The S4 crystal form of (1R,2S)-(E)-2-(3-(4-((cis-2,6-dimethylmorpholino)methyl)styryl)-1H-imidazol-6-yl)-5'-methoxispiro[cyclopropane-1,3'-indoline]-2'-one fumarate, which is a PLK4 inhibitor

Abstract

Disclosed is the S4 crystalline form of the fumarate salt of Compound (I), represented by the following structural formula: (I). The molar ratio of Compound (I) to fumaric acid is 1.0:1.0. The S4 crystalline form is characterized by an X-ray powder diffraction pattern containing peaks at 2θ of 6.6°, 9.8°, 16.3°, 21.1°, 28.7°, and 30.2°±0.2. TIFF2022530028000022.tif74128

Description

Technical Field

[0001] Cross - Reference to Related Applications This application claims priority based on U.S. Provisional Application No. 62 / 837,858, filed on April 24, 2019. The entire content of the above - mentioned application is incorporated herein by reference.

Background Art

[0002] Background The polo - like kinase (PLK) family, which are serine / threonine kinases, includes at least four known members: PLK1, PLK2 (also known as Snk), PLK3 (also known as Fnk or Prk), and PLK4 (also known as Sak). Agents that inhibit PLK4 have the potential to treat cancer. A number of potent PLK4 inhibitors are disclosed in U.S. Patent No. 8,263,596 (Patent Document 1); U.S. Patent No. 8,481,525 (Patent Document 2); and U.S. Patent No. 8,481,533 (Patent Document 3) (the entire teachings of which are incorporated herein by reference). The structure of one of the inhibitors disclosed in these patents is shown below as compound (I). TIFF0007705351000001.tif39128

[0003] There is a need for salt forms of this compound that are crystalline and have other physical properties that enable production - scale manufacturing. There is also a need for pharmaceutical formulations in which this drug candidate is stable and can be effectively delivered to patients.

[0004] In this regard, U.S. Patent No. 9,884,855 (Patent Document 4) discloses some crystalline forms of the 1:1 fumarate of compound (I), including the D - form, as potential anti - cancer drug candidates. The entire teachings of U.S. Patent No. 9,884,855 (Patent Document 4) are incorporated herein by reference. "1:1" refers to the molar ratio of fumaric acid to compound (I).

Prior Art Documents

Patent Documents

[0005] [Patent Document 1] U.S. Patent No. 8,263,596 [Patent Document 2] U.S. Patent No. 8,481,525 [Patent Document 3] U.S. Patent No. 8,481,533 [Patent Document 4] U.S. Patent No. 9,884,855 [Summary of the Invention]

[0006] The D form can be produced in a crystalline state, has low hygroscopicity, and is an effective anticancer agent with good solubility and favorable pharmacokinetic properties, as disclosed in U.S. Patent No. 9,884,855, and thus was selected for scale-up. However, it was later found that the production of the D form in the large-scale production process results in a mixture of different crystal forms (see Example 3).

[0007] It has been discovered in the present invention (see Examples 6 and 7) that the new crystal form called "S4" has the beneficial properties of the D form (e.g., has low hygroscopicity and good solubility and possesses excellent pharmacokinetic properties). More importantly, it overcomes the problems associated with the D form. Specifically, the S4 form can be prepared in high purity with good yield, enabling scale-up (see Examples 4 and 5).

[0008] Accordingly, the present disclosure provides a fumarate salt of compound (I) having a molar ratio of compound (I) to fumaric acid of 1:1, the S4 crystal form characterized by an X-ray powder diffraction (XRPD) pattern including peaks at 2θ of 6.6°, 9.8°, 16.3°, 21.1°, 28.7°, and 30.2° ± 0.2.

[0009] The present disclosure also provides a pharmaceutical composition comprising the S4 crystalline form of the 1:1 fumarate of compound (I) and a pharmaceutically acceptable carrier or diluent.

[0010] The present disclosure also provides a method for preparing the S4 crystalline form of the 1:1 fumarate of compound (I). The method includes dissolving fumaric acid and compound (I) in a dissolving solvent comprising 2-butanone, water, and ethanol for forming a crystallization solution, and then precipitating the S4 crystalline form from the crystallization solution, wherein the molar ratio of compound (I) to fumaric acid in the dissolving solvent is about 1:1 to about 1:1.3; and the molar ratio of compound (I) to fumaric acid in the compound (I) fumarate is 1:1. In one aspect, the dissolution of fumaric acid and compound (I) is carried out at an elevated temperature, and the precipitation of the S4 crystalline form from the crystallization solution is by cooling and / or addition of methylcyclohexane while cooling.

[0011] A method for treating cancer is also provided. The method includes administering an effective amount of the S4 crystalline form of the 1:1 fumarate of compound (I) to a subject having cancer.

[0012] The use of the S4 crystalline form of the fumarate of compound (I) for the production of a medicament for treating cancer is also provided.

[0013] The S4 crystalline form of the fumarate of compound (I) for use in the treatment of cancer is also provided. [Invention 1001] The following structural formula: TIFF0007705351000002.tif45128 A fumarate of compound (I) represented by the formula, wherein the molar ratio of compound (I) to fumaric acid is 1:1, and the fumarate contains an S4 crystal form characterized by an X-ray powder diffraction pattern including peaks at 2θ of 6.6°, 9.8°, 16.3°, 21.1°, 28.7°, and 30.2° ± 0.2. [Invention 1002] The fumarate of Invention 1001, containing an S4 crystal form characterized by an X-ray powder diffraction pattern including peaks at 2θ of 6.6°, 9.8°, 16.3°, 21.1°, 28.7°, 29.4°, and 30.2° ± 0.2. [Invention 1003] The fumarate of Invention 1001, containing an S4 crystal form characterized by an X-ray powder diffraction pattern including peaks at 2θ of 6.6°, 9.8°, 13.0°, 16.3°, 19.5°, 21.1°, 22.5°, 28.7°, 29.4°, and 30.2° ± 0.2. [Invention 1004] The fumarate of Invention 1001, containing an S4 crystal form characterized by an X-ray powder diffraction pattern including peaks at 2θ of 6.6°, 9.8°, 13.0°, 16.3°, 19.5°, 21.1°, 22.5°, 22.9°, 23.9°, 28.7°, 29.4°, and 30.2° ± 0.2. [Invention 1005] The fumarate of Invention 1001, containing an S4 crystal form characterized by an X-ray powder diffraction pattern including peaks at 2θ of 6.6°, 9.8°, 11.6°, 13.0°, 16.3°, 17.1°, 19.5°, 21.1°, 21.5°, 22.0°, 22.5°, 22.9°, 23.9°, 24.3°, 28.7°, 29.4°, and 30.2° ± 0.2. [Invention 1006] The fumarate of Invention 1001, containing an S4 crystal form characterized by an X-ray powder diffraction pattern substantially similar to that of Figure 1. [Invention 1007] The fumarate of any one of Inventions 1001 to 1006, wherein at least 90% by weight of the salt is in a single crystal form, the S4 form. [Invention 1008] The fumarate of any one of Inventions 1001 to 1006, wherein at least 99% by weight of the salt is in a single crystal form, the S4 form. [Invention 1009] The fumarate of any one of Inventions 1001 to 1008, containing less than 30% by weight of the D crystal form. [Invention 1010] The fumarate of any one of Inventions 1001 to 1008, containing less than 10% by weight of the D crystal form. [Invention 1011] A pharmaceutical composition comprising any fumarate of the present invention 1001 - 1010, and a pharmaceutically acceptable carrier or diluent. [The present invention 1012] A method for preparing the S4 crystal form of the fumarate of compound (I), comprising: dissolving fumaric acid and compound (I) in a dissolving solvent containing 2 - butanone, water, and ethanol for forming a crystallization solution; TIFF0007705351000003.tif38128 and then, a step of precipitating the S4 crystal form from the crystallization solution, wherein the molar ratio of compound (I) to fumaric acid in the dissolving solvent is about 1:1 to about 1:1.3; and the molar ratio of compound (I) to fumaric acid in the fumarate is 1:1. [The present invention 1013] The method of the present invention 1012, comprising: dissolving fumaric acid and compound (I) in a dissolving solvent at an elevated temperature, and precipitating the S4 crystal form from the crystallization solution by cooling and / or adding methylcyclohexane while cooling. [The present invention 1014] The method of the present invention 1012 or 1013, comprising the following steps: (i) Dissolving compound (I) in a mixture of 2 - butanone and water at an elevated temperature for forming a compound (I) solution; (ii) Dissolving fumaric acid in ethanol at an elevated temperature for forming a fumaric acid solution; (iii) Adding the fumaric acid solution to the compound (I) solution at an elevated temperature for forming a crystallization solution; (iv) Optionally, adding seed crystals to the crystallization solution at an elevated temperature; and (v) Lowering the temperature of the crystallization solution to precipitate the S4 crystal form. [The present invention 1015] The method of the present invention 1014, wherein the volume ratio of 2 - butanone to water is 80:20 to 98:2, the volume ratio of 2 - butanone / water to ethanol to methylcyclohexane is 6 - 7 to 4 - 5 to 11 - 22; the elevated temperature is 45°C - 55°C; and the lowered temperature is 20°C - 30°C. [The present invention 1016] The method of the present invention 1014, wherein the volume ratio of 2 - butanone to water is 90:10 to 98:2, the volume ratio of 2 - butanone / water to ethanol to methylcyclohexane is 6 - 7 to 4 - 5 to 11 - 22; the elevated temperature is 45°C - 55°C; and the lowered temperature is 20°C - 30°C. [The present invention 1017] The method of the present invention 1014, further comprising adding methylcyclohexane to the crystallization solution at a lowered temperature to precipitate the S4 crystal form. [The present invention 1018] The method of the present invention 1017 of further lowering the temperature of the crystallization solution to precipitate the S4 crystalline form. [The present invention 1019] The method of the present invention 1017 where the lowered temperature is 20°C to 30°C. [The present invention 1020] The method of the present invention 1018 where the further lowered temperature is 0°C to 10°C. [The present invention 1021] A method of treating a subject having cancer, comprising the step of administering to the subject an effective amount of any of the fumarates of the present invention 1001 to 1010 or a pharmaceutical composition thereof. [The present invention 1022] The method of the present invention 1021, wherein the cancer is selected from the group consisting of lung cancer, breast cancer, colon cancer, neuroblastoma, prostate cancer, melanoma, glioblastoma multiforme, ovarian cancer, lymphoma, leukemia, osteosarcoma, embryonal cell tumor, glioma, fibrosarcoma, gastrointestinal sarcoma, fibrous histiocytoma, round cell sarcoma, synovial sarcoma, cervical cancer, anogenital cancer, head and neck cancer, and hypopharyngeal cancer. [The present invention 1023] The method of the present invention 1021, wherein the cancer is selected from the group consisting of lung cancer, breast cancer, and colon cancer. [The present invention 1024] The method of the present invention 1021, wherein the cancer is breast cancer. [The present invention 1025] The method of the present invention 1021, wherein the cancer is triple-negative breast cancer. [The present invention 1026] The method of the present invention 1025, wherein the triple-negative breast cancer is inoperable or metastatic. [The present invention 1027] A method of treating a patient having pancreatic cancer, comprising the step of administering to the patient an effective amount of any of the fumarates of the present invention 1001 to 1010 or a pharmaceutical composition thereof.

Brief Description of the Drawings

[0014] [Figure 1] The X-ray powder diffraction pattern (XRPD) of the S4 crystalline form of the 1:1 fumarate salt of compound (I) from 100 g production is illustrated. [Figure 2] The X-ray powder diffraction pattern (XRPD) of the S4 crystalline form of the 1:1 fumarate salt of compound (I) from large-scale production in Example 5 is illustrated. [Figure 3] Exposures normalized by dose after dosing of the D-form and S4-form are shown.

Modes for Carrying Out the Invention

[0015] Detailed Description of the Invention The present disclosure provides a novel S4 crystalline form of the 1:1 fumarate salt of compound (I) and corresponding pharmaceutical compositions. The present disclosure also provides a novel method for preparing the S4 crystalline form in a reproducible and scale-upable manner with excellent yields and purity. Further, the present disclosure provides a method for treating cancer.

[0016] Crystal form of the 1:1 fumarate of compound (I) In some embodiments, at least a specified weight percentage of the 1:1 fumarate of compound (I) is in a single crystal form. Specified weight percentages include 70%, 72%, 75%, 77%, 80%, 82%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.5%, 99.9%, or weight percentage ranges of 70% - 75%, 75% - 80%, 80% - 85%, 85% - 90%, 90% - 95%, 95% - 100%, 70 - 80%, 80 - 90%, 90 - 100%. For example, in one embodiment, at least 80 wt% (e.g., at least 90 wt% or 99 wt%) of the 1:1 compound (I) fumarate is in a single crystal form. It should be understood that all values and ranges between these values and ranges are intended to be encompassed by this disclosure.

[0017] As used herein, "crystalline" refers to a solid having a crystal structure in which individual molecules have a highly uniform, regular, fixed chemical arrangement. The crystalline 1:1 fumarate of compound (I) can be a crystal of a single crystal form of the 1:1 fumarate of compound (I), or a mixture of crystals of different single crystal forms. A single crystal form means the 1:1 fumarate of compound (I) as a single crystal, or as a plurality of crystals each having the same crystal form.

[0018] When a specific weight percentage of the 1:1 fumarate of compound (I) is in a single crystalline form, the remaining fumarate is some combination of amorphous fumarate and / or one or more other crystalline forms of the 1:1 compound (I) fumarate excluding its single crystalline form. When the crystalline 1:1 fumarate of compound (I) is defined as a specified percentage of one specific crystalline form of the 1:1 fumarate of compound (I), the remainder is composed of the amorphous form and / or crystalline forms other than the specified one or more specific forms. Examples of single crystalline forms include the S4 form of the 1:1 compound (I) fumarate characterized by one or more properties described herein.

[0019] In another aspect, less than 30 wt%, 25 wt%, 20 wt%, 10 wt%, 9 wt%, 8 wt%, 7 wt%, 6 wt%, 5 wt%, 4 wt%, 3 wt%, 2 wt%, 1 wt%, 0.5 wt% of the 1:1 fumarate of compound (I) is in the D crystalline form. The amount of the S4 form relative to the D form in a sample can be assayed by preparing a series of mixtures of the S4 and D forms in known weight ratios and obtaining an XRPD spectrum for each. The relative amount of the S4 form relative to the D form in a sample is assayed by selecting one or more characteristic peaks of the S4 and D forms and correlating the relative intensities of the sample XRPD to the relative intensities of the mixture XRPD. Characteristic XRPD peaks for the D crystalline form are provided in Table 4 of U.S. Patent No. 9,884,855.

[0020] The purity of the 1:1 fumarate of compound (I) relative to other stereoisomers, i.e., the ratio of the weight of that stereoisomer to the weight of all stereoisomers, is at least 60 wt%, 70 wt%, 80 wt%, 90 wt%, 99 wt%, or 99.9 wt%.

[0021] Preparation of the S4 Crystalline Form of the 1:1 Fumarate of Compound (I) Solids of the 1:1 fumarate of compound (I) can be prepared, for example, by slow evaporation, slow cooling, and anti-solvent precipitation. The present disclosure provides a novel crystallization technique for preparing the S4 crystalline form in a reproducible and scalable manner with improved yields and purity.

[0022] As used herein, "poor solvent" refers to a solvent that causes the fumarate of compound (I) to precipitate from solution in the form of fine powder or crystals because the 1:1 fumarate of compound (I) has low solubility.

[0023] Alternatively, the 1:1 fumarate of compound (I) may be recrystallized from a suitable solvent, with or without the addition of seed crystals.

[0024] In one embodiment, the S4 crystal form of the 1:1 fumarate of compound (I) can be prepared by dissolving fumaric acid and compound (I) in a dissolving solvent containing 2-butanone, water, and ethanol for the formation of a crystallization solution, and then precipitating the S4 crystal form from the crystallization solution. The molar ratio of compound (I) to fumaric acid in the dissolving solvent is from about 1:1 to about 1:1.3; the molar ratio of compound (I) to fumaric acid in the compound (I) fumarate is 1:1. In one aspect of this embodiment, the dissolution of fumaric acid and compound (I) is carried out at an elevated temperature, and the precipitation of the S4 crystal form from the crystallization solution is by cooling and / or the addition of methylcyclohexane while cooling.

[0025] In another aspect, the S4 crystalline form of the 1:1 fumarate salt of compound (I) is prepared by the steps of: (i) dissolving compound (I) in a mixture of 2-butanone and water at an elevated temperature to form a compound (I) solution; (ii) dissolving fumaric acid in ethanol at an elevated temperature to form a fumaric acid solution; (iii) adding the fumaric acid solution to the compound (I) solution at an elevated temperature to form a crystallization solution; (iv) optionally, adding seed crystals to the crystallization solution at an elevated temperature; and (v) lowering the temperature of the crystallization solution to precipitate the S4 crystalline form. In one aspect of this embodiment, the volume ratio of 2-butanone to water is from 80:20 to 98:2 (e.g., from 90:10 to 98:2), and the volume ratio of 2-butanone / water to ethanol to methylcyclohexane is from 6 to 7 to 4 to 5 to 11 to 22; the elevated temperature is from 45 °C to 55 °C; and the lowered temperature is from 20 °C to 30 °C. In another aspect of this embodiment, the method further comprises the step of adding methylcyclohexane to the crystallization solution at the lowered temperature to precipitate the S4 crystalline form. In one specific aspect, the lowered temperature is from 20 °C to 30 °C. In another specific aspect, the temperature of the crystallization solution is further lowered to precipitate the S4 crystalline form. In a more specific aspect, the further lowered temperature is from 0 °C to 10 °C.

[0026] Characterization of the Single Crystal Form S4 of the 1:1 Fumarate Salt of Compound (I) In one embodiment, the S4 form exhibits a unique XRPD pattern that includes sharp peaks corresponding to angular peak positions (2θ) indicative of a highly crystalline material and a flat baseline (see Figure 1). The XRPD pattern is obtained from a copper radiation source (CuKα; λ = 1.54056 Å) operating at 40 V / 30 mA. In one specific embodiment, the S4 form is a single crystal form of the 1:1 fumarate of compound (I) characterized by an X-ray powder diffraction pattern that includes peaks at 6.6°, 9.8°, 16.3°, 21.1°, 28.7°, and 30.2°. In another specific embodiment, the S4 form is characterized by an X-ray powder diffraction pattern that includes peaks at 6.6°, 9.8°, 16.3°, 21.1°, 28.7°, 29.4°, and 30.2°. In another specific embodiment, the S4 form is characterized by an X-ray powder diffraction pattern that includes peaks at 6.6°, 9.8°, 13.0°, 16.3°, 19.5°, 21.1°, 22.5°, 28.7°, 29.4°, and 30.2°. In yet another specific embodiment, the S4 form is characterized by an X-ray powder diffraction pattern that includes peaks at 6.6°, 9.8°, 13.0°, 16.3°, 19.5°, 21.1°, 22.5°, 22.9°, 23.9°, 28.7°, 29.4°, and 30.2°. In yet another specific embodiment, the S4 form is characterized by an X-ray powder diffraction pattern that includes peaks at 6.6°, 9.8°, 11.6°, 13.0°, 16.3°, 17.1°, 19.5°, 21.1°, 21.5°, 22.0°, 22.5°, 22.9°, 23.9°, 24.3°, 28.7°, 29.4°, and 30.2°.

[0027] In yet another specific embodiment, the S4 form is characterized by an X-ray powder diffraction pattern that is substantially similar to Figure 1.

[0028] As used herein, an X-ray powder diffractogram is "substantially similar to that of [a particular] figure" when at least 90%, such as at least 95%, at least 98%, or at least 99% of the signals in two diffractograms are the same at ±0.2 (2θ). Those skilled in the art will understand that in the determination of "similarity", even with the same crystal form, there can be variations in intensity and / or signal position in the XRPD diffractogram. Thus, those skilled in the art will understand that the signal maximum in the XRPD diffractogram (at degree two theta (°2θ) as referred to herein) generally means the reported value ±0.2 degrees two theta (the variance recognized in the art as described below) for the reported value.

[0029] It is well known in the field of crystallography that even a particular crystal form can have a slight variation in the angular peak position due to factors such as temperature variations, sample substitution, and the presence or absence of an internal standard. In the present disclosure, the variability of the angular peak position is ±0.2 (2θ). Further, due to differences in crystallite size and non-random crystallite orientation in sample preparation for XRPD analysis, the relative peak intensity for a particular crystal form can vary. It is well known in the art that this variability can account for the above factors without interfering with the clear identification of the crystal form.

[0030] Method of treatment PLK4, as a member of the polo family of serine / threonine kinases, is known to be involved in cell mitotic progression. Thus, small molecule inhibitors of this enzyme can be potential anti-cancer agents.

[0031] The present disclosure provides a method of treating a subject having a disease that can be alleviated by inhibition of PLK4 by administering an effective amount of the S4 form to the subject, such as a method of treating cancer or inhibiting tumor growth. Thus, the S4 form inhibits tumor growth by inducing apoptosis of tumor cells or inhibiting the proliferation of tumor cells.

[0032] Specific cancers that can be treated by the disclosed method include lung cancer, breast cancer, colon cancer, brain cancer, neuroblastoma, prostate cancer, melanoma, glioblastoma multiforme, ovarian cancer, lymphoma, leukemia, melanoma, sarcoma, tumor-associated lesions, osteosarcoma, germ cell tumor, glioma, and mesothelioma. In one specific embodiment, the cancer is lung cancer, breast cancer, colon cancer, neuroblastoma, prostate cancer, melanoma, glioblastoma multiforme, ovarian cancer, lymphoma, leukemia, osteosarcoma, germ cell tumor, glioma, fibrosarcoma, gastrointestinal sarcoma, fibrous histiocytoma, round cell sarcoma, synovial sarcoma, cervical cancer, anogenital cancer, head and neck cancer, and hypopharyngeal cancer. In one specific embodiment, the cancer is lung cancer, colon cancer, brain cancer, neuroblastoma, prostate cancer, melanoma, glioblastoma multiforme, or ovarian cancer. In another specific embodiment, the cancer is lung cancer, breast cancer, colon cancer, brain cancer, neuroblastoma, prostate cancer, melanoma, glioblastoma multiforme, or ovarian cancer. In another specific embodiment, the cancer is lung cancer, breast cancer, and colon cancer. In yet another specific embodiment, the cancer is breast cancer. In yet another specific embodiment, the cancer is basal subtype breast cancer or luminal B subtype breast cancer. In one embodiment, basal subtype breast cancer is breast cancer that is negative for ER (estrogen receptor), HER2, and PR (progesterone receptor). In yet another specific embodiment, the cancer is triple-negative breast cancer. The term "triple-negative breast cancer" refers to breast cancer that has been determined to be negative for hormones, estrogen, progesterone, and HER2 under the ASCO / CAP Clinical Practice Guideline. In some embodiments, triple-negative breast cancer is inoperable or metastatic. In a specific embodiment, the aforementioned method is used to treat a subject having pancreatic cancer, which is an adenocarcinoma. In another specific embodiment, the pancreatic cancer is hypoxic. In yet another specific embodiment, the pancreatic cancer is non-hypoxic.

[0033] Soft tissue cancers can also be treated by the disclosed methods. "Soft tissue cancer" is a term recognized in the art that encompasses tumors originating from any soft tissue of the body. Such soft tissues connect, support, or surround various structures and organs of the body, such as, but not limited to, smooth muscle, skeletal muscle, tendons, fibrous tissue, adipose tissue, blood vessels and lymphatic vessels, perivascular tissue, nerves, mesenchymal cells, and synovial tissue. Thus, soft tissue cancers can be cancers of adipose tissue, muscle tissue, nerve tissue, joint tissue, blood vessels, lymphatic vessels, and fibrous tissue. Soft tissue cancers can be either benign or malignant. Generally, malignant soft tissue cancers are called sarcomas or soft tissue sarcomas. There are many types of soft tissue tumors, including lipoma, lipoblastoma, brown fat tumor, liposarcoma, leiomyoma, leiomyosarcoma, rhabdomyoma, rhabdomyosarcoma, neurofibroma, schwannoma (neurilemmoma), neuroma, malignant schwannoma, neurofibrosarcoma, neurogenic sarcoma, nodular tenosynovitis, synovial sarcoma, hemangioma, glomus tumor, hemangiopericytoma, hemangioendothelioma, angiosarcoma, Kaposi's sarcoma, lymphangioma, fibroma, elastofibroma, superficial fibromatosis, fibrous histiocytoma, fibrosarcoma, fibromatosis, dermatofibrosarcoma protuberans (DFSP), malignant fibrous histiocytoma (MFH), myxoma, granular cell tumor, malignant mesenchymoma, alveolar soft part sarcoma, epitheloid sarcoma, clear cell sarcoma, and fibromatosis of small cell tumor. In a specific aspect, the soft tissue cancer is a sarcoma selected from the group consisting of fibrosarcoma, gastrointestinal sarcoma, leiomyosarcoma, dedifferentiated liposarcoma, pleomorphic liposarcoma, malignant fibrous histiocytoma, round cell sarcoma, and synovial sarcoma. The term "effective amount" means an amount that, when administered to a subject, produces a beneficial or desired result, such as a clinical result, for example, an amount that inhibits, suppresses, or reduces cancer (e.g., as determined by clinical symptoms or the amount of cancer cells) in the subject compared to a control.

[0034] As used herein, "treatment of a subject having cancer" includes partial or substantial achievement of one or more of the following: inhibition of cancer growth, reduction in the extent of cancer (e.g., reduction in tumor size), inhibition of cancer growth rate, remission or improvement of clinical symptoms or cancer-related markers (such as components of tissue or serum), or increase in the lifespan of the subject; and reduction in the likelihood of cancer recurrence.

[0035] As used herein, the term "reduction in the likelihood of cancer recurrence" means inhibition or delay of cancer recurrence at the primary site or in the vicinity thereof and / or at a second site after a remission period. It also means that, due to the treatment described herein, the cancer is less likely to recur than in the untreated case.

[0036] As used herein, the term "remission" typically refers to the disappearance or undetectability of clinical symptoms or cancer-related markers after a subject has been successfully treated by anti-cancer therapy.

[0037] Generally, the effective amount of the compounds of the present disclosure can be determined by procedures determined by those skilled in the art, although it will vary depending on the particular drug or compound, pharmaceutical formulation, route of administration, type of disease or disorder, identity of the subject or host being treated, etc. The effective amount of the compounds of the present disclosure can be readily determined by those skilled in the art by methods of established procedures known in the art.

[0038] In one embodiment, the effective amount of the 1:1 fumarate salt of compound (I) is in the range of about 0.01 to about 1000 mg / kg body weight, or about 0.05 to about 500 mg / kg body weight, or about 0.1 to about 100 mg / kg body weight, or about 0.1 to about 15 mg / kg body weight, or about 1 to about 5 mg / kg body weight, or about 2 to about 3 mg / kg body weight. Those skilled in the art will recognize that certain factors can affect the dosage required to effectively treat a subject suffering from cancer, and these factors include, but are not limited to, the severity of the disease or disorder, past treatments, the general health and / or age of the subject, and other diseases present.

[0039] Furthermore, a "treatment" regimen for a subject with an effective amount of the fumarate salt of the disclosed compound (I) may consist of a single administration or may include a series of applications. For example, the 1:1 fumarate salt of compound (I) can be administered at least once a week. However, in another embodiment, the compound can be administered to the subject approximately once a week to once a day for a particular treatment. The length of the treatment period depends on various factors such as the severity of the disease, the age of the patient, the concentration and activity of the disclosed compound, or a combination thereof. The effective dosage of the compound used for treatment or prevention may increase or decrease during the course of a particular treatment or prevention regimen. Changes in dosage can occur and can be revealed by standard diagnostic assays known in the art. In some cases, chronic administration may be required.

[0040] A "subject" is a mammal, preferably a human, but may also be an animal in need of veterinary treatment, such as a pet (e.g., dog, cat, etc.), a livestock animal (e.g., cow, sheep, pig, horse, etc.), and a laboratory animal (e.g., rat, mouse, guinea pig, etc.).

[0041] As will be understood by those skilled in the art, the compounds of the present disclosure can be administered to a patient in various forms depending on the selected route of administration. The compounds of the present disclosure can be administered, for example, by oral administration, parenteral administration, buccal administration, sublingual administration, nasal administration, rectal administration, patch administration, pump administration, or transdermal administration, and the pharmaceutical compositions are formulated accordingly. Parenteral administration includes intravenous, intraperitoneal, subcutaneous, intramuscular, trans-epithelial, nasal, intrapulmonary, intrathecal, rectal, and topical modes of administration. Parenteral administration can be by continuous infusion over a selected period of time.

[0042] Pharmaceutical Compositions and Methods of Administration The S4 crystalline form of the 1:1 fumarate salt of compound (I) disclosed herein can be suitably formulated into a pharmaceutical composition for administration to a subject. In one aspect, the present disclosure provides a pharmaceutical composition comprising the S4 form described above and a pharmaceutically acceptable carrier or diluent, wherein at least 80% by weight (preferably 90% by weight, more preferably 99% by weight) of the salt is in the S4 crystalline form.

[0043] The pharmaceutical compositions of the present teachings optionally include one or more pharmaceutically acceptable carriers and / or diluents such as lactose, starch, cellulose, and dextrose. Other excipients may also be included, such as flavoring agents; sweetening agents; and preservatives such as methylparaben, ethylparaben, propylparaben, and butylparaben. A more complete list of suitable excipients can be found in the Handbook of Pharmaceutical Excipients (5 thIt can be found in Remington's Pharmaceutical Sciences, 20th Ed., Pharmaceutical Press (2005). Those skilled in the art will be aware of methods for preparing suitable formulations for various types of administration routes. Conventional techniques and ingredients for the selection and preparation of suitable formulations are described, for example, in Remington's Pharmaceutical Sciences (2003 - 20th edition) and United States Pharmacopeia: The National Formulary (USP 24 NF19) published in 1999. Carriers, diluents, and / or excipients are "acceptable" in the sense that they are compatible with the other ingredients of the pharmaceutical composition and not harmful to its recipient.

[0044] Typically, for oral therapeutic administration, the compounds of the present teachings can be incorporated and used with excipients in the form of ingestible tablets, buccal tablets, troches, capsules, elixirs, suspensions, syrups, wafers, etc.

[0045] Typically, for parenteral administration, generally, a solution of the compounds of the present teachings can be prepared in water appropriately mixed with a surfactant such as hydroxypropylcellulose. Dispersions may be prepared in glycerol, liquid polyethylene glycol, DMSO, and mixtures thereof, with or without alcohol, and in oils. Under normal storage and use conditions, these preparations contain preservatives to prevent microbial growth.

[0046] Typically, for injectable use, sterile aqueous solutions or dispersions of the compounds described herein, and sterile powders for the immediate preparation of sterile injectable solutions or dispersions are appropriate.

[0047] For nasal administration, the compounds of the present teachings can be formulated as aerosols, drops, gels, and powders. Aerosol formulations typically contain a solution or fine suspension of the active substance in a physiologically acceptable aqueous or non-aqueous solvent and are generally presented in a sealed container, in a sterile form, in single or multiple dose amounts. The sealed container can take the form of a cartridge or refill for use with a nebulizer. Alternatively, the sealed container can be an integrated dispensing device such as a single-dose nasal inhaler or an aerosol dispenser equipped with a metering valve, which is intended to be discarded after use. When the dosage form includes an aerosol dispenser, it will contain a propellant, which can be a compressed gas such as compressed air or an organic propellant such as a fluorochlorohydrocarbon. The aerosol dosage form can also take the form of a pump sprayer.

[0048] For buccal or sublingual administration, the compounds of the present teachings can be formulated as tablets, lozenges, or troches, with carriers such as sugar, gum arabic, tragacanth, or gelatin and glycerin.

[0049] For rectal administration, the compounds described herein can be formulated in the form of suppositories containing conventional suppository bases such as cocoa butter.

[0050] The present disclosure is illustrated by the following examples, which are in no way limiting.

Examples

[0051] Experiment TIFF0007705351000004.tif199154

[0052] Analysis conditions X-ray powder diffraction (XRPD): The samples were analyzed on a Panalytical CubiX-Pro X-ray powder diffractometer or a Bruker D8 Advance XRP diffractometer.

[0053] Panalytical conditions: The sample was placed in a silicon zero return ultra - micro sample holder. The sample was irradiated with copper Kα X - rays by an X - ray tube operating at 40 kV / 30 mA. The sample was scanned in continuous mode in the range of 3 - 45°.

[0054] Bruker D8 Advance XRP instrument parameters: A high - power Cu target operating at 40 kV / 40 mA was used. The Lynxeye detector was used with a PSD opening angle of 2.1°. The sample was scanned in the range of 4 - 40° (2σ) where typical peaks occur for most of the organic crystalline compounds.

[0055] Dynamic vapor sorption measurement (DVS): Approximately 10 mg of the sample was transferred to a DVS apparatus, and then hygroscopicity measurements were carried out at 25 °C by recording the weight change with respect to atmospheric humidity using the following parameters. TIFF0007705351000005.tif88141

[0056] Purity by HPLC: The purity of the sample was determined by HPLC. The HPLC operating parameters are listed below. TIFF0007705351000006.tif98152

[0057] Example 1: Small - scale crystallization in acetone produces the D - form, but it is not reproducible on a larger scale Several small-scale crystallization experiments were carried out in acetone. Here, on a 2 g scale, compound (I) and fumaric acid were dissolved in acetone at 50 - 60 °C. In Experiments 1 and 2, after mixing the compound (I) and the acid solution, the solvent was evaporated and concentrated, and then cooled to 0 °C in 2 - 5 hours. The D form was successfully produced from both seeded and unseeded crystallization. In both experiments, a D form that was stable in solution for at least 17 hours was given from both seeded and unseeded crystallization. In Experiment 3, when the concentration of the solvent was omitted from the procedure, no solid precipitated from the solution. From this experiment, it was concluded that the concentration of the solvent was necessary to give the D form (Table 1).

[0058] Next, the crystallization procedure in acetone was scaled up to 3.5 g and the total concentration time was extended to approximately 10 hours. In this experiment, the D form was converted to the S4 form after 3 hours of concentration.

[0059] An attempt was made to concentrate the fumaric acid solution before salt formation. After dissolving fumaric acid in acetone, the acid solution was concentrated to 8 volumes to obtain a suspension of fumaric acid. Then, compound (I) was dissolved in acetone (32 volumes) and added to the acid suspension at 55 °C with 1% seed and maintained at 55 °C for 1 hour. The solution was cooled to 5 °C and maintained for 16 - 17 hours. The D form was successfully produced (Table 2). However, it was found that the D form suspension converted to the S4 form after being maintained at 50 °C for less than 2.5 hours.

[0060] Since it rapidly transfers to the S4 form at 50 - 55 °C in acetone, it was decided to determine the effect of shortening the holding time at 50 - 55 °C. Therefore, for 1 - gram and 5 - gram batches, the holding time at 50 - 55 °C was shortened to 10 minutes, and then the temperature was cooled to 0 °C over 1.5 hours and maintained at that temperature for 16 - 17 hours. The D form was obtained with good quality, but the yields decreased to 63% and 65% respectively (Table 3). Before cooling to 5 °C, the experiment of lowering the temperature to 25 °C for 24 hours was repeated. This experiment also produced the D form with good quality, but the yield loss was still approximately 12%. It was also found that further cooling to 5 °C was not helpful for further reducing the loss (Table 4).

[0061] In an attempt to further increase the crystallization yield, a poor solvent was added during the cooling stage. Methyl t - butyl ether (MTBE) was selected as the poor solvent, and two experiments were carried out with 1 - gram and 5 - gram amounts of compound (I). A solution of compound (I) in acetone (34 volumes) was combined with a fumaric acid suspension in acetone (8 volumes) and 1% seeds at 50 °C and maintained for 0.5 hour. Then MTBE was added and cooled to 0 °C over 1.5 hours. Crystallization with the 1 - gram sample provided the D form, but the 5 - gram batch gave a mixture of D and S4. Neither experiment gave a satisfactory yield (64% and 60% respectively) (Table 5).

[0062] Next, the crystallization procedure was repeated with the temperature of mixing compound (I) and fumaric acid lowered to 25 °C and 35 °C. A mixture of the D form and the S4 form was obtained (Table 6).

[0063] (Table 1) Summary of preliminary crystallization experiments in acetone TIFF0007705351000007.tif80150

[0064] (Table 2) Scale - up of evaporation crystallization in acetone TIFF0007705351000008.tif51149

[0065] (Table 3) Shortening of the retention time after a certain addition TIFF0007705351000009.tif57156

[0066] (Table 4) Optimization of the crystallization yield by retention at 25 °C TIFF0007705351000010.tif38156

[0067] (Table 5) Addition of cold MTBE during cooling TIFF0007705351000011.tif62157

[0068] (Table 6) Salt formation in acetone at 25 °C and 35 °C TIFF0007705351000012.tif34128

[0069] Example 2: The crystallization quality depends on the starting material From the results of Example 1 above, salt formation in acetone at 50 °C seemed to be the most promising approach for producing the D form in reasonable yields. In the proposed method, first, at 50 °C, a solution of compound (I) in acetone was added to a fumaric acid acetone suspension to obtain a supersaturated acetone solution. Then, seeds were added to this supersaturated solution. After seed addition (before cooling), the system was maintained for no more than 2 hours; then, after cooling the system to 25 °C, it was maintained at 25 °C overnight. After filtration, the cake was washed with methyl t-butyl ether and dried at 50 °C under vacuum. This method was able to produce the desired D form in a yield of over 60%. To confirm the proposed method, several experiments were carried out on scales of 4 grams (purity 99.1%) and 9 grams (purity 99.8%). The experiment on the 4-gram batch produced the desired D form, but the experiment on the 9-gram scale produced the S4 form. Since the scales of the two experiments were similar, it was considered that the results might depend on the quality of the starting material (Table 7).

[0070] To study the influence of the quality of the starting material, several experiments were carried out using starting materials of different purities (Table 8). In Experiment 4, a lower purity compound (I) was used in the experiment. However, no obvious crystallization was found during the study. In Experiment 5, a batch of compound (I) with high purity was mixed with several batches of lower purity materials and used as the starting material. No crystallization was observed after seeding. The holding time at 50 °C was extended to 3 hours, but a mixture of D-form and S4-form was produced. In Experiment 6, a mixture of a typical pure compound (I) and the mother liquor from the preparation of compound (I) (the mother liquor from the synthesis step of compound (I)) was used as the starting material. The D-form was produced and could be maintained for a relatively long time.

[0071] To study the effect of residual solvents in compound (I), several "spiking" experiments were carried out (Table 9). The "spiked" solvents were the solvents used in the synthesis step. Furthermore, compound (I) was amorphous in several batches. To study whether the polymorphism of compound (I) affects the polymorphism of the salt, experiments were carried out starting with amorphous compound (I). In Experiment 7, 3% MeOH was added to the system and the D-form was transformed into the S4-form during maintenance at 50 °C. In Experiment 8, 1% methyl acetate was added to the system and the D-form was transformed into the S4-form during maintenance at 50 °C. In Experiment 9, amorphous compound (I) was used as the starting material and the D-form was transformed into the S4-form during maintenance at 50 °C.

[0072] In summary, an evaporation crystallization technique for giving the D-form was developed. However, it was then found that the success of the technique highly depends on the purity and form of the starting material. Therefore, starting materials with slightly different trace impurities or purity levels gave different polymorphs. The success of crystallization was also found to depend on the fine variation of crystallization conditions and the scale of crystallization. However, the critical factors controlling the formation of the D-form or the stable S4-form were not determined.

[0073] (Table 7) Repeatability of the proposed method in acetone TIFF0007705351000013.tif52156ML: Mother liquor

[0074] (Table 8) Effect of starting materials TIFF0007705351000014.tif75152

[0075] (Table 9) Different batches of starting materials TIFF0007705351000015.tif57150

[0076] Example 3: The large-scale production process aimed at pure D form resulted in a 1:1 mixture of D form and S4 form 1.05 kg of compound (I) was dissolved in 5 - 10 kg of acetone at 55 - 60 °C with stirring for 30 minutes to prepare a compound (I) solution. A solution of 0.32 kg of fumaric acid dissolved in 35 kg of acetone was gradually added to the compound (I) solution at 60 °C. After concentrating the resulting mixture, 3.5 g of seed crystals containing D form were added. The mixture was stirred at 60 °C for 1 - 2 hours. Then, the mixture was concentrated and cooled to 50 °C in 2 hours. Next, 4 kg of MTBE was added to the 50 °C mixture, and the resulting mixture was stirred at the same temperature for an additional 1 - 2 hours. The mixture was gradually cooled to 25 °C over 20 - 25 hours. A polymorphic transition resulting in a 1:1 mixture of D polymorph and S4 polymorph was detected at the end of the cooling process. Then, the fumarate was collected, washed twice with 3 - 4 kg of MTBE, filtered, and dried at 65 - 70 °C for 8 - 24 hours. 1.02 kg of a 1:1 mixture of D polymorph and S4 polymorph was obtained as the final product.

[0077] Example 4: Small-scale preparation of 1:1 compound (I) fumarate S4 crystal form A novel small-scale crystallization method in the MEK / water / EtOH / MCH solvent system was developed as follows: 0.282 g of fumaric acid was dissolved in 5 ml of ethanol at 50 °C to prepare an acid solution. 1.0 g of compound (I) was dissolved in 7 ml of 2-butanone / water (95 / 5 volume ratio) at 50 °C to prepare a compound (I) solution. 20% of the acid solution was added to the compound (I) solution, 20 mg of seed crystals were added to the mixed solution, and then the remainder of the acid was added dropwise. The temperature was maintained at 50 °C for 3 hours, then cooled to 25 °C in 2.5 hours and further maintained at 25 °C for 17 hours. Next, 22 volume equivalents of methylcyclohexane were added over 2 hours. The temperature was cooled to 5 °C in 30 minutes and further maintained at 5 °C for 3 hours. The sample was filtered and dried under vacuum at 50 °C to obtain the final product, which was confirmed by XRPD to be in a single S4 crystal form. The XRPD pattern for this final product was essentially identical to that shown in Figure 1 and listed in Table 10 for the final product prepared by using 100 g of compound (I) based on the same method. Due to its unique XRPD pattern with sharp peaks and a flat baseline, the S4 form was considered to be a single crystal form with a crystal purity of at least 90%.

[0078] To improve the production yield, methylcyclohexane needs to be appropriately added as a poor solvent. Specifically, when the volume ratio of 2-butanone / water (95 / 5), ethanol, and methylcyclohexane is 6 - 7 to 4 - 5 to 11 - 22, the S4 form can consistently be obtained in a yield higher than 90% (Table 11). Also, mainly to improve the yield through a decrease in the solubility of the product, the molar ratio of fumaric acid to compound (I) can also be finely adjusted. When the molar ratio of fumaric acid to compound (I) was changed from 1.1 to 1.5, the loss in the yield of the S4 form decreased from 7% to 3% (Table 12). A molar ratio of 1.3 was later found to have performance equivalent to 1.5 and was thus applied to the crystallization method.

[0079] From the preparation of each of the fumaric acid solution or the compound (I) solution and maintenance at 50 °C for an extended period (up to 24 hours), rapid addition (within 30 minutes) of fumaric acid or a methylcyclohexyl poor solvent, and rapid cooling, the crystallization method was concluded to be very robust (Table 13). Even under these stress conditions, consistently, the S4 form was obtained with extremely high purity; the yield, although slightly decreased, was still much higher than that obtained for the D form from the methods disclosed in the present disclosure (see Examples 1-2) and U.S. Patent No. 9,884,855 (see Example 6).

[0080] (Table 10) XRPD of the S4 form TIFF0007705351000016.tif92128

[0081] (Table 11) Pre-crystallization in MEK / water / EtOH (MCH) TIFF0007705351000017.tif91152

[0082] (Table 12) Effect of the amount of added acid TIFF0007705351000018.tif77153

[0083] (Table 13) Stress tests TIFF0007705351000019.tif94157

[0084] Example 5: Large-scale production process for the S4 form To produce the S4 form on a large scale, for the formation of the compound (I) solution, 17.0 kg of compound (I) was dissolved in 90 kg of 2-butanone / water (95 / 5 volume ratio) at 45 - 55 °C with stirring for 1 - 4 hours. For the formation of the acid solution, 4.3 kg of fumaric acid was dissolved in 74 kg of EtOH at 45 - 55 °C with stirring for 1 - 4 hours. A portion of the acid solution was added to the compound (I) solution at 45 - 55 °C and stirred at 45 - 55 °C for 0.5 - 4 hours. Seed crystals containing the S4 form were added to the mixed solution. The seeded mixture was stirred at 45 - 55 °C for 2 - 6 hours, and then the remaining acid solution was added. The resulting mixture was stirred at 45 - 55 °C for 3 - 12 hours and then cooled to 20 - 30 °C over 5 - 12 hours. Next, 284 kg of methylcyclohexane was gradually added over 6 - 8 hours. After the addition, the mixture was stirred at 20 - 30 °C for 3 - 12 hours, then cooled to 0 - 10 °C over 3 - 12 hours and maintained at 0 - 10 °C with stirring for an additional 6 - 8 hours. The desired salt was then collected, washed twice with 16 kg of MCH, filtered, and dried at 50 °C for 8 - 24 hours to obtain 17.96 kg of the final product, which was confirmed by XRPD to be in a single S4 crystal form (see Figure 2). Due to its unique XRPD pattern with sharp peaks and a flat baseline, the S4 form was considered to be a single crystal form with a crystal purity of at least 90%.

[0085] Example 6: Hygroscopicity and Solubility Tests Based on the previously described DVS measurements, the S4 form showed low hygroscopicity at 0 - 80% RH, with a weight gain at 80% RH of only 0.41%, which was slightly lower than that of the D form, which had a weight gain of 0.56% at 80% RH.

[0086] To evaluate the dissolution behavior of the S4 form in comparison with the D form, a number of tests were carried out. The intrinsic dissolution rate at pH 2 was 429.3 μg / cm 2 -min for the S4 form and 440.4 μg / cm 2Similar results were shown for ( - min). Equilibrium solubility was also measured in various biorelevant media at atmospheric temperature. The S4 form showed a high equilibrium solubility (2.52 mg / ml) in fasting state simulated gastric fluid (FaSSGF) and a low equilibrium solubility (0.05 mg / ml) in fasting state simulated intestinal fluid (FaSSIF), which were similar to those observed for the D form (2.08 and 0.08 mg / ml, respectively).

[0087] In accordance with FDA guidance, in order to establish an in vivo - in vitro correlation (IVIVC), i.e., the correlation between the in vitro properties of a dosage form and the in vivo response, the in vitro dissolution test of the drug substance in the oral dosage form can be properly designed. According to this guidance, two batches of fumarate containing the D crystal form and the S4 crystal form were used to prepare tablets in order to evaluate the impact on the performance of the drug product. Each of the two batches of fumarate and the excipients were individually sieved and then weighed. Next, the desired amounts of each component were transferred to a shaker mixer and mixed. Then, tableting for the two batches was carried out using a single - punch tableting machine.

[0088] Next, the performance of these tablets was evaluated using a properly validated dissolution test method. The results showed that both batches of tablets had very similar dissolution profiles: for all batches, the final average concentration reached 94% after 60 minutes (see Table 14). These data indicate that the drug product prepared from the S4 crystal form shows performance equivalent to that of the D form with respect to the in vitro dissolution test, which means bioequivalence as suggested by FDA guidance.

[0089] (Table 14) In vitro dissolution tests for S4 form and D form in each oral dosage form TIFF0007705351000020.tif52148

[0090] Example 7: Pharmacokinetic analysis In the clinical development of compound (I), tablets were prepared according to Current Good Manufacturing Practice (cGMP) rules using two batches of fumarates each containing either the D polymorph or the S4 polymorph. These are referred to in the following sections as D-form tablets and S4-form tablets.

[0091] Method: Drug administration, blood collection, and plasma preparation Patients were dosed in a clinical trial approved by Health Canada and the hospital's Research Ethics Board where the drug was administered. Eight patients were dosed with S4-form tablets, six at a dose level of 48 mg and two at a dose level of 160 mg. Sixty-four patients were dosed with D-form tablets at doses in the range of 3 - 160 mg.

[0092] Blood was collected for analysis before dosing and at 2, 4, and 6 hours after dosing. Briefly, approximately 6 mL of blood was collected into a K3EDTA blood collection tube using standard venipuncture techniques and then inverted 8 - 10 times to mix. The sample was then centrifuged at approximately 5°C at 1,000 g for 10 - 15 minutes. The plasma fraction of the blood was removed using a disposable pipette, transferred to a new tube, frozen at -70°C, and then transported on dry ice for analysis.

[0093] Human plasma analysis The level of compound (I) in plasma was measured using a method validated according to all applicable USFDA, OECD, and MHLW rules, as well as the USFDA Guidance for Industry: Bioanalytical Method Validation May 2001. Briefly, compound (I) as the internal standard 13A human plasma sample containing compound (I) together with C6 and K3EDTA as an anticoagulant was prepared by liquid-liquid extraction. After evaporation of the extraction solvent, the sample extract was regenerated and analyzed by reversed-phase HPLC using Waters Atlantis dC18. The mobile phase was sprayed using heated nitrogen in a Z-spray ion source / interface set to electrospray positive ionization. The ionized compounds were detected using MS / MS. To determine the compound concentration, the experimental samples were compared with standard curve samples prepared using a reference standard material. Pharmacokinetic parameters were determined using the Excel add-in "PK Functions".

[0094] Results To compare the doses at different dose levels, the exposure (AUC 0-6h value) (ng*h / mL) was divided by the administered dose (mg). The results are plotted in Figure 3. The mean value of the exposure normalized by the dose after dosing with D-shaped tablets was 3.11 ± 2.19 ng*h / mL / mg. The mean value of the exposure normalized by the dose after dosing with S4-shaped tablets was 5.99 ± 2.48 ng*h / mL / mg. The p-value for significance calculated using a two-sided t-test was 0.0009.

[0095] Conclusion Dosing with tablets containing S4 results in a higher exposure level in patients than dosing with tablets containing D. In the presented data, the mean value of the doubling rate of exposure after dosing with S4-shaped tablets compared to D-shaped tablets is 5.99 / 3.11 = 1.9-fold.

Claims

1. A crystal of the fumarate of the compound (I) represented by the following structural formula: wherein the molar ratio of compound (I) to fumaric acid in the fumarate of compound (I) is 1:1, and the crystal is characterized by an X-ray powder diffraction pattern including peaks at 2θ of 6.6°, 9.8°, 16.3°, 21.1°, 28.7°, and 30.2° ± 0.

2.

2. The crystal according to claim 1, characterized by an X-ray powder diffraction pattern including peaks at 2θ of 6.6°, 9.8°, 16.3°, 21.1°, 28.7°, 29.4°, and 30.2° ± 0.

2.

3. The crystal according to claim 1, characterized by an X-ray powder diffraction pattern including peaks at 2θ of 6.6°, 9.8°, 13.0°, 16.3°, 19.5°, 21.1°, 22.5°, 28.7°, 29.4°, and 30.2° ± 0.

2.

4. The crystal according to claim 1, characterized by an X-ray powder diffraction pattern including peaks at 2θ of 6.6°, 9.8°, 13.0°, 16.3°, 19.5°, 21.1°, 22.5°, 22.9°, 23.9°, 28.7°, 29.4°, and 30.2° ± 0.

2.

5. The crystal according to claim 1, characterized by an X-ray powder diffraction pattern including peaks at 2θ of 6.6°, 9.8°, 11.6°, 13.0°, 16.3°, 17.1°, 19.5°, 21.1°, 21.5°, 22.0°, 22.5°, 22.9°, 23.9°, 24.3°, 28.7°, 29.4°, and 30.2° ± 0.

2.

6. The crystal according to any one of claims 1 to 5, wherein at least 90% by weight of the crystal is in a single crystal form.

7. The crystal of the compound according to any one of claims 1 to 5, wherein at least 99% by weight of the crystal is in a single crystal form.

8. The crystal according to any one of claims 1 to 5, containing less than 30% by weight of a crystal characterized by an X-ray powder diffraction pattern including peaks at 2θ of 9.6°, 12.8°, 16.0°, 16.9°, 20.7°, 20.8°, 21.2°, and 22.0° ± 0.

2.

9. The crystal according to any one of claims 1 to 5, containing less than 10% by weight of a crystal characterized by an X-ray powder diffraction pattern including peaks at 2θ of 9.6°, 12.8°, 16.0°, 16.9°, 20.7°, 20.8°, 21.2°, and 22.0° ± 0.

2.

10. A crystal according to any one of claims 1 to 9, and a pharmaceutically acceptable carrier or diluent, A pharmaceutical composition comprising the same.

11. A method for preparing the crystal according to claim 1, comprising dissolving fumaric acid and compound (I) in a dissolving solvent containing 2-butanone, water, and ethanol for forming a crystallization solution, and then precipitating crystals from the crystallization solution, A method wherein the molar ratio of compound (I) to fumaric acid in the dissolving solvent is 1:1 to 1:1.

3.

12. The method according to claim 11, comprising dissolving fumaric acid and compound (I) in a dissolving solvent at an elevated temperature, and precipitating crystals from the crystallization solution by cooling and / or adding methylcyclohexane while cooling.

13. The method according to claim 11 or 12, comprising the following steps: (i) dissolving compound (I) in a mixture of 2-butanone and water at an elevated temperature for forming a compound (I) solution; (ii) dissolving fumaric acid in ethanol at an elevated temperature for forming a fumaric acid solution; (iii) adding the fumaric acid solution to the compound (I) solution at an elevated temperature for forming a crystallization solution; (iv) optionally, adding seed crystals to the crystallization solution at an elevated temperature; and (v) lowering the temperature of the crystallization solution to precipitate crystals.

14. The method according to claim 13, wherein the volume ratio of 2-butanone to water is 80:20 to 98:2, the elevated temperature is 45°C to 55°C; and the lowered temperature is 20°C to 30°C.

15. The method according to claim 13, wherein the volume ratio of 2-butanone to water is 90:10 to 98:2, the elevated temperature is 45°C to 55°C; and the lowered temperature is 20°C to 30°C.

16. The method according to claim 13, further comprising adding methylcyclohexane to the crystallization solution at a lowered temperature to precipitate crystals.

17. The method according to claim 16, wherein the volume ratio of 2-butanone to water is 80:20 to 98:2, the elevated temperature is 45°C to 55°C; and the volume ratio of 2-butanone / water, ethanol, and methylcyclohexane is 6 to 7 to 4 to 5 to 11 to 22.

18. The method according to claim 16, further lowering the temperature of the crystallization solution to precipitate crystals.

19. The method according to claim 16, wherein the lowered temperature is 20°C to 30°C.

20. The method according to claim 18, wherein the further lowered temperature is 0°C to 10°C.

21. A pharmaceutical composition for use in treating a subject having cancer, comprising an effective amount of the crystal according to any one of claims 1 to 9.

22. The pharmaceutical composition according to claim 21, wherein the cancer is selected from the group consisting of lung cancer, breast cancer, colon cancer, neuroblastoma, prostate cancer, melanoma, glioblastoma multiforme, ovarian cancer, lymphoma, leukemia, osteosarcoma, germ cell tumor, glioma, fibrosarcoma, gastrointestinal sarcoma, fibrous histiocytoma, round cell sarcoma, synovial sarcoma, cervical cancer, anogenital cancer, head and neck cancer, and hypopharyngeal cancer.

23. The pharmaceutical composition according to claim 21, wherein the cancer is selected from the group consisting of lung cancer, breast cancer, and colon cancer.

24. The pharmaceutical composition according to claim 21, wherein the cancer is breast cancer.

25. The pharmaceutical composition according to claim 21, wherein the cancer is triple-negative breast cancer.

26. The pharmaceutical composition according to claim 25, wherein the triple-negative breast cancer is inoperable or metastatic.

27. A pharmaceutical composition for use in treating a patient having pancreatic cancer, comprising an effective amount of the crystal according to any one of claims 1 to 9.

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