Crystalline salt forms of EP4 antagonists

Abstract

Providing a crystalline salt form of an EP4 antagonist. A crystalline free acid form of (S)-4-(1-(3-(difluoromethyl)-1-methyl-5-(3-(trifluoromethyl)phenoxy)-1H-pyrazole-4-carboxamido)ethyl)benzoic acid is provided. Methods of making and using the crystalline free acid form of (S)-4-(1-(3-(difluoromethyl)-1-methyl-5-(3-(trifluoromethyl)phenoxy)-1H-pyrazole-4-carboxamido)ethyl)benzoic acid are also provided. JPEG2024545695000025.jpg46149

Description

[Technical field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of U.S. Provisional Patent Application No. 63 / 293 / 183, filed December 23, 2021, which is incorporated by reference as if fully set forth herein. [Background technology]

[0002] Blockade of prostaglandin E2 (PGE2) signaling by antagonists through the interaction of PGE2 with prostaglandin E receptor 4 (EP4) has been shown to be effective in reducing inflammation (Chen et al. (2010) British J. Pharmacol. 160, 292-310). PGE2 has also been implicated as a key component in the immunosuppressive environment created by many solid tumors (Whiteside (2010) Expert Opinion in Biological Therapy. 2010.10, 1019-1035), and inhibition of EP4 signaling by antagonists has been shown to reduce tumor growth (Terada et al. (2010) Cancer Res. 70, 1606-1615) and tumor metastasis in tumor animal models (Yang et al. (2006) Cancer Res. 66, 9665-9672). Further reports of PGE2-type EP4 antagonism used in oncology can be found in Ma X. et al. (2013) Oncoimmunology 2:e22647 and Cherukuri, DP et al. (2007) Exp. Cell Res. 313, 2969-2979; Majumder, M. et al. (2018) Int. J. Mol. Sci. 19, 1019; and Terada, N. et al. (2010) Cancer Research 70, 1606-1615.

[0003] Specific examples of EP4 antagonists include compounds having a chemical structure according to Formula I, and are described in US Pat. No. 8,686,018, which is incorporated herein by reference in its entirety. [ka]

[0004] Although the usefulness of EP4 antagonism has been demonstrated in the art, their preparation and use as active pharmaceutical ingredients in pharmaceuticals must be in a form that can be conveniently operated and processed.In this regard, the chemical stability and physical stability of active compounds are important considerations.Preferably, compounds and pharmaceutical compositions containing them can be effectively stored for long periods of time without showing significant changes in physiochemical properties. Summary of the Invention [Means for solving the problem]

[0005] An embodiment of the present invention may provide a crystalline free acid form of a compound according to Formula II (hereinafter Compound 1). [ka] (S)-4-(1-(3-(difluoromethyl)-1-methyl-5-(3-(trifluoromethyl)phenoxy)-1H-pyrazole-4-carboxamido)ethyl)benzoic acid

[0006] Please note that in the event of a discrepancy between the depicted IUPAC name of Compound 1, or any other compound reported herein, and its fully drawn chemical structure, the structure is controlling.

[0007] In some embodiments, the crystalline free acid form of the compound of Formula II provides at least one, two, three, four, five, six or more peaks in an X-ray powder diffraction (XRPD) spectrum that fall within the following ranges of values ​​in degrees 2θ (±0.2°): 8.2, 10.3, 12.6, 14.2, 14.5, 14.7, 14.8, 15.3, 15.8, 16.4, 17.8, 18.3, 18.4, 19.2, 19.5, 20.4, 23.0, 23.4, 24.2, 25.4, 25.8, 26.3, 26.5, 27.7, 28.9, 29.4, 30.0, 30.4, 31.2, 32.0, 32.6, 33.3, 34.0 and 34.2. In some embodiments, the crystalline free acid form of Compound 1 is characterized by an XRPD pattern substantially as depicted in Figure 1. In some embodiments, the crystalline free acid form of the compound provides peaks in a powder X-ray diffraction spectrum in the ranges of 2θ° (±0.2°): 8.2, 10.3, 12.6, 14.2, 14.5, 14.7, 14.8, 15.3, 15.8, 16.4, 17.8, 18.3, 18.4, 19.2, 19.5, 20.4, 23.0, 23.4, 24.2, 25.4, 25.8, 26.3, 26.5, 27.7, 28.9, 29.4, 30.0, 30.4, 31.2, 32.0, 32.6, 33.3, 34.0, and 34.2. In other embodiments, the crystalline free acid form of the compound provides peaks in its powder X-ray diffraction spectrum in the ranges of 2θ° (±0.2°): 14.7, 14.8, 15.3, 16.4, 18.3, 18.4, 19.5, 20.4, 23.0, 24.2, 27.7, 33.3, and 34.2. In further embodiments, the crystalline free acid form of the compound provides peaks in its powder X-ray diffraction spectrum in the ranges of 2θ° (±0.2°): 14.7, 14.8, 18.4, 19.5, 20.4, 23.0, 24.2, and 33.3. In some embodiments, the crystalline free acid form of the compound provides peaks in its powder X-ray diffraction spectrum in the ranges of 2θ° (±0.2°): 18.4, 20.4, and 24.2.

[0008] The 2θ° values ​​of the crystalline free acid form of the compound according to formula II were determined by using a 50 kV, 300 mA X-ray diffractometer, model: RINT TTR-III (Rigaku), using CuKα radiation at 50 kV and 300 mA run in continuous scan mode and parallel beam measurement mode at a scan rate of 5.00° / min. The detector was a scintillation counter and the tube voltage was 50 kV with a tube current of 300 mA. The step width was 0.02° and an entrance slit box of 0.50 mm was used. The diffractometer was also operated at room temperature with a scan range of 5 to 35°.

[0009] In some embodiments, the crystalline free acid form is characterized by a differential scanning calorimetry (DSC) thermogram substantially similar to that shown in Figure 2. In other embodiments, the crystalline free acid form of Compound 1 is characterized by an endothermic peak at 150°C.

[0010] Embodiments may provide pharmaceutical compositions comprising a crystalline form of Compound 1 described herein and a pharma- ceutically acceptable carrier. In some embodiments, the compositions are formulated for oral or parenteral administration.

[0011] Embodiments may provide a method of making the crystalline free acid form of Compound 1. Such a method may include the following steps: a) Preparing a pyrazole according to Reaction Scheme A: [ka] ; b) oxidizing the pyrazole according to Reaction Scheme B: [ka] ; c) Functionalization of the pyrazole oxide according to Reaction Scheme C: [ka] ; d) coupling the functionalized pyrazole to an amine to form an amide according to Reaction Scheme D: [ka] ; e) Preparing the crystalline free acid form of Compound 1 according to Reaction Scheme E: [ka] may include one or more of:

[0012] Embodiments may provide a method of treating cancer in a subject in need thereof, comprising administering a therapeutically effective amount of a crystalline free acid form of Compound 1 to the subject.

[0013] An embodiment may provide a method of treating cancer in a subject in need thereof, comprising detecting an altered EP4 status (e.g., increased expression of EP4) in a biological sample of cells, and if the biological sample contains cancerous cells having an altered EP4 status, administering a therapeutically effective amount of the crystalline free acid form of Compound 1 to the subject.

[0014] An embodiment may provide for the use of the crystalline free acid form of Compound 1 in a method for the treatment of cancer.

[0015] An embodiment may provide for the use of a crystalline free acid form of Compound 1 in the preparation of a medicament for treating cancer. [Brief description of the drawings]

[0016] [Figure 1] FIG. 1 shows the XRPD spectrum obtained from the crystalline free acid form of Compound 1. [Diagram 2] FIG. 2 shows the DSC thermogram of the crystalline free acid form of Compound 1. [Diagram 3] FIG. 3 shows the hygroscopicity of the crystalline free acid form of Compound 1. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0017] Compound 1: [ka] Provided herein are crystalline forms of which are useful as selective EP4 antagonists.

[0018] In some embodiments, the crystalline form of Compound 1 is a free acid form. One embodiment of the crystalline free acid form of Compound 1 exhibits peaks in an X-ray powder diffraction (XRPD) spectrum in one, two, three, four, five, six or more of the following ranges of 2θ° (±0.2°) values: 8.2, 10.3, 12.6, 14.2, 14.5, 14.7, 14.8, 15.3, 15.8, 16.4, 17.8, 18.3, 18.4, 19.2, 19.5, 20.4, 23.0, 23.4, 24.2, 25.4, 25.8, 26.3, 26.5, 27.7, 28.9, 29.4, 30.0, 30.4, 31.2, 32.0, 32.6, 33.3, 34.0, and 34.2. For example, the crystalline free acid form of Compound 1 may exhibit at least one, two, three, four, five, or six values ​​selected from the group consisting of 2θ° (±0.2°): 14.7, 14.8, 15.3, 16.4, 18.3, 18.4, 19.5, 20.4, 23.0, 24.2, 27.7, 33.3, and 34.2. In further embodiments, the crystalline free acid form of the compound exhibits peaks in its powder X-ray diffraction spectrum in the ranges of 2θ° (±0.2°): 14.7, 14.8, 18.4, 19.5, 20.4, 23.0, 24.2, and 33.3. In some embodiments, the crystalline free acid form of the compound exhibits peaks in its powder X-ray diffraction spectrum in the ranges of 2θ° (±0.2°): 18.4, 20.4, and 24.2. In some embodiments, the crystalline free acid form of Compound 1 is characterized by an XRPD pattern substantially as shown in FIG.

[0019] In some embodiments, the crystalline free acid form is characterized by a differential scanning calorimetry (DSC) thermogram substantially similar to that shown in FIG.

[0020] In some embodiments, the crystalline free acid form is characterized by hygroscopicity substantially similar to that shown in FIG.

[0021] As used herein, "substantially as shown," "substantially the same," or "substantially as indicated," with respect to data such as spectra, means that one of skill in the art, when comparing such spectra obtained using the same data collection methods, e.g., as shown in FIG. 1, would conclude that the spectra are sufficiently similar to represent the same crystalline free acid form of the compounds taught herein.

[0022] Also provided are methods of synthesizing and crystallizing the compounds taught herein. In the case of the crystalline free acid form, the methods comprise the following steps: a) Preparing a pyrazole according to Reaction Scheme A: [ka] ; b) oxidizing the pyrazole according to Reaction Scheme B; [ka] ; c) Functionalization of the pyrazole oxide according to Reaction Scheme C: [ka] ; d) coupling the functionalized pyrazole to an amine to form an amide according to Reaction Scheme D: [ka] ; e) Producing the crystalline free acid form of Compound 1 according to Reaction Scheme E: [ka] may include one or more of the following:

[0023] The crystalline compounds described herein can be combined with a pharma- ceutically acceptable carrier to provide a pharmaceutical formulation thereof. The particular choice of carrier and formulation depends on the particular route of administration for which the composition is intended. In some embodiments, the carrier is selected to maintain the crystalline form of the compound prior to administration.

[0024] As used in the specification and claims, the singular forms "a," "an," and "the" include plural references unless the content dictates otherwise. Thus, for example, reference to the "HCl salt" refers to the monohydrochloride, dihydrochloride, 1.5hydrochloride, as well as other stoichiometric and non-stoichiometric hydrochloride salts.

[0025] As used herein, "pharmaceutical acceptable carrier" refers to a non-toxic carrier, adjuvant, or vehicle that does not destroy the pharmacological activity of the compound in which it is formulated. In some embodiments, the pharmaceutical acceptable carrier is selected to maintain the crystalline free acid form of the compound. Pharmaceutically acceptable carriers, adjuvants, or vehicles include, but are not limited to, sorbic acid, potassium sorbate, partial glyceride mixtures of saturated vegetable fatty acids, water, salts or electrolytes, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts, colloidal silica, magnesium trisilicate, polyvinylpyrrolidone, cellulose-based materials, polyethylene glycol, sodium carboxymethylcellulose, polyacrylates, waxes, polyethylene glycol, and wool fat.

[0026] The compositions of the present invention may be suitable for oral, parenteral, topical or implanted reservoir administration. In some embodiments, the formulation comprises components derived from natural or non-natural sources. In some embodiments, the formulation or carrier may be provided in a sterile form. Non-limiting examples of sterile carriers include endotoxin-free water or pyrogen-free water.

[0027] The term "parenteral" as used herein includes subcutaneous, intravenous, intramuscular, intra-articular, intrasynovial, intrasternal, intrathecal, intrahepatic, intralesional and intracranial injection or infusion techniques. In certain embodiments, the compound is administered by intravenous, oral, subcutaneous or intramuscular administration. Sterile injectable forms of the compositions of the present invention may be aqueous or oily suspensions. These suspensions may be formulated according to techniques known in the art using suitable dispersing or wetting agents and suspending agents. Sterile injectable preparations may also be sterile injectable solutions or suspensions in non-toxic parenterally acceptable diluents or solvents. Among the acceptable vehicles and solvents that may be used are water, Ringer's solution and isotonic sodium chloride solution. In addition, sterile fixed oils are conventionally used as solvents or suspending media.

[0028] For this purpose, any non-irritating fixed oil can be used, including synthetic mono- or diglycerides. Fatty acids and their glyceride derivatives, especially in their polyoxyethylated versions, are useful in the preparation of injectables, as are natural pharmaceutically acceptable oils, such as olive oil or castor oil. These oil solutions or suspensions may also contain long-chain alcohol diluents or dispersants, such as carboxymethylcellulose or similar dispersants commonly used in the preparation of pharmaceutically acceptable dosage forms, including emulsions and suspensions. Other commonly used surfactants, such as Tween, Spans and other emulsifiers, commonly used in the manufacture of pharmaceutically acceptable solid, liquid, or other dosage forms, may also be used for formulation purposes.

[0029] For oral administration, the compound or salt may be provided in any acceptable oral dosage form, including, but not limited to, capsules, tablets, aqueous suspensions or solutions. For tablets for oral use, commonly used carriers include lactose and cornstarch. Lubricants such as magnesium stearate may also be added. For oral administration in capsule form, useful diluents include lactose and dried cornstarch. When aqueous suspensions are required for oral use, the active ingredient may be combined with emulsifying and suspending agents. If desired, certain sweetening, flavoring or coloring agents may be added. In addition, preservatives may be added. Suitable examples of pharmaceutically acceptable preservatives include, but are not limited to, various antibacterial and antifungal agents, such as solvents, such as ethanol, propylene glycol, benzyl alcohol, chlorobutanol, quaternary ammonium salts and parabens (e.g., methylparaben, ethylparaben, propylparaben, etc.).

[0030] In some embodiments, the crystalline free acid form of Compound 1 is formulated into a capsule, and 5 mg to 50 mg, 10 mg to 40 mg, or 20 mg to 30 mg of Compound 1 is present in the capsule.

[0031] In other embodiments, the capsule comprises a capsule shell made from hypromellose.

[0032] In some embodiments, the crystalline free acid form of Compound 1 is formulated into a tablet, and Compound 1 undergoes a wet granulation process during tablet formation.

[0033] In other embodiments, the tablet comprises lactose monohydrate, low substituted hydroxypropyl cellulose, hydroxypropyl cellulose, microcrystalline cellulose, magnesium stearate, and / or water.

[0034] The tablets may have a size of 1 mm to 7 mm, 2 mm to 7 mm, 4 mm to 6.5 mm, or 5.5 mm to 6.5 mm. In some embodiments, the tablets have a size of 6.5 mm.

[0035] Target and usage The crystalline forms of the compounds taught herein may be used to treat cancers such as skin cancer, breast cancer, colorectal cancer, prostate cancer, kidney cancer, cervical cancer, ovarian cancer, endometrial cancer, glioblastoma, head and neck cancer, medulloblastoma, lung cancer, or urinary tract cancer.

[0036] "Treate", "treating" and "treatment" refer to reversing, alleviating, delaying the onset of, inhibiting the progression of, or otherwise ameliorating a disease or disorder as described herein. In some embodiments, treatment may be administered after one or more symptoms have developed. In other embodiments, treatment may be administered in the absence of symptoms. For example, treatment may be administered to a susceptible individual prior to the onset of symptoms (e.g., in light of a medical history and / or in light of genetic or other susceptibility factors). Treatment may also be continued after symptoms have resolved, e.g., to prevent or delay recurrence.

[0037] As used herein, "patient" or "subject" refers to an animal subject, preferably a mammalian subject, particularly a human subject (including both male and female subjects, including neonatal, infant, juvenile, adolescent, adult and geriatric subjects). Subjects also include other mammalian subjects (e.g., dogs, cats, horses, cows, sheep, goats, monkeys, birds, etc.) for experimental or veterinary purposes.

[0038] In some embodiments, treatment is provided to a subject having a cancer with altered EP4 status.

[0039] In some embodiments, treatment may include or may be performed in conjunction with analyzing (e.g., measuring or evaluating) EP4 status in a biological sample containing cells of the cancer, and if the cancer exhibits EP4 alterations, treating the subject with a therapeutically effective amount of an active agent described herein.

[0040] As used herein, an "altered state" with respect to EP4 includes increased activity of the encoded protein, such as as a result of its increased expression (e.g., increased mRNA levels or increased protein levels), increased copy number in the genome, and / or mutation, as compared to corresponding noncancerous tissue. In some embodiments, an altered state of EP4 includes gene and / or encoded protein mutations that result in increased activity or are otherwise associated with more aggressive forms of hepatocellular carcinoma.

[0041] "Expression" of EP4 means that the gene encoding it is transcribed and preferably translated. Typically, expression of the coding region results in production of the encoded polypeptide.

[0042] EP4 protein is known and its altered state and / or expression may be measured using techniques standard in the art, such as genomic analysis for mutations or copy number abnormalities, such as by nucleic acid amplification, sequencing analysis and / or hybridization-based techniques, RNA expression analysis such as Northern blots or qRT-PCR, Western blots or other immunoblots or immunoassays, fluorescence activated cell sorting (FACS), and the like.

[0043] In order that the invention described herein may be more fully understood, the following examples are set forth. It should be understood that these examples are for illustrative purposes only and are not to be construed as limiting. EXAMPLES

[0044] Definitions: The following abbreviations have the indicated meanings. ACN Acetonitrile DCM Dichloromethane DMF Dimethylformamide K2CO3 Potassium Carbonate MeOH Methanol EtOAc Ethyl acetate Preparative HPLC Preparative High Performance Liquid Chromatography TEA Triethylamine THF Tetrahydrofuran MTBE Methyl tert-butyl ether AcOH Acetic acid POCl3 Phosphoryl chloride t-BuOH tert-butanol IPA Isopropanol V volume equivalent

[0045] Example 1: Procedure for the synthesis of the free acid crystalline form of Compound 1: [ka] Example 1A: General synthetic scheme I: [ka] Compounds 2-4 can be prepared as described in US Pat. No. 8,686,018, which is incorporated herein by reference in its entirety.

[0046] Step 1.1 (Stage 4A) Compound 3 (1.0 kg) and 5 L of DMF were charged into the reactor at 20-30° C. and then the reaction mass was cooled to −5° C.-10° C. Compound 4 (0.56 kg) and HBTU (1.36 kg) were added sequentially into the reactor at −5-10° C. Triethylamine (TEA) (0.6019 kg) was added slowly at −5-10° C. The reaction mass was stirred at 0-10° C. for 1-3 hours. The progress of the reaction was monitored by HPLC.

[0047] Sampling procedure: Take 2 ml of the reaction mass and subject it to HPLC.

[0048] Pass criteria: Compound 3 is (NMT) 1.0% or less.

[0049] If the reaction is not complete: Stir the reaction mass further at 0-10 °C for 1-3 h and check the content of compound 3 according to the acceptance criteria.

[0050] On completion, 4 L of water was slowly charged into the reaction mass at 0-20°C and the reaction mass was extracted with 10 L of ethyl acetate and again with 6 L of ethyl acetate (Note: If separation of layers does not occur during the first extraction, increase the temperature to 10-20°C, add 2 L of ethyl acetate, stir for 10-30 minutes, allow the ethyl acetate layer to settle and separate). The combined ethyl acetate layer was washed sequentially with 8% NaHCO3 solution (5 L of water + 0.4 Kg of NaHCO3) and 10% sodium chloride solution (5 L of water + 0.5 Kg of sodium chloride) at 20-30°C. The ethyl acetate layer was distilled under vacuum below 55°C to obtain compound 2 (yield = 1.48 kg w / w wrt compound 3, assuming 100% yield) which was used in step 2.1 without isolation.

[0051] Chirality Control: The chirality of compound 2 is commercially available from (S)-methyl 4-(1-aminoethyl)benzoate (compound 4) starting material.

[0052] Step 2.1 (Stage 4B) THF (2.96 L) was charged to the reactor with compound 2 (crude, 1.48 kg) at a temperature below 55° C. THF was distilled off from the reactor under vacuum and the reactor was recharged with THF (2.96 L) at a temperature below 55° C. THF was again distilled off from the reactor under vacuum and the reactor was recharged with THF (7.4 L). The reaction mass was cooled to 10-30° C. NaOH (0.3 kg in 7.4 L water) was charged to the reaction mass at 10-30° C. The temperature was increased to 35-50° C. and the reaction mass was stirred for 8-14 hours. The progress of the reaction was monitored by HPLC.

[0053] Sampling Procedure: Take 2 ml of the reaction mass for HPLC.

[0054] Pass criteria: Compound 2 is 1.0% or less.

[0055] If the reaction is not complete: Stir the reaction mass further at 35-50°C for 1-3 hours.

[0056] On completion, the reaction mass was cooled to a temperature of 0-10°C. The pH of the reaction mass was adjusted to 1.5-2.5 by slowly adding 10 L of HCl solution (1.48 L concentrated HCl in 13.32 L water) to the reaction mass at 0-30°C. The reaction mass was extracted with ethyl acetate (11.84 L) and again with ethyl acetate (10.36 L). The combined ethyl acetate layer was washed with 10% sodium chloride solution (7.4 L water + 0.74 Kg sodium chloride). The organic layer was transferred to another reactor by particle free filtration. The ethyl acetate was distilled off completely under vacuum at a temperature below 55°C. IPA (1.48 L) was added to the residue at a temperature below 55°C. IPA was distilled off from the reaction mass under vacuum. IPA (7.4 L) was again charged into the reaction mass and the reaction mass was heated to a temperature of 55-65°C to get a clear solution. The reaction mass was stirred at 55-65°C for 10-30 minutes and then cooled to 40-50°C. Water (10.36 L) was slowly added to the reaction mass at 40-50°C and the reaction mass was stirred for 30-60 minutes at 40-50°C. The reaction mass was cooled to 20-30°C and stirred for 60±15 minutes at 20-30°C. The reaction mass was further cooled to 5-15°C and stirred at 5-15°C for 2-4 hours. The precipitated solid was then filtered and the solid was washed sequentially with 2.96 L of (1:2 V / V) IPA+water mixture, then 1.48 L of water, followed by 1.48 L of (1:2 v / v IPA+water mixture. The wet material was dried at 55-65° C. for 10-20 hours. The progress of drying of the material was monitored by the moisture content using Karl Fischer titration. The pass criterion for moisture content is 1.0% or less. If the moisture content does not meet the pass criterion, continue drying at 55-65° C. for 2-4 hours and check the moisture content. After meeting the target drying criterion, the material is removed from the dryer to obtain crude title compound 1 (0.9-1.3 kg, yield=70-90%) (Step 4B).

[0057] Step 3.1 (Stage 4C) Methanol (5 L) was charged into the reactor at 20-30°C. Product from crude compound 1 step 4B (1.0 kg) was added into the reactor. The reaction mass was heated to 55-65°C and stirred for 30-45 minutes to get a clear solution. Then the reaction mass was cooled to 30-50°C and transferred to another reactor by particle free filtration (5 micron filter). The reaction mass was rinsed with methanol (2 L) at 30-50°C and transferred to another reactor by particle free filtration (5 micron filter). The reaction mass was cooled to 5-15°C and water (13 L) was added slowly to the reaction mass at 5-25°C. After the addition of water was completed, the reaction mass was heated to a temperature of 20-30°C and stirred at the same temperature for 2-4 hours. The precipitated solid was filtered and the solid was washed with 3 L of (1:2 V / V) MeOH+water mixture. The wet material was dried at 55-65°C for 10-20 hours. The progress of drying of the material was monitored by (i) water content using Karl Fischer titration and (ii) methanol content by GC. The pass criteria for water content is 1.0% or less and the pass criteria for methanol content is 3000 ppm. If the water and methanol content do not meet the pass criteria, continue drying at 55-65°C for 2-4 hours and check the water and methanol content. After meeting the target criteria for drying, the material was removed from the dryer to obtain the crystalline free acid form of compound 1 as a white solid (0.9-0.98 kg, yield = 90-98%).

[0058] Example 1B: [ka] Alternatively, compound 1 can be prepared from the following synthesis: Compound 2 can be prepared as described in U.S. Patent No. 8,686,018, the entirety of which is incorporated herein by reference.

[0059] The reactor was charged with compound 2 (1 equiv.) followed by THF (5V). The reaction was stirred and 1M NaOH (5V) was added at 20-25°C. The reaction was heated to 60-65°C and monitored for completion (approximately 4-6 hours). Upon completion, the reaction was cooled to 0-10°C and slowly charged with 1M HCl (6V). The reaction was then charged with EtOAc (8V), stirred, allowed to phase separate, and the organic phase was then removed from the aqueous phase. The aqueous phase was back extracted with EtOAc (8V). The organic phases were then combined and washed with 12% brine (5V). The aqueous phase was removed and the organic phase was concentrated. The concentrated organic phase was then azeotroped twice with MeOH (1V). The crude organic material was then charged with MeOH (2V) and heated to 50-60°C until the solution was clear. Water (6V) was then slowly added to the solution. The solution was cooled at a rate of 0.1 °C / min to a temperature of 15-20 °C. The precipitated solid was then filtered from the solution and washed twice with 1 V MeOH:water (1:3 v / v). The collected solid was dried until a constant weight was achieved, giving overall compound 1 (0.85 W, 87%) as a white solid.

[0060] Example 2: XRPD characterization of the crystalline free acid form of Compound 1: The X-ray powder diffraction (XRPD) pattern for the crystalline free acid form of Compound 1 is shown in FIG.

[0061] Example 3: Characterization of the crystalline free acid form of TG-DTA Compound 1: Thermogravimetry and differential thermal analysis (TG-DTA) of the crystalline free acid form of Compound 1 are shown in Figure 2. In the DTA curve, an endothermic peak was observed near 150°C (onset). In the TG curve, no measurable weight loss was observed up to 180°C. The melting range of the crystalline free acid form of Compound 1 was observed at 152°C-154°C according to USP-NF 37.

[0062] Example 4: Characterization of the hygroscopicity of the crystalline free acid form of Compound 1: The dynamic vapor sorption (DVS) method was used to measure the hygroscopicity of the crystalline free acid form of Compound 1. The adsorption and desorption isotherms for the crystalline free acid form of Compound 1 are shown in Figure 3. No significant weight change was observed for the crystalline free acid form of Compound 1 over the range of 25 °C and relative humidity from 5% to 95% RH. The results showed that the crystalline free acid form of Compound 1 is non-hygroscopic.

[0063] Summary data for the crystalline free acid form of Compound 1: The physical and chemical properties of the crystalline free acid of Compound 1 are summarized in Table 1.

[0064] [Table 1]

Claims

1. Compound 1: 【Chemistry 1】 Crystalline form of.

2. 2. The crystalline compound of claim 1 which is the crystalline free acid form of Compound 1.

3. 3. The crystalline free acid compound of claim 2, wherein the crystalline free acid form compound exhibits at least the following powder X-ray diffraction peaks at 2θ° (±0.2°): 18.4, 20.4, and 24.

2.

4. 4. The crystalline free acid compound of claim 3, wherein the crystalline free acid form compound exhibits at least the following powder X-ray diffraction peaks 2θ° (±0.2°): 14.7, 14.8, 18.4, 19.5, 20.4, 23.0, 24.2, and 33.

3.

5. 5. The crystalline free acid compound of claim 4, wherein the crystalline free acid form compound exhibits at least the following powder X-ray diffraction peaks 2θ° (±0.2°): 14.7, 14.8, 15.3, 16.4, 18.3, 18.4, 19.5, 20.4, 23.0, 24.2, 27.7, 33.3, and 34.

2.

6. 6. The crystalline free acid compound of claim 5, wherein the crystalline free acid form compound exhibits at least the following powder X-ray diffraction peaks 2θ° (±0.2°): 8.2, 10.3, 12.6, 14.2, 14.5, 14.7, 14.8, 15.3, 15.8, 16.4, 17.8, 18.3, 18.4, 19.2, 19.5, 20.4, 23.0, 23.4, 24.2, 25.4, 25.8, 26.3, 26.5, 27.7, 28.9, 29.4, 30.0, 30.4, 31.2, 32.0, 32.6, 33.3, 34.0, and 34.

2.

7. 3. The crystalline free acid compound of claim 2, wherein the crystalline free acid form is characterized by an X-ray powder diffraction (XRPD) pattern substantially as shown in FIG.

8. 3. The crystalline free acid compound of claim 2, wherein the crystalline free acid form is characterized by a differential scanning calorimetry (DSC) thermograph substantially similar to that shown in FIG.

9. 3. The crystalline free acid compound of claim 2, wherein the crystalline free acid form is characterized by a hygroscopicity substantially similar to that shown in FIG.

10. A pharmaceutical composition comprising the crystalline compound of any one of claims 1 to 9 and a pharma- ceutically acceptable carrier.

11. 11. The pharmaceutical composition of claim 10, formulated for oral administration.

12. 10. A method for preparing the crystalline free acid form compound of claim 2, comprising the steps of: a) producing a pyrazole according to Reaction Scheme A: 【Chemistry 2】 ； b) oxidizing said pyrazole according to Reaction Scheme B: 【Chemistry 3】 ； c) functionalizing said pyrazole oxide according to Reaction Scheme C: 【Chemistry 4】 ； d) coupling said functionalized pyrazole to an amine to form an amide according to Reaction Scheme D: 【Chemistry 5】 ； e) producing said crystalline free acid form of Compound 1 according to Reaction Scheme E: 【Chemistry 6】 The method includes one or more of the following:

13. 12. A method of treating cancer in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a compound according to any one of claims 1 to 9 or a pharmaceutical composition according to claim 10 or claim 11.

14. 14. The method of claim 13, wherein the cancer is skin cancer, breast cancer, colorectal cancer, prostate cancer, renal cancer, cervical cancer, ovarian cancer, endometrial cancer, glioblastoma, head and neck cancer, medulloblastoma, lung cancer, or urinary tract cancer.

15. 14. The method of claim 13, wherein the cancer has an altered EP4 status.

16. The method of claim 15, wherein the altered EP4 status comprises increased expression of EP4.

17. 1. A method of treating cancer in a subject in need thereof, comprising: detecting an altered EP4 status in a biological sample containing cells of said cancer, and if said cancer has said altered EP4 status, administering to said subject a therapeutically effective amount of a crystalline form of the compound according to any one of claims 1 to 9 or a pharmaceutical composition according to claim 10 or claim 11. A method comprising:

18. The method of claim 17, wherein the altered EP4 status comprises increased expression of EP4.

19. Use of a crystalline form of a compound according to any one of claims 1 to 9 or a pharmaceutical composition according to claim 10 or claim 11 in a method for the treatment of cancer.

20. 20. The use of claim 19, wherein the cancer has an altered EP4 status.

21. The use of claim 20, wherein the altered EP4 status comprises increased expression of EP4.

22. Use of a crystalline form of a compound according to any one of claims 1 to 9 in the preparation of a medicament.

23. 23. The use according to claim 22, wherein the medicament is for the treatment of cancer.

24. 24. The use of claim 23, wherein the cancer has an altered EP4 status.

25. 25. The use of claim 24, wherein the altered EP4 status comprises increased expression of EP4.

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