Besylate Hydrate Crystals (Form E) Compound 1 free solvate crystals (Form A, 52.8 mg) were dissolved in DMSO (0.307 mL), and 2 M benzenesulfonic acid aqueous solution (0.073 mL) was added. This solution (0.015 mL) was lyophilized at −20°C for 3 days. Acetone (0.015 mL) was added to the resulting lyophilized product, and the mixture was shaken and stirred at room temperature for 7 days to obtain a wet solid (Sample E1). Benzenesulfonic acid monohydrate (54.7 mg) was dissolved in acetone (1.03 mL). The aforementioned benzenesulfonic acid acetone solution (1.03 mL) was added to Compound 1 free solvate crystals (Form A, 51.6 mg) in a reaction vessel. The previously described sample E1 (approximately 0.1 mg) was then added and stirred at room temperature for 9 hours. After adding acetone (1.0 mL), the solid was collected by filtration, washed with acetone, and dried under reduced pressure to obtain a solid (sample E2). Benzenesulfonic acid monohydrate (67.2 mg) was dissolved in acetone (1.53 mL). The previously described benzenesulfonic acid acetone solution (1.53 mL) was added to the free solvate crystals of Compound 1 (Form A, 76.5 mg) in a reaction vessel. The previously described sample E2 (approximately 0.1 mg) was then added and stirred at room temperature for 3 hours. Acetone (1.53 mL) was then added and stirred for 2 hours, after which acetone (1.53 mL) was added and stirred for 1 hour. The resulting solid was collected by filtration, washed with acetone, and then vacuum-dried for 1 day to obtain a solid of the title compound. Powder X-ray diffraction analysis (Example β3) of the resulting solid was performed. The measurement results are shown in Figure 71. It was confirmed to be a crystalline solid. The results of simultaneous thermogravimetry and differential thermal analysis of the obtained solid (Example β9) are shown in Figure 72. Weight loss due to the solvent was detected around 70°C. １ The results of H-NMR (Example β18) are shown in Figure 73. A peak corresponding to benzenesulfonic acid equivalent to 1 equivalent of Compound 1 was observed, while no solvent peaks other than those of the measurement solvents DMSO and water, the internal standard TMS, and acetone, which is thought to be a residual solvent, were observed. From the above, it was confirmed that the obtained solid was the besylate hydrate crystal of Compound 1 (Form E).

[0189] Example α48 (1) Preparation of Compound 1 Hydrobromide Hydrate Crystals (Form A) DMSO (0.112 mL) was added to Compound 1 free solvate crystals (Form A, 22.5 mg) in a reaction vessel, and the mixture was heated and stirred at 70°C for 10 minutes to dissolve. After adding 2M aqueous hydrobromic acid (0.56 mL), a wet solid of Compound 1 hydrochloride hydrate (approximately 0.1 mg) was added and stirred at room temperature for 17 hours. The resulting solid was collected by filtration and vacuum-dried for 4 days to obtain a solid of the title compound. The results of powder X-ray diffraction analysis of the resulting solid (Example β3) are shown in Figure 74. It was confirmed to be a crystalline solid. The results of simultaneous thermogravimetry and differential thermal analysis of the resulting solid (Example β13) are shown in Figure 75. Ion peaks corresponding to the m / z of water and HBr were detected at around 100°C and around 310°C, respectively, and the weight loss values ​​correspond to the amounts of one water molecule and one HBr molecule per molecule of Compound 1. １ The results of H-NMR (Example β19) are shown in Figure 76. The content of Compound 1 was calculated to be 75%, which corresponds to the amount of Compound 1 contained in the hydrobromide hydrate of Compound 1. Furthermore, no solvent peaks were observed other than those of the measurement solvent DMSO, water, and the internal standard substances TMS and 1,3,5-trimethoxybenzene. From the above, it was confirmed that the obtained solid was the hydrobromide hydrate crystal of Compound 1 (Form A).

[0190] Preparation of Formulation for Dissolution Test Example α49 Preparation of Formulation (Sample F) Using Free-Form Hydrate Crystals of Compound 1 (Form B) The free-form hydrate crystals of Compound 1 (Form B, 25.35 mg) obtained in Example α6 and Pharmatose 200M (226.16 mg) were weighed into an agate mortar and mixed with a pestle for 15 minutes. The mixture was sieved through a sieve with 500 μm openings and then vacuum-dried overnight to obtain the title formulation.

[0191] Example α50 Preparation of a formulation (Sample G) using hydrochloride hydrate crystals of Compound 1 (Form A) The hydrochloride hydrate crystals of Compound 1 (Form A, 20.12 mg) obtained in Example α13 and Pharmatose 200M (181.07 mg) were weighed into an agate mortar and mixed with a pestle for 15 minutes. The mixture was sieved through a 500 μm mesh sieve and then vacuum dried overnight to obtain the title formulation.

[0192] Example α51 Preparation of a formulation (Sample H) using sulfate hydrate crystals of Compound 1 (Form A) The sulfate hydrate crystals of Compound 1 (Form A, 25.02 mg) obtained in Example α19 and Pharmatose 200M (225.44 mg) were weighed into an agate mortar and mixed with a pestle for 15 minutes. The mixture was sieved through a 500 μm mesh sieve and then vacuum dried overnight to obtain the title formulation.

[0193] Example α52 Preparation of a formulation (Sample J) using mesylate hydrate crystals of Compound 1 (Form F) The mesylate hydrate crystals of Compound 1 (Form F, 25.75 mg) obtained in Example α30 and Pharmatose 200M (240.22 mg) were weighed into an agate mortar and mixed with a pestle for 15 minutes. The mixture was sieved through a 500 μm mesh sieve and then vacuum dried overnight to obtain the title formulation.

[0194] Dissolution Test Example α53 Sample F (free form hydrate preparation), Sample G (hydrochloride hydrate preparation), Sample H (sulfate hydrate preparation), and Sample J (mesylate hydrate preparation) obtained in Examples α49 to α52 were subjected to a dissolution test according to the conditions described in Example β23 to evaluate the solubility of the crystals. The results are shown in Figure 77. The initial dissolution (dissolution concentration at 0-15 minutes) was similar for Sample F and Sample G, followed by Sample J and Sample H, in that order. However, it was confirmed that Sample G showed a higher dissolution concentration at 60-240 minutes than Sample F, Sample H, and Sample J.

[0195] The samples obtained above were analyzed under the following conditions: Powder X-ray diffraction measurements were carried out in a laboratory with a set temperature of 23°C.

[0196] Powder X-ray diffraction measurements of Example β1, Example α1, Example α2, Example α6, Example α16, Example α17, Example α19, Example α28, Example α30, and Example α35 were carried out under the following conditions: Measurement device: SmartLab System, D / Tex Ultra detector (manufactured by Rigaku Corporation) Radiation source: CuKα1 Tube voltage: 45 kV Tube current: 200 mA Scanning range: 5 to 31° Scanning speed: 0.5° / min Sampling width: 0.005°

[0197] Example β2 Powder X-ray diffraction measurement of Example α13 was carried out under the following conditions: Measurement device: SmartLab System, D / Tex Ultra detector (manufactured by Rigaku Corporation) Radiation source: CuKα1 Tube voltage: 45 kV Tube current: 200 mA Scanning range: 1.5 to 50° Scanning speed: 0.5° / min Sampling width: 0.02°

[0198] Powder X-ray diffraction measurements of Example β3, Example α2, Example α3, Example α7, Example α8, Example α9, Example α10, Example α11, Example α14, Example α19, Example α20, Example α22, Example α24, Example α25, Example α29, Example α30, Example α31, Example α32, Example α39, Example α41, Example α42, Example α44, Example α46, and Example α48 were carried out under the following conditions: Measurement equipment: D8 Discover, 2D VANTEC-500 solid state detector (manufactured by Bruker) Radiation source: CuKα Tube voltage, tube current: 50 kV, 1000 μA Measurement range: 5 to 31° Exposure time: 40 seconds or 120 seconds

[0199] Example β4 Powder X-ray diffraction measurements of Examples α4, α5, and α15 were carried out under the following conditions: Apparatus: Empyrean (PANalytical) Radiation source: CuKα Tube voltage, tube current: 45 kV, 40 mA Scanning range: 3 to 25° Scanning speed: 1.3° / sec Sampling width: 0.026°

[0200] Example β5 Powder X-ray diffraction measurement of Example α12 was carried out under the following conditions, and the crystal form at each temperature was evaluated under a nitrogen atmosphere. The sample temperature setting of the measurement device was raised from 35°C to 100°C, and the powder X-ray diffraction patterns were measured at 100°C (Figure 24) and 35°C (Figure 25) during the period until the temperature was lowered again to 35°C. Measurement device: SmartLab System, D / Tex Ultra detector, DSC attachment (manufactured by Rigaku Corporation) Radiation source: CuKα1 Tube voltage: 45 kV Tube current: 200 mA Scanning range: 5 to 31° Scanning speed: 0.5° / min Sampling width: 0.02° or 0.005° Measurement device temperature setting change conditions:

[0201]

[0202] Example β6 Powder X-ray diffraction measurement of Example α33 was carried out under the following conditions, and the crystal form at each temperature was evaluated under a nitrogen atmosphere. The sample temperature setting of the measurement device was raised from 30°C to 100°C, and the powder X-ray diffraction pattern at 100°C (Figure 54) was measured at the time point before the temperature was lowered again to 30°C. Measurement device: SmartLab System, D / Tex Ultra detector, DSC attachment (manufactured by Rigaku Corporation) Radiation source: CuKα1 Tube voltage: 45 kV Tube current: 200 mA Scanning range: 7 to 30° Scanning speed: 5.0° / min Sampling width: 0.02° Temperature setting change conditions of the measurement device:

[0203]

[0204] Example β7 Powder X-ray diffraction measurement of Example α40 was carried out under the following conditions, and the crystal form at each temperature was evaluated under a nitrogen atmosphere. The sample temperature setting of the measuring device was raised from 25°C to 150°C, and the powder X-ray diffraction pattern at 150°C (Figure 61) was measured at the time point before the temperature was lowered to 25°C again. Measuring device: SmartLab System, D / Tex Ultra detector, DSC attachment (manufactured by Rigaku Corporation) Radiation source: CuKα1 Tube voltage: 45 kV Tube current: 200 mA Scanning range: 5 to 40° Scanning speed: 5.0° / min Sampling width: 0.02° Temperature setting change conditions of the measuring device:

[0205]

[0206] Example β8 Powder X-ray diffraction measurement of Example α43 was carried out under the following conditions, and the crystal form at each temperature was evaluated under a nitrogen atmosphere. The sample temperature setting of the measuring device was raised from 30°C to 150°C, and the powder X-ray diffraction pattern at 150°C (Figure 68) was measured at the time point before the temperature was lowered again to 30°C. Measuring device: SmartLab System, D / Tex Ultra detector, DSC attachment (manufactured by Rigaku Corporation) Radiation source: CuKα1 Tube voltage: 45 kV Tube current: 200 mA Scanning range: 5 to 30° Scanning speed: 5.0° / min Sampling width: 0.02° Temperature setting change conditions of the measuring device:

[0207]

[0208] Example β9 Thermogravimetry and simultaneous differential thermal analysis (TG-DTA) for Examples α2, α6, α17, α19, α22, α28, α30, α35, α39, α41, and α46 were performed under the following conditions: Measurement device: STA7200RV+AS-3T (Hitachi High-Tech Science) Measurement range: 30 to 350°C Heating rate: 10°C / min Atmosphere: Nitrogen Measurement: The sample was weighed into an open aluminum pan, covered with a mesh, and then the measurement was performed.

[0209] Example β10 Thermogravimetry and simultaneous differential thermal analysis (TG-DTA) of Example α1 were carried out under the following conditions: Measuring device: STA7200RV+AS-3T (Hitachi High-Tech Science) Measurement range: 30 to 500°C Heating rate: 10°C / min Atmosphere: Nitrogen Measurement: The sample was weighed in an open aluminum pan, covered with a mesh, and then the measurement was carried out.

[0210] Example β11 Thermogravimetry and simultaneous differential thermal analysis (TG-DTA) of Example α1 were carried out under the following conditions. Measuring device: STA7200RV+AS-3T (Hitachi High-Tech Science) Measurement range: 30 to 310°C Heating rate: 20°C / min, left standing at 310°C for 20 minutes Atmosphere: Nitrogen Measurement: The sample was weighed into an open aluminum pan and the measurement was carried out. After the measurement was completed, the sample was recovered.

[0211] Example β12 Thermogravimetry, differential thermal analysis, and simultaneous mass spectrometry (TG-DSC-MS) of Examples α7, α8, α9, and α44 were performed under the following conditions. Measurement equipment: STA449F1 Jupiter (manufactured by NETZSCH) + JMS-Q1500GC (manufactured by JEOL) Measurement range: 50 to 350°C Heating rate: 10°C / min Atmosphere: Helium, 50 mL / min Transfer line temperature: 300°C Ion source temperature: 250°C Ionization method, voltage, current: EI, 70 eV, 20 μA Relative EM voltage: +700 V Measurement mode: scan (m / z 10-400) Measurement: The sample was filtered through a mesh or sintered filter, dried in vacuum overnight, and then placed on an open aluminum pan for measurement.

[0212] Example β13 Thermogravimetry, differential thermal analysis and simultaneous mass spectrometry (TG-DSC-MS) of Examples α10, α11, α13, α16 and α48 were carried out under the following conditions. Measurement equipment: STA449F1 Jupiter (manufactured by NETZSCH) + JMS-Q1500GC (manufactured by JEOL) Measurement range: 50 to 350°C Heating rate: 10°C / min Atmosphere: Helium, 50mL / min Transfer line temperature: 300°C Ion source temperature: 250°C Ionization method, voltage, current: EI, 70eV, 20μA Relative EM voltage: +700V Measurement mode: scan (m / z 10-400) Measurement: The sample was weighed into an open aluminum pan and the measurement was carried out.

[0213] Example β14 Single crystal X-ray structural analysis of Example α3 and Example α42 was carried out under the following conditions: Measuring device: Rigaku XtaLAB Synergy Custom with a VariMax Cu Diffractometer (manufactured by Rigaku Corporation) Radiation source: CuKα Tube voltage: 40 kV Tube current: 30 mA Temperature: −180° C. (Example α3) or 24° C. (Example α42) Measurement: Measurement was carried out using a strategy and exposure time that were considered to be sufficient to obtain diffraction spots for structural analysis. Structural analysis: Using the Olex2 program, initial structure determination was performed using the Intrinsic Phasing method (SHELXT-2018 / 2), and structure refinement was performed using the Full-matrix least-squares method (SHELXL-2018 / 3). Data collection and refinement statistics are shown in Table 7 below.

[0214]

[0215] Example β15 Single crystal X-ray structure analysis of Example α14 was carried out under the following conditions. Measurement equipment: High Energy Accelerator Research Organization Photon Factory BL-5A Radiation source: Synchrotron radiation (λ = 0.92 Å) Temperature: -178 ° C Measurement: Measurement was carried out using a strategy and exposure time that was thought to be sufficient to obtain diffraction spots for structural analysis. Structural analysis: Using the Olex2 program, initial structure determination was performed using the Intrinsic Phasing method (SHELXT-2018 / 2), and structural refinement was performed using the Full-matrix least-squares method (SHELXL-2018 / 3). Data collection and refinement statistics are shown in Table 8 below.

[0216]

[0217] Example β16 Analysis of Cl content in Examples α13 and α17 by ultra high performance liquid chromatography was carried out under the following conditions. HPLC analysis condition 1 Measurement apparatus: Shimadzu Nexera X3 UHPLC (Shimadzu), Corona Veo RS Charged Aerosol Detector (Thermo Fisher Scientific) Column: Acclaim Trinity P1 3 μm, 2.1 × 100 mm (Thermo Fisher Scientific) Detector: Corona Veo RS Charged Aerosol Detector Mobile phase: 20 mM ammonium acetate solution (pH = 6.6) / acetonitrile = 55 / 45 (isocratic conditions) Post-added solvent: acetonitrile Flow rate: 0.3 mL / min Injection volume: 5 μL Analysis time: 7 minutes Column temperature: 40°C CAD temperature: 35°C Sample treatment: 1 mg of sample was accurately weighed, 1 mL of 0.1 mg / mL hippuric acid methanol solution and 1 mL of methanol were added, and the mixture was stirred with a vortex mixer to prepare a sample solution (sample concentration: 0.5 mg / mL).

[0218] Example β17 Analysis of sulfate ion content by ultra high performance liquid chromatography in Examples α19 and α22 was carried out under the following conditions: HPLC analysis condition 2 Measurement equipment: Waters ACQUITY UPLC H-Class Plus (manufactured by Waters), Corona Veo RS Charged Aerosol Detector (manufactured by Thermo Fisher Scientific) Column: Acclaim Trinity P1 3 μm, 2.1 × 100 mm (manufactured by Thermo Fisher Scientific) Detector: Corona Veo RS Charged Aerosol Detector Mobile phase: Analysis was performed using the gradient shown in Table 9.

[0219] Post-addition solvent: water Flow rate: mobile phase 0.6 mL / min, post-addition solvent phase 0.4 mL / min Injection volume: 10 μL Analysis time: 15 min Column temperature: 40°C CAD temperature: 20°C Sample treatment: 1 mg of sample was accurately weighed, 1 mL of DMSO was added, and the mixture was stirred with a vortex mixer to prepare a sample solution (sample concentration 1.0 mg / mL).

[0220] Example β18 Example α1, Example α28, Example α30, Example α35, Example α39, Example α46 １ H-NMR measurements were carried out under the following conditions. Measurement apparatus: JNM-ECX500II (manufactured by JEOL Corporation) Measurement solvent: DMSO-d6, contains 0.03% (v / v) TMS Measurement temperature: 294-295K Sample preparation: A commercially available deuterated solvent was mixed with the compound to be measured to prepare the sample. Number of integrations, relaxation waiting time: Measurements were carried out the number of times (8 or 128 times) that were considered to provide a sufficient signal-to-noise ratio, and for a time (60 seconds) that was sufficiently longer than the relaxation time. Integration value: The equivalent ratio (mol %) of the salt was calculated based on the signal area intensity ratio of each signal.

[0221] Example β19 Example α6, Example α41, Example α48 １ H-NMR measurements were carried out under the following conditions. Measurement apparatus: JNM-ECX500II (manufactured by JEOL Corporation). Measurement solvent: DMSO-d6, contains 0.03% (v / v) TMS. Measurement temperature: 294-295 K. Sample preparation: 1,3,5-trimethoxybenzene was dissolved in a commercially available deuterated solvent. This solution was mixed with the compound to be measured to prepare the sample. Number of integrations and relaxation waiting time: Measurements were carried out the number of times (64) considered to obtain a sufficient signal-to-noise ratio, and for a time (60 seconds) sufficiently longer than the relaxation time. Integration value: Based on the signal area intensity ratio of each signal, the equivalent ratio (mol %) of the salt and the content of Compound 1 (wt %) in the sample were calculated.

[0222] Example β20 Example α22 １H-NMR measurements were carried out under the following conditions. Measurement apparatus: JNM-ECX500II (manufactured by JEOL Corporation) Measurement solvent: Methanol-d4: Deuterium oxide, contains 0.05% (v / v) 3-(trimethylsilyl)propionic-acid sodium salt mixed solution (9:1 (v / v)) Measurement temperature: 294 K Sample preparation: A commercially available deuterated solvent was mixed with the compound to be measured. Number of integrations, relaxation waiting time: Measurements were carried out the number of times (512) considered to obtain a sufficient S / N ratio, for a time (60 seconds) sufficiently longer than the relaxation time. Integration value: The equivalent ratio (mol%) of the salt was calculated based on the signal area intensity ratio of each signal.

[0223] Example β21 Example α4 １ H-NMR measurements were carried out under the following conditions. Measurement apparatus: JNM-ECZ500R (manufactured by JEOL Corporation) Measurement solvent: DMSO-d6, contains 0.03% (v / v) TMS Measurement temperature: 295 K Sample preparation: 1,3,5-trimethoxybenzene was dissolved in a commercially available deuterated solvent. This solution was mixed with the compound to be measured to prepare a sample. Number of integrations and relaxation waiting time: Measurements were carried out the number of times (16) considered to obtain a sufficient signal-to-noise ratio, and for a time (30 seconds) sufficiently longer than the relaxation time. Integration value: Based on the signal area intensity ratio of each signal, the equivalent ratio (mol %) of the salt and the content of Compound 1 (wt %) in the sample were calculated.

[0224] Example β22 The Karl Fischer titration of Examples α16 and α17 was carried out under the following conditions: Measuring device: MKC-710M (Kyoto Electronics Manufacturing Co., Ltd.) Anolyte: Chem-Aqua anolyte AGE Catholyte: Chem-Aqua catholyte CGE Measurement temperature: 25°C Measurement method: Approximately 10 mg of the sample was weighed into a weighing funnel, and the water content was measured by coulometric titration.

[0225] Example β23 The formulation components of the samples used in Example α53 are as follows. Sample F: Free form hydrate crystals of Compound 1 (Form B): Pharmatose 200M (1:9 (w / w)) Sample G: Hydrochloride hydrate crystals of Compound 1 (Form A): Pharmatose 200M (1:9 (w / w)) Sample H: Sulfate hydrate crystals of Compound 1 (Form A): Pharmatose 200M (1:9 (w / w)) Sample J: Mesylate hydrate crystals of Compound 1 (Form F): Pharmatose 200M (1:9 (w / w)) The dissolution test was carried out under the following conditions. Each of Sample F, Sample G, Sample H, and Sample J was weighed out in an amount of 4.55 mg, calculated as the weight of the free form of Compound 1. Each weighed sample was placed in a dissolution tester, and the solution was collected at each sampling point shown in Table 8. The collected solution was diluted three times with a DMSO solution containing 2-(1-naphthyl)ethanol, an internal standard, and subjected to analysis by high performance liquid chromatography. The dissolution test equipment and conditions used are shown in Table 10.

[0226]

[0227] Furthermore, the content analysis of Compound 1 in Example α53 by high performance liquid chromatography was carried out under the following conditions. Apparatus name: Waters ACQUITY UPLC H-Class Plus (manufactured by Waters) Column: ACQUITY UPLC BEH Shield RP18 Column, 130 Å, 1.7 μm, 2.1 × 50 mm (manufactured by Waters) Detector: ACQUITY UPLC PDA detector (manufactured by Waters) Mobile phase A: 0.05% trifluoroacetic acid / ultrapure water Mobile phase B: 0.05% trifluoroacetic acid / acetonitrile Sample temperature: 20°C Column temperature: 40°C Injection volume: 1 μL Absorption wavelength: compound (339 nm), internal standard (282 nm) Gradient table

[0228]

[0229] Test Example 1: Binding test of Compound 1 to the ATP-binding site of MYT1 and MYT1 kinase activity inhibition test 1. Experimental materials and methods (1) Binding test to the ATP-binding site of MYT1 The binding ability of Compound 1 to the ATP-binding site of MYT1 kinase protein was evaluated. 5X Kinase Buffer A (PV3189, manufactured by Thermo Fisher Scientific) was diluted 5-fold with Milli-Q water to prepare 1X Kinase Buffer. MYT1 protein (manufactured by Carnabiosciences) was diluted to a concentration of 0.005 μM, and Eu-Anti-GST Antibody was diluted to a concentration of 1 μM with 1X Kinase Buffer. In addition, Kinase Tracer 178 (PV5593, manufactured by Thermo Fisher Scientific) was diluted with 1x Kinase Buffer to a concentration of 0.1 μM. 2.5 μL of Compound 1 diluted with DMSO was added to a 96-well plate, followed by 5 μL of a 1:1 mixture of the diluted MYT1 protein and Eu-GST-Antibody. 2.5 μL of the diluted Kinase Tracer 178 was then added, mixed well, and allowed to stand at room temperature for 30 minutes. Subsequently, fluorescence at 665 nm and 615 nm generated by irradiation with 340 nm excitation light was detected using Envision (manufactured by PerkinElmer). The intensity of the fluorescence wavelength at 665 nm was divided by the intensity of the fluorescence wavelength at 615 nm to calculate the percentage of the tracer bound to the MYT1 protein for each condition. The signal without Compound 1 was set to 100% and the signal without MYT1 protein was set to 0%, and the inhibition rate was calculated when Compound 1 was added at each concentration, and the IC ５０ The results are shown in Table 12. IC of 10 μM or less ５０ When the above expression is observed, it can be determined that the compound exhibits binding activity to the ATP binding site of the MYT1 protein.

[0230] (2) MYT1 Kinase Activity Inhibition Test The ability of Compound 1 to inhibit MYT1 kinase activity was evaluated by assessing the phosphorylation level of Y15 of CDK1 protein by MYT1 protein using ELISA. The Cyclex Wee1 Kinase Assay / Inhibitor Screening Kit Ver. 3 (manufactured by MBL, CY-1172V3) kit was used. 10 μL of 5 nM MYT1 protein (manufactured by Carnabiosciences) was prepared using Kinase Buffer. 30 μL of 83.3 μM ATP was also prepared using Kinase Buffer. Compound 1 was diluted to each concentration in 10 μL increments using Kinase Buffer. 30 μL of ATP was added to the kit plate, followed by 10 μL of compound. 10 μL of MYT1 protein dilution was then added, mixed, and allowed to stand at room temperature for 60 minutes. The reaction solution was removed from each well, and the wells were washed with Wash Buffer. After removing the Wash Buffer, 100 μL of HRP-conjugated anti-phosphotyrosine antibody was added and allowed to stand at room temperature for 60 minutes. The reaction solution was then removed from the wells, and the wells were washed with Wash Buffer. After removing the Wash Buffer, 100 μL of Substrate Reagent was added to each well, and the plate was allowed to stand at room temperature for 8 minutes. Then, 100 μL of Stop Solution (1N sulfuric acid) was added to each well according to the protocol. The absorbance at 450 nm and 590 nm was measured using Envision (PerkinElmer). The absorbance at 590 nm was subtracted from the absorbance at 450 nm to evaluate the phosphorylation level of CDK1 protein in each well. The signal without Compound 1 was set to 100% and the signal without MYT1 protein was set to 0%, and the inhibition rate was calculated when Compound 1 was added at each concentration, and the IC ５０ The results are shown in Table 12. IC of 10 μM or less ５０ When the above expression is observed, it can be determined that the kinase activity of MYT1 protein is inhibited.

[0231]

[0232] 2. Results As shown in Table 3, Compound 1 had an IC value of 10 μM or less in the binding assay to the ATP binding site of MYT1. ５０ This suggests that it binds to the ATP binding site of the MYT1 protein. In addition, in a MYT1 kinase activity inhibition test, it had an IC value of 10 μM or less. ５０ These results suggest that the compound is capable of inhibiting the kinase activity of MYT1 protein, i.e., is a MYT1 inhibitor.

[0233] Test Example 2: Test of combined cytotoxic activity of pemetrexed and compound 1 1. Experimental materials and methods (1) Cell line The cell line LU65 (provided by the JCRB Cell Bank) was used for the evaluation of the present invention. LU65 is a lung cancer cell line with a truncated S82* deletion mutation in the RB1 gene (see Nature. 2019 May; 569 (7757): 503-508 (Non-Patent Document 22)), resulting in reduced RB1 function. Cell culture was performed using RPMI-1640 medium (manufactured by SIGMA) containing 10% FBS (manufactured by SIGMA).

[0234] (2) Cell Damage Test by Pemetrexed with Compound 1 Added Pemetrexed (Tokyo Chemical Industry Co., Ltd.) and Compound 1 at various concentrations were diluted with DMSO, and 0.04 μL of the dilution series and DMSO were added to each well of a cell culture plate using an Echo 555 Liquid Handler (Beckman Coulter). 1,000 LU65 cells (40 μL) were added to each well, and the cells were incubated at 37°C and 5% CO ２The cells were cultured in an incubator for 7 days. The amount of intracellular ATP, a marker of cell viability, was measured and used to monitor cell number. After culture, 20 μL of CellTiter-Glo® 2.0 (Promega) was added to each well, and luminescence intensity was measured using a Multimode Plate Reader EnVision® Xcite (PerkinElmer). The luminescence in the DMSO-added wells minus the luminescence in the non-seeded wells was set to 100%, and the viability under each condition was calculated by calculating the percentage of the luminescence detected in each well minus the luminescence in the non-seeded wells. Furthermore, the cytotoxic activity under each condition was calculated by subtracting the viability for each combination of pemetrexed treatment concentration and Compound 1 treatment concentration from 100%. In addition, based on the cytotoxic activity of wells without compound 1 at each concentration of pemetrexed and the cytotoxic activity of wells without pemetrexed at each concentration of compound 1, the Bliss score based on Bliss independence, which is an index of the combined effect at each combination of each concentration of pemetrexed and each concentration of compound 1 (PLoS Comput Biol. 2019 May 20; 15 (5): e1006752. (Non-Patent Document 6)), was calculated, and the maximum value and the concentrations of pemetrexed and compound 1 showing the maximum value are listed in Table 13. In addition, the HSA score, which is an index of combined effect different from Bliss independence (PLoS Comput Biol. 2019 May 20; 15 (5): e1006752. (Non-Patent Document 6)), was also calculated, and the maximum value and the concentrations of pemetrexed and compound 1 showing the maximum value are listed in Table 13. When both the maximum Bliss score and the maximum HSA score are 10% or more, it can be determined that the combined use is effective.

[0235]

[0236] 2. Results As shown in Figure 78 and Table 13, in the pemetrexed cytotoxicity test when Compound 1 was added, the maximum Bliss score and the maximum HSA score were both 10% or more, suggesting that Compound 1 inhibits the function of MYT1 in cells and exerts a combined effect with pemetrexed.

[0237] The present invention provides a compound having MYT1 inhibitory activity, or a salt thereof, or a crystal of a solvate thereof, and also provides a pharmaceutical useful for treating and preventing cancer, which contains the compound or salt as an active ingredient.

Claims

The following formula (1) A crystal of a compound represented by the formula: 　 The crystal according to claim 1, wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is a salt of the compound represented by formula (1). 　 The crystal according to claim 1, wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is a solvate of the compound represented by formula (1) or a solvate of a salt of the compound. 　 The crystal according to claim 1 or 3, wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is at least one solvate selected from the group consisting of the compound represented by formula (1) and hydrochloride, hydrobromide, hydroiodide, sulfate, mesylate, esylate, tosylate, besylate, phosphate, nitrate, acetate, oxalate, maleate, fumarate, citrate, malate, malonate, gluconate, mandelate, salicylate, fluoroacetate, trifluoroacetate, camsylate, tartrate, propionate, glutarate, lithium salt, sodium salt, potassium salt, cesium salt, rubidium salt, magnesium salt, calcium salt, strontium salt, barium salt, alkylammonium salt, dialkylammonium salt, trialkylammonium salt, and tetraalkylammonium salt of the compound. 　 The crystal according to claim 1 or 3, wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is a solvate of the compound represented by formula (1) or a salt of the compound with a solvent selected from the group consisting of water, methanol, ethanol, 1-propanol, 2-propanol, 1-butanol, 2-butanol, benzyl alcohol, acetonitrile, ethyl acetate, n-propyl acetate, n-butyl acetate, tetrahydrofuran, 1,4-dioxane, t-butyl methyl ether, dichloromethane, dimethyl sulfoxide, acetic acid, formic acid, acetone, 2-butanone, methyl isobutyl ketone, anisole, toluene, chlorobenzene, formamide, dimethylformamide, and dimethylacetamide. 　 The crystal according to claim 1 or 3, wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is a solvate of at least one selected from the group consisting of the compound represented by formula (1) and hydrochloride, hydrobromide, sulfate, mesylate, tosylate, and besylate salts of the compound, and a solvent selected from the group consisting of water, ethanol, 2-propanol, 2-butanol, dimethyl sulfoxide, acetic acid, and formic acid. 　 The crystal according to claim 1 or 3, wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is a hydrate of the hydrochloride of the compound represented by formula (1). 　 The crystal according to claim 1 or 3, wherein the compound represented by formula (1) or a salt thereof, or a solvate thereof is a monohydrate of the hydrochloride salt of the compound represented by formula (1). 　 The crystal according to any one of claims 1 to 8, wherein the crystal comprises at least one peak selected from the group consisting of 6.59°, 9.19°, 9.70°, 12.70°, 13.15°, 13.29°, 13.80°, 14.93°, 15.80°, and 16.45° (±0.2) as a diffraction angle (2θ value) in powder X-ray diffraction at 23°C. 　 A pharmaceutical composition comprising a crystal of the compound or a salt thereof, or a solvate thereof, according to any one of claims 1 to 9. 　 The pharmaceutical composition according to claim 10 for the prevention and / or treatment of cancer. 　 A method for preventing and / or treating cancer, comprising administering to a subject an effective amount of the compound according to any one of claims 1 to 9, or a salt thereof, or a crystal of a solvate thereof. 　 A crystalline compound according to any one of claims 1 to 9, or a salt thereof, or a solvate thereof, for use in the prevention and / or treatment of cancer. 　 Use of a crystal of the compound or a salt thereof, or a solvate thereof according to any one of claims 1 to 9, for the manufacture of a pharmaceutical composition for the prevention and / or treatment of cancer. 　 The following formula (1) or a salt thereof, or a solvate thereof, as an active ingredient, the method comprising the step of mixing a crystal of the compound or salt thereof, or a solvate thereof, according to any one of claims 1 to 9, with a pharmaceutically acceptable carrier or medium.

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