2,2,2-trifluoroacetic acid 1-(2,4-dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide polymorph and method for producing the same

Abstract

Crystalline polymorphic forms of the neurotrophic agent 2,2,2-trifluoroacetic acid 1-(2,4-dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147) and methods for producing the crystalline polymorphic forms are provided.

Description

Technical Field

[0001] The present disclosure relates to polymorphic forms of active agents. Specifically, the present disclosure relates to the polymorphic form of the neuroprotective agent 1-(2,4-dimethylphenyl)-2-[(3-methoxyphenyl)methylene] hydrazide 2,2,2-trifluoroacetate (J147).

Background Art

[0002] 1-(2,4-Dimethylphenyl)-2-[(3-methoxyphenyl)methylene] hydrazide 2,2,2-trifluoroacetate (J147) is a potent orally active neurotrophic agent found in the efficacy screening in a cellular model of age-related pathology, and its structure is shown in Formula I.

[0003]

Chemical Formula

[0004] J147 has a broad neuroprotective effect and showed an activity with an EC 50 of 10 - 200 nM in assays showing different neurotoxic pathways related to aging and neurodegenerative diseases. It has been shown to improve memory in normal rodents and prevent the loss of synaptic proteins and the decline of cognitive function in transgenic AD mouse models. Furthermore, neuroprotective, neuroanti-inflammatory and LTP-enhancing activities have been shown.

[0005] The neurotrophic effect and nootropic effect are associated with an increase in BDNF levels and BDNF-responsive proteins. Interestingly, despite this mechanism of action, the neuroprotective effect of J147 has been observed to be independent of the activation of the TrkB receptor. J147 has been shown to reduce the levels of soluble Aβ40 and Aβ42 and is currently being studied for its potential use in the treatment of ALS.

Summary of the Invention

Problems to be Solved by the Invention

[0006] This disclosure relates to crystalline form II of 2,2,2-trifluoroacetic acid 1-(2,4-dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147) and a method for producing the same. [Means for solving the problem]

[0007] Some embodiments of the present disclosure provide a method for producing a crystalline form II of 2,2,2-trifluoroacetic acid 1-(2,4-dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147) having a powder X-ray diffraction pattern including peaks located at 13.37, 18.47, and 23.34 (±0.2)° at 2θ values, the method comprising the steps of supplying a slurry containing saturated amorphous or crystalline form I of 2,2,2-trifluoroacetic acid 1-(2,4-dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147) in a solvent / poor solvent mixture, and mixing the slurry to obtain crystalline form II.

[0008] Some embodiments of the present disclosure provide an isolate of crystalline form I of 2,2,2-trifluoroacetic acid 1-(2,4-dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147) having a powder X-ray diffraction pattern including peaks located at 11.85, 17.11, 17.79 and 23.40 (±0.2)° at 2θ values.

[0009] The present patent or application file shall include at least one drawing made in color. A copy of this patent or patent application publication containing the color drawing shall be provided by the Office upon request and payment of the required fees.

[0010] Various embodiments of this disclosure are described below with reference to the figures herein. [Brief explanation of the drawing]

[0011] [Figure 1] Figure 1 shows the X-ray diffraction (XRD) diffraction chart of crystalline form I of 2,2,2-trifluoroacetic acid 1-(2,4-dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147). [Figure 2A] Figure 2A shows the differential scanning calorimetry (DSC) thermogram of crystal form I of J147. [Figure 2B] Figure 2B shows the thermogravimetric analysis (TGA) thermogram of crystal form I of J147. [Figure 3A] Figure 3A shows the dynamic water vapor sorption isotherm of crystal form I of J147 using TGA. [Figure 3B] Figure 3B shows the dynamical plot of crystal form I of J147 using TGA. [Figure 4] Figure 4 shows the Fourier transform infrared (FTIR) spectrum of crystal form I of J147. [Figure 5] Figure 5 shows the Raman spectrum of crystal form I of J147. [Figure 6] Figure 6 shows the nuclear magnetic resonance (NMR) spectrum of crystal form I of J147. [Figure 7] Figure 7 shows the graphical output using a Debye ring integral with a two-dimensional detector during XRD analysis of a single sample of J147. [Figure 8] Figure 8 shows the phylogenetic tree of all XRD results from the screening of the J147 sample. [Figure 9] Figure 9 shows a cell plot of all XRD results from the screening of the J147 sample. [Figure 10] Figure 10 shows the cluster plot of all XRD results from the screening of the J147 sample. [Figure 11A] Figure 11A shows the DSC thermogram of crystal form I of J147. [Figure 11B] Figure 11B shows the XRD spectrum of crystal form IJ147. [Figure 12A] Figure 12A shows a micrograph of a representative example of crystal form IJ147. [Figure 12B] Figure 12B shows another representative micrograph of crystalline form IJ147. [Figure 13] Figure 13 shows the XRD spectrum of crystalline form IIJ147. [Figure 14A] Figure 14A shows the DSC thermogram of crystalline form II of J147. [Figure 14B] Figure 14B shows the TGA thermogram of crystalline form II of J147. [Figure 15] Figure 15 shows a micrograph of a representative example of crystalline form IIJ147. [Figure 16A] Figure 16A shows the moisture adsorption / desorption isotherm of crystalline form IIJ147 using dynamic vapor sorption (DVS) analysis. [Figure 16B] Figure 16B shows the kinetic plot of crystalline form IIJ147 using DVS. [Figure 17] Figure 17 shows the computer-predicted powder XRD pattern of a single crystal of form II of J147. The predicted pattern matched the pattern of the measured pattern.

Mode for Carrying Out the Invention

[0012] Embodiments herein are directed to various polymorphic forms of 1-(2,4-dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147) of 2,2,2-trifluoroacetic acid and methods for preparing those forms. The discovery of polymorphic forms of an active pharmaceutical ingredient is recognized as being of practical importance in product development. The polymorphic forms of J147 disclosed herein can affect various physicochemical properties of J147, such as, without limitation, hardness, stability, filterability, solubility, hygroscopicity, melting point, solid density, and fluidity. Altering one or more of the physicochemical properties of J147 can lead to improvements in the impact not only on subsequent manufacturing but also on pharmacokinetics and other aspects of administering the API.

[0013] In some embodiments, isolated crystalline form I of 2,2,2-trifluoroacetic acid 1-(2,4-dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147) is provided, having a powder X-ray diffraction pattern including peaks at positions of 11.85, 17.11, 17.79 and 23.40 (±0.2)° at 2θ values.

[0014] As used herein, “isolated” means the separation of the polymorphic forms, in particular J147, the other forms, and the substantial removal of impurities and, if applicable, the solvent. For example, the purity of the isolated polymorphs is at least about 95%, or about 98%, or about 99%, or about 99.5%, including up to the detection limit of impurities or 100% purity on paper.

[0015] In some embodiments, the isolated crystalline form I may further contain X-ray diffraction peaks at 2θ values ​​of 8.64, 13.36, 19.25, 21.64, and 26.81 (±0.2)°.

[0016] In some embodiments, isolated crystalline form II of 2,2,2-trifluoroacetic acid 1-(2,4-dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147) is provided, having a powder X-ray diffraction pattern including peaks at positions of 13.37, 18.47, and 23.34 (±0.2)° at 2θ values.

[0017] In some embodiments, the isolated crystal form II may further have X-ray diffraction peaks at 17.74, 20.39, 26.25, and 28.74 (±0.2)° 2θ values.

[0018] As disclosed herein in the examples described later, the spectra of Form I and Form II may contain other XRD minor peaks.

[0019] In some embodiments, a method is provided for preparing a crystalline form I of 2,2,2-trifluoroacetic acid 1-(2,4-dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147) having a powder X-ray diffraction pattern including peaks at 11.85, 17.11, 17.79 and 23.40 (±0.2)° 2θ values, the method comprising recrystallizing 2,2,2-trifluoroacetic acid 1-(2,4-dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide from an organic solvent selected from the group consisting of nitromethane, methyl ethyl ketone, tetrahydrofuran, acetone, acetonitrile, heptane, isopropyl ether, isopropyl acetate, and chloroform.

[0020] In carrying out the methods disclosed herein, generally, recrystallization techniques well known to those skilled in the art are applied. For example, a sample of J147 can be dissolved in a small amount of solvent, such as by dissolving a given solvent at a high temperature. J147 has been found to be moderately soluble in various types of solvents, including polar protic and polar aprotic solvents. J147 is generally poorly soluble in highly hydrophobic hydrocarbon solvents such as heptane, and conversely, poorly soluble in water. Therefore, such solvents can function as co-solvents or poor solvents in recrystallization. Recrystallization can be carried out with or without stirring, mixing, or agitation.

[0021] In some embodiments, the method for preparing Form I can use water as the poor solvent. In some embodiments, the ratio of the organic solvent to the poor solvent water is in the range of about 4:1 to about 1:4.

[0022] According to the methods disclosed herein, the yield of Form I can be obtained in some embodiments in the range of about 50% to about 100%, or about 90% to about 100%, as illustrated below. In some embodiments, the yield may be at least about 95%, at least about 98%, at least about 99%, or quantitatively recovered relative to the amount of J147 to be recrystallized.

[0023] A method is provided for preparing a crystalline form I of 2,2,2-trifluoroacetic acid 1-(2,4-dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147) having a powder X-ray diffraction pattern including peaks at 11.85, 17.11, 17.79 and 23.40 (±0.2)° at 2θ values, the method comprising recrystallizing 2,2,2-trifluoroacetic acid 1-(2,4-dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide from a solvent / poor solvent mixture containing an alcohol as the solvent. In some such embodiments, the solvent-to-poor solvent ratio is in the range of about 4:1 to about 1:4. In some embodiments, the solvent alcohol is a C1-C4 alcohol. That is, for example, the alcohol may be selected from the group consisting of methanol, ethanol, 1-propanol, 2-propanol, trifluoroethanol, 1-butanol, and 2-butanol. In some embodiments, the poor solvent is water or heptane.

[0024] According to the methods disclosed herein, the yield of Form I can be obtained in some embodiments in the range of about 50% to about 100%, or about 90% to about 100%, as illustrated below. In some embodiments, the yield may be at least about 95%, at least about 98%, at least about 99%, or quantitatively recovered relative to the amount of J147 to be recrystallized.

[0025] In some embodiments, a method is provided for producing crystalline form II of 2,2,2-trifluoroacetic acid 1-(2,4-dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147) having a powder X-ray diffraction pattern including peaks at 13.37, 18.47, and 23.34 (±0.2)° 2θ values, the method comprising supplying a slurry containing saturated crystalline form I of 2,2,2-trifluoroacetic acid 1-(2,4-dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147) in a solvent / poor solvent mixture containing water as the poor solvent, mixing the slurry at ambient temperature to produce crystalline form II.

[0026] In this specification, "mixing" is used to broadly include mixing, stirring, agitation, and the like.

[0027] In some embodiments, form II can be obtained via amorphous J147.

[0028] In some embodiments, the method for obtaining Form II of J147 can use a solvent selected from the group consisting of alcohols, dimethylformamide (DMF), and dimethylacetamide (DMA).

[0029] In some embodiments, the method for obtaining Form II of J147 involves the use of a solvent alcohol selected from the group consisting of methanol, ethanol, trifluoroethanol, 1-propanol, and 2-propanol.

[0030] In some embodiments, the method for obtaining Form II of J147 involves a solvent / poor solvent mixture ratio in the range of approximately 1:2 to approximately 1:1.

[0031] In some embodiments, the method for obtaining Form II of J147 involves mixing a saturated slurry of J147 for several days, for example, about 6 days. In some embodiments, the mixing may be combined with heating as needed. However, laboratory environmental conditions, i.e., about 25°C, are usually sufficient. In some embodiments, Form II may reach sufficient purity and volume after about 3 days, or about 4 days, or about 5 days. Consequently, Form II can be isolated at intervals of 6 days or more, and optionally about 7 or 8 days.

[0032] In some embodiments, the method for forming Foam II from a slurry involves the use of an apparatus including a container connected to a recirculation system. In some embodiments, recirculation can be carried out via a homogenizer apparatus to which a shear force is applied. The homogenizer apparatus may include a stator and a rotatable rotor, to which a high shear mixing force is applied by the rotation of the rotor at a speed of 250 rpm or more. The rotor may also rotate at a speed of 500 rpm or more, and even 1,000 rpm or more.

[0033] Slurry recirculation may involve controlling the slurry flow through the outlet and inlet of the container. Recirculation energy may be supplied by a pump. For this purpose, conventional flow control mechanisms such as metering pumps and valves may be used. The above method can also be carried out in continuous mode.

[0034] In some embodiments, form I or amorphous J147 may be present in a supersaturated solution dissolved in a solvent. This solution can be mixed with a poor solvent solution. A poor solvent is a solvent in which the chemical substance has low solubility. It may also be a mixture of the poor solvent and the solvent. For example, the poor solvent can include water or heptane. When the solutions are mixed, the solubility of the substance in the solvent mixture decreases, causing it to precipitate.

[0035] In some embodiments, the method may also include the step of introducing seed crystals into a container to promote crystallization. The seed crystals may be placed in a supersaturated solution or a poor solvent. Such seed crystals are selected to be insoluble in the individual solvents and solvent mixtures.

[0036] Mixing may include controlling the flow of the solution into the container. For this purpose, conventional flow control mechanisms such as metering pumps and valves may be used.

[0037] The temperature can be adjusted before they are introduced into the container. This may be achieved by any conventional temperature control device, such as a heater or cooling bath with a solution source.

[0038] In some embodiments, the recirculation system may include a homogenizer, an outlet means for transferring slurry from a container to the homogenizer, and an inlet means for receiving slurry from the homogenizer into the container. The homogenizer may include a stator and a rotatable rotor, and means for applying a high shear mixing force by the rotation of the rotor. The high shear mixing force is applied by the rotation of the rotor at a speed of 250 rpm or more. The rotor may also rotate at a speed of 500 rpm or more, and even 1,000 rpm or more.

[0039] The apparatus may include means for controlling the slurry flow via a homogenizer. For this purpose, conventional flow control mechanisms such as metering pumps and valves may be used. The apparatus may also include means for regulating the slurry temperature in the container. This may be achieved by any conventional temperature control device, such as a heater or cooling bath, with a solution source or container.

[0040] According to the methods disclosed herein, in some embodiments, as illustrated below, the yield of Form II can be obtained in the range of about 50% to about 100%, or about 90% to about 100%. In some embodiments, the yield may be at least about 95%, at least about 98%, at least about 99%, or quantitatively recovered relative to the amount of J147 recrystallized.

[0041] The following examples are provided to illustrate embodiments of the present disclosure. These examples are intended solely to illustrate the present disclosure and are not intended to limit the scope of the present disclosure in any way. Parts and percentages are based on mass unless otherwise specified. As used herein, “room temperature” refers to a temperature of about 20°C to about 25°C. [Examples]

[0042] This example describes the screening of J147 for polymorphic behavior. Screening was performed using solvent recrystallization, hydration experiments, competitive and non-competitive slurry experiments, and grinding to control the solid-state form of the test material. The resulting samples were characterized using differential scanning calorimetry (DSC), polarized light microscopy, thermogravimetric analysis (TGA), Fourier transform nuclear magnetic resonance (NMR), powder X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR), Raman spectroscopy, and dynamic vapor sorption / desorption (DVS). The polymorphic screening revealed that J147 is polymorphic, and several solid forms were identified and characterized. Thermodynamically stable forms were confirmed.

[0043] Experimental method Microscopy: The optical properties of the samples were evaluated using a Zeiss Universal microscope and / or an Olympus BX53 microscope, which consist of a polarized visible light source and a polarization analyzer. The specimens were typically mounted on microscope slides with coverslips. Observations of particle / crystal size and shape, as well as birefringence, were recorded.

[0044] Hot Stage Microscopy (Hsm): Linkam hot stage accessories were used in combination with a microscope. The specimen was placed on a microscope slide with a coverslip. The sample was heated from temperature to melting point using a Linkam TMS94 temperature control and Linksys 32 data capture software system. Observations of measurable phase changes, melting, recrystallization, and decomposition were recorded.

[0045] Proton nuclear magnetic resonance spectroscopy ( 1 (H-NMR): Samples were prepared by dissolving 1-10 mg of API in deuterated chloroform containing 0.05% (v / v) tetramethylsilane (TMS). Spectra were collected at ambient temperature using a Bruker 400 MHz NMR spectrometer.

[0046] Differential Scanning Calorimetry (DSC): Differential scanning calorimetry (DSC) is a technique used to measure the characteristic heat flux of a test specimen when scanned through a temperature gradient under a controlled atmosphere. Thermal phase transitions such as endothermic melting and exothermic decomposition were recorded. DSC data were collected using TA Instruments DSC. Generally, samples in the mass range of 1–10 mg were pressed into an aluminum sample pan and scanned from 25 to approximately 150°C at heating rates of 2, 10, 20, and 50°C / min using a nitrogen purge of 50 mL / min.

[0047] Thermogravimetric Analysis (TGA): Thermogravimetric analysis involves determining the mass of a sample as a function of temperature. TGA data were acquired using a TA Instruments Q500TGA. Generally, samples in the mass range of 2–10 mg were placed in a tare-weighted empty platinum sample pan and attached to a microbalance with a thin wire. The sample was suspended in a furnace and heated from 25 to approximately 250°C at 10°C / min using a nitrogen purge of 100 mL / min. The change in sample weight as a function of temperature was observed.

[0048] Powder X-ray diffraction (XRD): X-ray diffraction is an analytical technique used to study the crystallinity of solid materials. X-rays incident on a crystalline material are scattered in all directions. In specific directions, the scattered X-rays are structurally enhanced to form a diffracted beam. The conditions for structural diffraction are described by Bragg's law and depend on the intrinsic composition and spatial arrangement of the crystal structure. Thus, each molecular solid diffracts X-rays in different directions and with different intensities, resulting in a unique X-ray diffraction pattern. In some experiments, the sample temperature was controlled using a variable temperature hot stage.

[0049] Powder X-ray diffraction patterns were obtained using a Bruker D8 discovery diffractometer equipped with an XYZ stage, a positioning laser video microscope, and a two-dimensional HiStar region detector or scintillation detector. A CuKα 1.5406 Å source was used to irradiate the sample at 40 kV and 40 mA. The X-ray optics consisted of a 0.5 mm pinhole collimator and a Gobel mirror. θ-θ continuous scanning was performed at a sample-detector distance of approximately 30 cm, yielding an effective 20 range from 4–40. The sample was placed on a low-background quartz plate.

[0050] Hygroscopic Dynamic Vapor Adsorption (DVS): DVS is a gravimetric screening technique that measures the rate and amount of solvent (water) adsorbed onto a sample. The relative humidity or vapor concentration around the sample is varied while measuring the mass change of the sample. Vapor adsorption isotherms show the equilibrium amount of adsorbed vapor as a function of relative humidity. Isotherms are constructed using the mass values ​​at each relative humidity step. The isotherms are divided into two elements: adsorption for the step that increases humidity and desorption for the step that decreases humidity. Plots of dynamic data showing the changes in mass and humidity as a function of time are also provided.

[0051] The samples were analyzed using a TA Q2000 automated dynamic vapor sorbent analyzer. After drying the samples at 40°C for 5 hours, they were cooled to 25°C using a dry nitrogen purge until no more mass was lost at 0% RH. The samples were then exposed to 0–95% RH and returned to 0% RH at 25°C in a 5% RH step.

[0052] Fourier transform infrared spectroscopy (FTIR): Infrared spectra were acquired using a Nicolet 510M-O Fourier transform infrared spectrometer equipped with a Harrick Splitpea® total reflection attenuation device. Resolution 4 cm for each analysis. -1 Then I scanned 128 times, 4000~400cm -1 The spectrum was obtained.

[0053] Raman spectroscopy: Raman spectra were acquired using a ThermoDXR Raman dispersion spectrometer with 780 nm laser excitation. 3300~100 cm⁻¹ -1 The spectrum was obtained. The sample was analyzed as a bulk powder.

[0054] Screening of J147 samples: Initial tests were performed using XRD, DSC, TGA, proton NMR, FTIR, and Raman spectroscopy. Powder X-ray diffraction was used to test the material and determine if it was crystalline. Figure 1 shows the XRD pattern of the material that was crystalline and named Form I. The corresponding peaks and their distributions are shown in Table A below. TIFF0007883832000002.tif144114

[0055] The thermal behavior of Form I was measured by DSC and TGA. The DSC thermogram showed endothermic fusion beginning at 62.6°C. The endothermic fusion was split into peaks at 66.5°C and 74.7°C. The heat of fusion was 123.8 J / g. This split endothermic fusion was shown to be due to the coexistence of two polymorphic forms by hot stage microscopy, as described later. Scans of the TGA thermogram showed that the weight loss of the sample from 25°C to 104.7°C was less than 0.4 wt%, indicating the absence of volatile substances. Figures 2A and 2B show the DSC and TGA thermograms, respectively.

[0056] The isotherms and dynamic plots of dynamic vapor sorption are shown in Figures 3A and 3B, respectively. The material exhibited strong hydrophobicity and did not appear to have a tendency to form hydrates. A total weight loss of approximately 0.5% was observed at 95% RH. This unusual phenomenon (weight loss at high humidity) may not be due to the J147 sample, but rather to differences in the adsorption characteristics of the sample and the reference pan.

[0057] The Fourier transform infrared (FTIR) spectrum is shown in Figure 4. Visual confirmation confirms that the spectrum is consistent with the structure. The Raman spectrum matches the FTIR spectrum and is shown in Figure 5. The proton NMR data is consistent with the structure of J147 and is shown in Figure 6. The proton NMR data is also shown in tabular format in Table B below. TIFF0007883832000003.tif191163

[0058] Polymorph screening: Screening was performed using solvent-based recrystallization, and the solids were analyzed by X-ray diffraction. Further solid-state forms were explored using suspension slurry experiments and grinding. Visual Solubility Measurement: Approximately 80 mg of J147 was placed in each of 25 vials. Solvent was added, and the vials were stirred at ambient temperature for several minutes. The remaining solid was visually observed. The experiment was terminated by gradually adding solvent until the solid dissolved or until the maximum volume of solvent had been added. These stock solutions were used to set up further panels for the experiment. J147 was found to be well soluble in all solvents tested except water. The visually determined solubility is shown in Table 1.

[0059] [Table 1]

[0060] Solvent Recrystallization: To perform the solvent-based portion of polymorph screening, the test material was recrystallized using various solvents under approximately 150 different crystal growth conditions. The scale of the recrystallization experiments ranged from approximately 6 mL to 15 mL. Crystal growth conditions were varied using solvent mixtures in a binary gradient sequence. Saturated solutions were prepared by contacting the test material with various solvent systems at saturation temperature and stirring excessively (as much as possible). If the solid did not completely dissolve in the solvent, the mother liquor was filtered to separate the remaining solid. The mother liquor was then heated above saturation temperature (superheated) to dissolve the remaining solid. Subsequently, the temperature of each solution was adjusted to the growth temperature, and solvent evaporation was initiated by introducing a controlled nitrogen shear flow. Due to the solubility of J147 in most solvents, the ambient growth temperature was used in all experiments. The recrystallization conditions used in the six solvent-based panels are summarized in Table 2. Each recrystallization panel contains 15–27 wells. Within each panel, the wells contain different solvent compositions. Due to the different solvent compositions in each well, each well functioned as a different crystal growth experiment. Based on the XRD analysis performed, novel polymorphs of J147 were discovered. The first polymorph was named Form I, and the second polymorph was named Form II. Table 2 is an overview of the recrystallization panel for solvent-based polymorph screening. Tables 3-8 show the solvent matrices of the six recrystallization panels used in the solvent-based portion of the polymorph screening.

[0061] [Table 2]

[0062] [Table 3]

[0063] [Table 4]

[0064] [Table 5]

[0065] [Table 6]

[0066] [Table 7]

[0067] [Table 8]

[0068] Non-competitive slurry experiments: In addition to solvent recrystallization experiments, non-competitive slurry experiments were conducted to find new solid-state forms. This experiment utilizes the differences in solubility of different polymorphic forms (if different polymorphic forms exist for the compound). Therefore, only polymorphs with lower solubility (i.e., more stable) than the original crystalline form can arise from non-competitive slurry experiments.

[0069] When a solid is mixed with a solvent to form a slurry, a saturated solution is eventually produced. For the dissolved polymorphic form, the solution is saturated. However, for any polymorphic form that is more stable than the initially dissolved form (more stable forms have lower solubility), the solution becomes supersaturated. Therefore, any more stable polymorphic form can nucleate and precipitate from the solution. Furthermore, non-competitive slurry experiments are often useful in identifying the solvent that forms solvates with the compound.

[0070] Slurry experiments were conducted by exposing an excess supply of material to a solvent and stirring the resulting suspension at ambient temperature for several days. The solid was filtered (Whatman grade 1, pore size 11 μm), and the resulting form was determined by XRD analysis. To avoid the possibility of desolvation or physical changes after separation, the samples were not dried before X-ray analysis. A summary of the non-competitive slurry experiments is shown in Table 9.

[0071] [Table 9]

[0072] Based on these X-ray dispersion behaviors, all slurry experiments with Form I yielded Form II after 6 days of slurring. These data indicate that Form II is more stable than Form I at ambient temperature and pressure. No new polymorphs, solvates, or hydrates were separated in these experiments.

[0073] Solids synthesized from solvent-based recrystallization panels were analyzed by powder XRD. To mitigate preferential particle effects, all XRD screening data were collected using a two-dimensional detection system. The two-dimensional detector integrates along concentric Debye rings, reducing pattern variation. An example of Debye ring integration using a two-dimensional detector is shown in Figure 7. Where bright spots appear in the conical ring, it indicates a strong preferential particle effect, which can lead to significant variation in the observed diffraction pattern, including changes in peak intensity. Several samples of J147 showed preferential particle effects based on the appearance of scattering behavior.

[0074] The results of this analysis revealed that the material exists as two distinct polymorphs. These polymorphs were named Form I and Form II. The first material obtained was named Form I. The names of the forms obtained for individual (solvent-based) recrystallization experiments are shown in Tables 3-8. XRD data collected during the experiment were evaluated using full-profile chemometrics to determine whether the crystalline form of the sample changed during the experiment. To simplify evaluation, X-ray amorphous samples were not included in the chemometrics treatment. The analysis inevitably involved cluster analysis and multivariate statistics to group all patterns deemed statistically identical. The results of this analysis are summarized in the dendrogram in Figure 8, the cell plot in Figure 9, and the cluster plot in Figure 10. These figures provide image cluster analysis of product similarity.

[0075] Chemometric analysis of diffraction data classified the samples into three distinct groups (or clusters) labeled A to C. A summary showing the number of members in each group (A to C) is shown in Table 10.

[0076] [Table 10]

[0077] The characteristics of each group / form are as follows. After classifying the data into different forms based on diffraction behavior, each form was studied to determine whether other properties of the forms could be distinguished. Characterization of each form began by comparing the diffraction data represented by each form with the diffraction data of the other forms. This was followed by typical DSC, TGA, DVS analysis and microscopic examination.

[0078] Form I (Group B): The characteristic diffraction behavior of this form is shown in Figure 1. The XRD patterns of all Form I samples are crystalline and very similar. This form was obtained from various solvents in approximately 50% of the experiments. The DSC profiles of Form I samples showed endothermic fusion starting at approximately 63°C. In all but one of the Form I samples analyzed, the maximum endothermic peak split in the range of 66.5°C to 74.7°C. This splitting endothermic fusion is thought to be due to the mixing of two polymorphic forms observed with a hot stage microscope. A representative DSC thermogram is shown in Figure 2A. The Form I sample showed a single endothermic fusion starting at approximately 63°C in its DSC profile. The XRD pattern was consistent with the patterns of the other Form I samples. Figures 11A and 11B show the DSC and XRD profiles of this sample, respectively. The TGA thermogram scan of Form I (Figure 2B) showed a weight loss of less than 0.4% by weight between 25°C and 104.7°C, indicating the absence of volatile substances.

[0079] Microscopic observation of Form I revealed a mixture of cylindrical and needle-shaped particles ranging in length from approximately 10 to 300 microns. Upon heating, the larger particles appeared to melt at approximately 67°C, while the smaller needles remained unmelted. Figures 12A and 12B show representative micrographs at room temperature and 67°C. This, along with multiple endothermic findings in the DSC profile, suggests that the majority of Form I samples may actually be mixtures of Form I and Form II.

[0080] The dynamic vapor sorption isotherms and kinetic plots of representative samples of Form I are shown in Figures 3A and 3B, respectively. This material was highly hydrophobic and showed no tendency to form hydrates. A total weight loss of approximately 0.5% was observed at 95% RH. This unusual event (weight loss at high humidity) is most likely due to differences in the adsorption properties of the sample and the reference pan, rather than being caused by the J147 sample. The Form I sample was placed in an oven at 40°C over the weekend. XRD data showed a transition to Form II. Variable temperature (VT) XRD experiments were performed on Form I. The Form I sample was held for XRD at 35°C over the weekend without any form transition being observed. Overall, Form I was considered to be due to the dry crystalline nature and polymorphic foam of the compound.

[0081] Form II (Group A): Although the Form II polymorph was not obtained from recrystallization screening experiments, it was observed in approximately 50% of all experiments. The XRD patterns representing Form II samples indicate that the samples were crystalline and very similar. Figure 13 shows the characteristic XRD pattern of Form II. A summary of the peaks is shown in Table C below. TIFF0007883832000014.tif132114

[0082] The DSC thermogram of this form shows specific endothermic melting with extrapolation start at approximately 74°C, maximum peak at approximately 75°C, and melt enthalpy of approximately 68 J / g. Figure 14A shows the DSC profile of form II. The TGA thermogram scan of form II (Figure 2B) showed less than 0.1% weight loss from 25°C to 75°C and no volatile substances. Microscopic observation of Form II revealed birefringent, needle-shaped particles ranging in length from approximately 10 to 50 microns. Figure 15 shows a representative micrograph of Form II crystals at room temperature. Upon heating, one melting event was observed at approximately 75°C. Figures 16A and 16B show the absorption and desorption isotherms and kinetic plots, respectively, for Form II. As seen in Form I, this form was also highly hydrophobic and showed no tendency to form hydrates. At 95% RH, a total weight loss of approximately 0.5% was observed. This unusual event (weight loss at high humidity) is most likely due to differences in the adsorption properties of the sample and the reference pan, rather than being due to the J147 sample. The single-crystal predicted powder XRD pattern was consistent with the Form II pattern obtained in the experiment shown in Figure 17. The Form II sample was placed in a 40°C oven over the weekend. The XRD data did not show any form transition. Overall, Form II was considered to be due to a dry crystalline polymorph form of the compound.

[0083] In this example, several J147 samples were generated. Polymorph screening recrystallization experiments produced either form I or amorphous form of J147 (shown in Table 11). By generating a saturated slurry of form I and stirring / agitating it at ambient temperature for several days, form II polymorphs were generated. Many of the form I samples contained some form II, indicating a tendency for the two polymorphs to nucleate and co-grow. Based on the results of competitive slurry experiments, form II appears to be a thermodynamically stable form under ambient conditions.

[0084] [Table 11]

[0085] Competitive Slurry Experiments: In addition to solvent recrystallization experiments, two competitive slurry experiments were also performed to identify the most stable form. This experiment utilizes the difference in solubility of different polymorphic forms. Therefore, only polymorphic forms (and solvates) with lower solubility (more stable) than the first dissolved form can be produced by the competitive slurry experiment. The slurry experiments were performed by exposing excess material of Form I and Form II to a small amount of solvent / water and stirring the resulting suspension at ambient temperature for several days. The solid was filtered and the obtained forms were identified by analysis by XRD and DSC. Samples were not dried before X-ray analysis to avoid the possibility of desolvation or physical changes after separation. The results of the competitive slurry experiments are shown in Table 12.

[0086] [Table 12]

[0087] Competitive slurry experiments resulted in the transformation of the sample to form II. This result suggests that, at ambient temperature and pressure, form II is less soluble and a more thermodynamically stable polymorph than form I.

[0088] Grinding: Form I was ground using a Crescent Wig-L-Bug ball mill at 4800 vibrations / min (3.2 m / s) for 1 minute. XRD analysis showed no transformation under these conditions.

[0089] Recrystallization using heating: Amorphous / glass samples from Panel 1 were placed in a vacuum oven at 60°C for 6 days. XRD analysis of this solid showed an XRD pattern of Form II. A sample of Form I was placed in an oven at 40°C over the weekend. XRD data showed a transition to Form II. Variable temperature (VT) XRD experiments were performed on Form I. A sample of Form I was held for XRD at 35°C over the weekend without any form transition being observed.

[0090] Thermodynamic Relationships: DSC experiments were performed to obtain heat of fusion and melting data. This data is often used to determine whether polymorphs exist in an enantiotropic or monotropic relationship. The law of heat of fusion states that if the heat of fusion of a high-melting-point polymorph is low, the two forms are enantiotropic. Conversely, if the heat of fusion of a high-melting-point polymorph is high, the two forms are monotropic. In a monotropic system, the transition from one polymorph to another is irreversible. In an enantiotropic system, a reversible transition is possible between the two polymorphs by heating or cooling.

[0091] Pure Form I samples and two Form II samples were analyzed by DSC using small sample sizes at a low heating rate of 2°C / min. Table 13 shows the average melting point temperature and heat of fusion data for 10 experiments. These data indicate that Form I has a low melting temperature and Form II has a high melting temperature. The heat of fusion is very close to that of the Form I sample with a high standard deviation.

[0092] [Table 13]

[0093] Static vapor sorption studies: Dynamic vapor sorption tests were conducted in a sealed humidity chamber equipped with an automatic moisture absorption balance. Data collected during dynamic vapor sorption tests are often not in thermodynamic equilibrium. Samples were observed in a 75% static humidity chamber to determine whether the Form I and Form II materials formed hydrates over time.

[0094] During these studies, samples were stored in open petri dishes in a chamber containing a saturated salt solution to maintain relative vapor pressure. A saturated sodium chloride solution (75% RH) was used at ambient temperature.

[0095] The samples were gravimetrically analyzed at 5 and 12 days. After both periods, neither Form I nor Form II showed a significant weight change, indicating no hydrate formation.

[0096] The raw diffraction data generated in the polymorph screening experiment was classified into two polymorphic forms. Additional experiments (DSC, TGA, HSM, etc.) were performed using samples of these different forms to refine the forms. A summary of the discovered polymorphic forms is provided in Table 14.

[0097] [Table 14]

[0098] Table 14 summarizes the different solid-state forms of J147 observed during the study. The main result is the discovery of two anhydrous polymorphic forms of J147. The polymorphic forms isolated in this example were named I and II. At room temperature and atmospheric pressure, form II is the thermodynamic form of J147. Form I is the metastable form at room temperature and atmospheric pressure.

[0099] Various experimental results confirmed that form I is a metastable form at room temperature and pressure. Evidence of the transition from form I to form II was observed after storage at 40°C for approximately 3 days. Competitive slurries (at ambient temperature) containing a 50 / 50 mixture of form I and form II in two different solvent systems showed the transformation from form I to form II after 4 and 8 days. Non-competitive slurry experiments of form I in seven different solvent systems showed the transition to form II after 6 days.

[0100] Screening required solvent crystallization, heating, grinding, sorption experiments, and competitive and non-competitive slurry experiments on the material. Overall, J147 was recrystallized under more than 150 different crystal growth conditions and analyzed using powder X-ray diffraction. The samples were classified into different groups using the X-ray data. These groups were studied using thermal, optical, spectroscopic, and other means to elucidate the intrinsic solid-state forms of the API. In general, J147 exhibits two distinct polymorphic forms, named Form I and Form II, in addition to the amorphous form. No solvates or hydrates were found in this example. Of the two polymorphic forms, Form II was the thermodynamically stable polymorph under environmental conditions.

[0101] Preparation example of Form II of J147 Batch method: Approximately 100 kg of crude J147 derived from the synthetic preparation was concentrated twice with approximately 80 kg of ethanol. The crude product was incorporated into approximately 48 kg of ethanol, and the batch temperature was adjusted to 28°C. Approximately 37 kg of water was gradually added to the batch. The batch was held at approximately 30°C for approximately 1.7 hours. The batch sample was removed from the reactor, and 45 mL of water was added to precipitate the solid. The obtained solid was returned to the batch as a seed crystal, and the mixture was stirred at 30°C for 40 minutes. Approximately 34 kg of water was added further. The batch was held at approximately 18°C ​​for approximately 58 hours, then cooled to approximately 10°C and held for a further approximately 5.5 hours. Analysis of the obtained solid showed the presence of form I. Form I was converted to form II by heating the slurry to approximately 45°C for approximately 16 hours, then cooling again to approximately 10°C, and holding the batch at this temperature for approximately 3 hours. After washing, drying, and filtration, approximately 17.7 kg of J147 solid foam II was recovered.

Claims

1. A method for producing crystals of crystalline form II of 2,2,2-trifluoroacetic acid 1-(2,4-dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147), having a powder X-ray diffraction pattern including peaks at 13.37, 18.47, and 23.34 (±0.2)° at 2θ values, (a) A step of preparing a slurry containing 2,2,2-trifluoroacetic acid 1-(2,4-dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147) of crystals of crystal form I having a powder X-ray diffraction pattern with saturated amorphous or 2θ peaks at 11.85, 17.11, 17.79 and 23.40 (±0.2)° in a mixture containing a solvent and a poor solvent, and (b) A step of mixing the slurry to obtain crystals of crystal form II, Methods that include...

2. The method according to claim 1, wherein the slurry contains saturated J147 of the crystal form I.

3. The method according to claim 1 or 2, wherein the mixing step (b) is carried out at a temperature in the range of 25°C to 50°C.

4. The method according to claim 1 or 2, wherein the mixing step (b) is performed at a temperature in the range of 40°C to 50°C.

5. The method according to any one of claims 1 to 4, wherein the poor solvent is water.

6. The method according to any one of claims 1 to 5, wherein the solvent is an alcohol.

7. The method according to claim 6, wherein the alcohol is selected from the group consisting of methanol, ethanol, trifluoroethanol, 1-propanol, and 2-propanol.

8. The method according to claim 6, wherein the alcohol is ethanol.

9. The method according to any one of claims 1 to 5, wherein the solvent is dimethylformamide (DMF) or dimethylacetamide (DMA).

10. The method according to any one of claims 1 to 4, wherein the solvent is ethanol and the poor solvent is water.

11. The method according to any one of claims 1 to 10, wherein the ratio of the solvent to the poor solvent is in the range of 4:1 to 1:

4.

12. The method according to any one of claims 1 to 11, wherein the ratio of the solvent to the poor solvent is 1:

2.

13. The method according to any one of claims 1 to 12, wherein the mixing step (b) continues for 6 hours to 6 days.

14. The method according to any one of claims 1 to 13, wherein the yield of crystals in the crystal form II is in the range of 50% to 100%.

15. The method according to any one of claims 1 to 14, wherein the purity of the crystals of the crystal form II is at least 98%.

16. The method according to any one of claims 1 to 15, wherein the crystals of the crystal form I of J147 are obtained by recrystallizing 2,2,2-trifluoroacetic acid 1-(2,4-dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide from an organic solvent selected from the group consisting of nitromethane, methyl ethyl ketone, tetrahydrofuran, acetone, acetonitrile, heptane, isopropyl ether, isopropyl acetate, and chloroform, in the presence of water as a poor solvent if necessary.

17. A method for producing crystals of crystalline form I of 2,2,2-trifluoroacetic acid 1-(2,4-dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147), having a powder X-ray diffraction pattern including peaks at 11.85, 17.11, 17.79 and 23.40 (±0.2)° at 2θ values, A method comprising recrystallizing 2,2,2-trifluoroacetic acid 1-(2,4-dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide from an organic solvent selected from the group consisting of nitromethane, methyl ethyl ketone, tetrahydrofuran, acetone, acetonitrile, heptane, isopropyl ether, isopropyl acetate, and chloroform, and optionally further containing water as a poor solvent.

18. The method according to claim 17, wherein the recrystallization is carried out using water as the poor solvent, and the ratio of the organic solvent to the poor solvent is in the range of 4:1 to 1:

4.

19. A method for producing crystals of crystalline form I of 2,2,2-trifluoroacetic acid 1-(2,4-dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147), having a powder X-ray diffraction pattern including peaks at 11.85, 17.11, 17.79 and 23.40 (±0.2)° at 2θ values, A method comprising the step of recrystallizing 2,2,2-trifluoroacetic acid 1-(2,4-dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide from a mixture containing a solvent and a poor solvent, wherein the solvent is an alcohol and the poor solvent is water or heptane.

20. Crystals of isolated crystalline form I of 2,2,2-trifluoroacetic acid 1-(2,4-dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147) having a powder X-ray diffraction pattern including peaks at 11.85, 17.11, 17.79 and 23.40 (±0.2)° at 2θ values, wherein the powder X-ray diffraction pattern further includes X-ray diffraction peaks at 8.64, 13.36, 19.25, 21.64 and 26.81 (±0.2)° at 2θ values.

21. Crystals of isolated crystalline form II of 2,2,2-trifluoroacetic acid 1-(2,4-dimethylphenyl)-2-[(3-methoxyphenyl)methylene]hydrazide (J147) having a powder X-ray diffraction pattern including peaks at 13.37, 18.47 and 23.34 (±0.2)° at 2θ values, wherein the powder X-ray diffraction pattern further includes X-ray diffraction peaks at 17.74, 20.39, 26.25 and 28.74 (±0.2)° at 2θ values.

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