Denatonium acetate monohydrate

Abstract

Novel monohydrate salt forms of denatonium acetate are disclosed. More specifically, the novel salt and hydrate crystal forms are useful for the treatment and prevention of diseases and conditions such as metabolic syndrome, obesity, NASH, glycemic control / diabetes and IBD (intestinal disease).

Description

[Technical field]

[0001] (CROSS REFERENCE TO RELATED APPLICATIONS) This disclosure claims the benefit of priority to U.S. Provisional Application No. 63 / 255,947, filed October 14, 2021, which is incorporated herein by reference in its entirety for all purposes.

[0002] FIELD OF THEINVENTION The present disclosure provides novel monohydrate salt forms of denatonium acetate. More specifically, the novel salt and hydrate crystal forms are useful for the treatment and prevention of diseases and conditions, including, inter alia, metabolic syndrome, obesity, NASH, glycemic control / diabetes, Prader-Willi syndrome, and IBD (intestinal bowel disease). [Background technology]

[0003] Denatonium acetate is a salt described in US2015 / 0252305 ("The bittering agent is preferably a denatonium salt or derivative thereof. In one embodiment, the bittering agent is a denatonium salt selected from the group consisting of denatonium chloride, denatonium citrate, denatonium saccharide, denatonium carbonate, denatonium acetate, denatonium benzoate, and mixtures thereof. In one embodiment, the liquid composition comprises a first denatonium salt and the film comprises a second denatonium salt different from the first denatonium salt."). However, it is believed that denatonium acetate has not been synthesized and has only been described as a number of theoretical salts with denatonium as the cation. A series of patent applications subsequently assigned to Aardvark Therapeutics, Inc. described desodium acetate as a pharmaceutical composition and included a synthetic process for synthesizing desodium acetate from lidocaine. See, e.g., WO2021 / 062061. This synthesis process produces denatonium acetate anhydrate (DAA). However, it was subsequently determined that DAA is not suitable for use as an API (active pharmaceutical ingredient) due to its insufficient stability and FDA regulatory requirements for the stability of active pharmaceutical ingredients that DAA does not meet. Thus, there is a need in the art for a more stable salt form of denatonium acetate (DA). Desodium acetate is known to be useful in the treatment of various conditions, such as metabolic syndrome, obesity and hyperglycemia. See, for example, WO2021 / 133908. Summary of the Invention

[0004] The present disclosure is based on the discovery that DAA is unstable and decomposes under at least some conditions. DAA is hydrated and recrystallized into a stable salt, namely, denatonium acetate monohydrate (DAM). The present disclosure provides a novel denatonium acetate monohydrate salt and its crystalline form. Desodium acetate monohydrate and crystalline form can provide advantages in the manufacture of desodium acetate, such as higher stability, handling and administration. In particular, DAM can show improved physical and chemical stability against stress, high temperature and humidity, for example, compared to DAA.

[0005] The present disclosure provides compounds of formula 1: [ka] The present invention provides a denatonium acetate monohydrate salt having the following structure: Preferably, the salt is a monohydrate crystal. The salt may be characterized by an X-ray powder diffraction (XRPD) spectrum substantially as shown in FIG. 5A or FIG. 5B.

[0006] The present disclosure further provides a pharmaceutical composition comprising a therapeutically effective amount of denatonium acetate monohydrate salt crystals in combination with one or more pharma- ceutical acceptable carriers. The one or more pharma-ceutical acceptable carriers can comprise a biocompatible polymer. The biocompatible polymer can be cellulose. The one or more pharma-ceutical acceptable carriers can comprise a saccharide. The saccharide can be a sugar alcohol. The sugar alcohol can be mannitol. The one or more pharma-ceutical acceptable carriers can comprise talc. The one or more pharma-ceutical acceptable carriers can comprise an organic acid. The organic acid can be acetic acid. The pharmaceutical composition can be formulated for oral administration. The pharmaceutical composition can comprise a solid granule.

[0007] The present disclosure also provides a method for producing denatonium acetate monohydrate salt crystals, comprising contacting an equivalent amount of DAA (denatonium acetate anhydride) with a lower alkyl isobutyl ketone, such as methyl isobutyl ketone, and water such that the concentration of water is greater than 10% by weight, recovering the resulting solid phase, and removing the solid therefrom. The present disclosure provides a method for producing denatonium acetate monohydrate salt crystals described herein, comprising contacting denatonium acetate anhydride with a lower alkyl isobutyl ketone and water to form a solid phase comprising denatonium acetate monohydrate. The lower alkyl isobutyl ketone can be methyl isobutyl ketone or ethyl isobutyl ketone. The contact of denatonium acetate anhydride with the lower alkyl isobutyl ketone and water forms a composition having a water concentration greater than 10% by weight. Denatonium acetate monohydrate is a monohydrate crystal. The monohydrate crystal can be characterized by an X-ray powder diffraction (XRPD) spectrum substantially as shown in FIG. 5A or FIG. 5B. [Brief description of the drawings]

[0008] [Figure 1] FIG. 1 shows the synthetic process for producing DAM from DAA.

[0009] [Diagram 2] FIG. 2 shows a form diagram illustrating the relationship between the polymorphic patterns observed for DAM.

[0010] [Diagram 3] Figure 3 shows the overlaid diffractograms of the DAMs of pattern 1 (lower trace) and pattern 2 (upper trace).

[0011] [Figure 4] Figure 4 shows the overlaid diffractograms of the DAMs of pattern 1 (lower trace) and pattern 5 (red) (upper trace).

[0012] [Figure 5A] FIG. 5A shows the diffractogram of the DAM of pattern 1.

[0013] [Figure 5B] FIG. 5B shows a zoomed-in version of the diffractogram of the DAM of pattern 1 from 0 to approximately 5500 counts. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0014] The process of synthesizing DAM begins with lidocaine base, producing denatonium hydroxide, which then produces DAA.

[0015] Step 1: Synthesis of denatonium hydroxide from lidocaine In a reflux apparatus, add 25 g of lidocaine, 60 mL of water and 17.5 g of benzyl chloride and, with stirring, heat to 70-90 ° C. The solution must be heated and stirred at the aforementioned values ​​for 24 hours, and then cooled to 30 ° C. The unreacted reagents are removed with 3 × 10 mL of toluene. With stirring, 65 g of sodium hydroxide is dissolved in 65 mL of cold water and added to the aqueous solution with stirring over 3 hours. The mixture is filtered, washed with water and dried in air. Recrystallize in hot chloroform or hot ethanol. [ka]

[0016] Step 2: Preparation of denatonium acetate anhydrous (DAA) from denatonium hydroxide To a reflux apparatus, add 10 g of denatonium hydroxide (MW: 342.475 g / mol, 0.029 mol), 20 mL of acetone, and 2 g of glacial acetic acid (0.033 mol) dissolved in 15 mL of acetone, stir the mixture and heat to 35° C. for 3 h, then evaporate to dryness and recrystallize in hot acetone. [ka]

[0017] It was found that DAA decomposed and DAA decomposed into (A) lidocaine and benzyl acetate or (B) 2-(diethylamino)-3-phenyl-N-(2,6-dimethylphenylpropionamide). Therefore, DAA required low temperature to be used as an intermediate. In step 3, DAA was kept in the organic layer and distilled under vacuum until the temperature reached 65-67° C. under less than 150 torr. Methyl isobutyl ketone was added and refluxed under vacuum to remove water by azeotropic distillation to form DAM. DAM was crystallized by adding isopropyl alcohol. Residual salts were removed. The mixture was distilled under vacuum. Methyl isobutyl ketone was then added followed by water. In some embodiments, a lower alkyl isobutyl ketone is used in place of methyl isobutyl ketone. In some embodiments, the lower alkyl is 1-3 In some embodiments, the lower alkyl is methyl or ethyl. The temperature was reduced to below 10° C. The remaining solid was isolated and washed with methyl isobutyl ketone to obtain the final DAM (denatonium acetate monohydrate).

[0018] The present disclosure provides denatonium acetate monohydrate crystals. In some embodiments, the denatonium acetate monohydrate crystals are characterized by an X-ray powder diffraction (XRPD) spectrum substantially as shown in FIG. 5A or FIG. 5B.

[0019] In some embodiments, a pharmaceutical composition is provided that includes denatonium acetate monohydrate and a pharma- ceutically acceptable carrier. The pharma- ceutically acceptable carrier may include, for example, a biocompatible polymer, such as cellulose, and / or a saccharide, such as a sugar alcohol, such as mannitol. In some embodiments, the pharma- ceutical composition includes talc. In some embodiments, the pharma- ceutical composition includes talc, cellulose, and a saccharide. In some embodiments, the pharmaceutical composition is an oral formulation. In some embodiments, the pharmaceutical composition (e.g., oral formulation) is a sustained release cellulose-based and mannitol additive formulation. In some embodiments, the pharma- ceutical composition includes an organic acid, such as acetic acid, which may be present in combination with any of the above-mentioned components or combinations of components. The pharmaceutical composition may be formulated for oral administration. The pharmaceutical composition may include solid granules. EXAMPLES

[0020] Example 1 This example describes the characterization of a batch of denatonium acetate monohydrate (DAM) and the polymorph screening studies performed with the material. DAM is highly crystalline and has an XRPD pattern designated Pattern 1 (FIG. 5A, enlarged version in FIG. 5B).

[0021] Gravimetric vapor sorption (GVS) experiments performed on DAM and storage under stress conditions show that the material is highly hygroscopic at relative humidity (RH) levels above 80%. Deliquescence was observed after 5 days at 96% RH. Mass loss occurred when relative humidity levels dropped below 10%. DAM samples kept at 0% RH transformed to pattern 5. This pattern was metastable and transformed to pattern 1 within a few hours under ambient laboratory conditions.

[0022] DAM was thermally unstable when heated to 150° C. (slightly above its melting point) in the DSC and UPLC analysis showed that only about 54.9% of the input material remained.

[0023] Attempts to produce amorphous material by lyophilization, rapid evaporation or anti-solvent precipitation from various solvents did not yield amorphous material. Pattern 1 as supplied was used as input material for polymorph screening studies with 48 solvents as well as solvent / anti-solvent screening studies. Four additional XRPD patterns were found during all investigations of DAM.

[0024] Pattern 2 was obtained from an attempt to produce amorphous material by lyophilization of a solution of DAM in t-butanol. After 7 weeks of storage in a closed vial, it was observed to transform into pattern 1.

[0025] Pattern 3 (from evaporation of a solution of DAM in ethyl formate) 1 The pattern was found to be a mixture of denatonium acetate and formate salts based on 1 H NMR and is therefore not representative of that which would be representative of denatonium acetate alone.

[0026] Pattern 4 was obtained from a polymorph screening by evaporation of a solution of DAM in 2-methoxyethanol. Samples reanalyzed by XRPD after storage in closed vials for 6 weeks show conversion to pattern 1.

[0027] Pattern 5 was observed by storing DAM at 0% RH, drying under vacuum at room temperature, or by heating to 120 °C and rapid cooling in the DSC. When left on an XRPD disc under ambient laboratory conditions, it was observed to readily transform into pattern 1 after 4 h. A form diagram showing the relationship between the patterns observed is shown in Figure 2.

[0028] A summary of the initial characterization of the DAM is shown in Table 1. [Table 1]

[0029] Measure the particle size distribution and D 90 The ES+ mass spectrum of DAM showed a bimodal distribution at approximately 65 μm. The M+ The sample was highly crystalline. Thermal analysis of DAM by differential scanning calorimetry (DSC) at 10 °C / min showed a broad and shallow endothermic event between 44 and 124 °C, a broad endothermic event with onset at 135 °C consistent with possible melting, and a further endothermic event between 149 and 206 °C. Thermogravimetric analysis (TGA) at 10 °C / min showed mass losses of 4.3% from 40 to 153 °C, 12.3% from 154 to 192 °C, and 7.8% from 193 to 220 °C, followed by further mass loss thereafter.

[0030] Several gravimetric vapor sorption (GVS) experiments were performed to investigate the behavior of DAM for various relative humidity levels. A GVS experiment (GVS1) over a range of relative humidity (40-90-0-90-0-40%) showed that DAM was extremely hygroscopic, with a 23% mass gain in the 0-90% relative humidity range (second sorption cycle). Most of the mass gain was observed above 80% RH. Of note, the experiment did not reach equilibrium at the highest humidity stage (even using a method that allowed up to 24 h at each stage if equilibrium was not reached), so the 23% mass gain is likely an underestimate of hygroscopicity.

[0031] A GVS experiment (GVS2) with DAM (DAM, Lot: ED21356-001-001-00) was carried out to investigate the behavior of the material at relative humidity levels up to 80%. In this experiment, a similar method to GVS1 was used, except for the step involving 90% RH, where a large mass change was observed and equilibrium was not reached. GVS2 over the relative humidity range (40-80-0-80-0-40%) starting from the adsorption cycle showed a mass gain of 2.1% (at 0-80% relative humidity in the second adsorption cycle), with the majority of this mass change (+1.4%) occurring at 0-10% RH. The material remained a free-flowing powder after the experiment, and XRPD analysis showed a diffractogram consistent with pattern 1.

[0032] A further GVS experiment (GVS3) of DAM over a range of relative humidities (40-0-80-0-80-0-40%) was carried out to determine if there was a noticeable difference in behavior when the high (80%) exposure was not allowed prior to the start of the 0-80% RH adsorption cycle. This experiment (GVS3), starting with the desorption cycle, showed a mass increase of 2% (at 0-80% relative humidity in both the first and second adsorption cycles). This was the same for both adsorption cycles in the experiment and was comparable to the 2.1% result obtained in GVS2. The majority of the mass change (an increase of 1.3-1.4%) occurred between 0-10% RH. The material remained a free-flowing powder after the experiment and XRPD analysis showed a diffractogram consistent with pattern 1.

[0033] GVS experiments performed with DAM and storage under stress conditions show that the material is extremely hygroscopic at relative humidity levels above 80%. It showed a mass gain of 1.4% at 0-10% RH, 2.1% at 0-80% RH (GVS2) and 23% at 0-90% RH (GVS1). After 5 days of storage at 96% RH, it was observed to deliquesce. The mass also decreases when the relative humidity level drops below 10%.

[0034] After observing the mass loss of DAM going from 10% RH to 0% RH in the GSV experiment described above, a portion of the DAM was placed in an open vial and kept at 0% RH in the GSV instrument for 2 days. It was then removed from the instrument and a portion was immediately analyzed by XRPD. It showed a diffractogram consistent with pattern 5. Reanalysis of the sample after standing on a disk under ambient laboratory conditions for 4 hours showed conversion to pattern 1.

[0035] GVS and stressed storage experiments carried out with DAM show that the material is highly hygroscopic at relative humidity levels above 80%. Relative humidity levels below 10% result in mass loss. DAM samples kept at 0% RH transformed to pattern 5, which is metastable and transforms to pattern 1 within hours under ambient laboratory conditions.

[0036] A series of additional DSC experiments were performed to further investigate the thermal behavior of DAM. A sample of DAM (5 mg) was heated to 150° C. in the DSC at 10° C. / min and then cooled to 30° C. After the experiment, the pan was removed from the instrument, opened, and the material (DAM, Lot: ED21356-003-001-00) examined for signs of decomposition. UPLC showed significant signs of decomposition, with only about 54.9% of the input material remaining. 1 1 H NMR also shows evidence of substantial decomposition.

[0037] A sample of DAM (4.55 mg) was heated to 120 in the DSC at 10° C. / min, held at 120° C. for 2 min, and then cooled to 30° C. After the experiment, the pan was removed from the instrument, opened, and the material examined for signs of decomposition. UPLC and 1 1 H NMR showed no noticeable signs of decomposition.

[0038] DSC experiments performed with DAM indicate that the material is thermally unstable when heated to 150° C., with significant decomposition indicated by NMR and UPLC. No significant chemical decomposition was observed when the sample was heated briefly to 120° C., but a change from pattern 1 to pattern 5 was observed in the XRPD diffractogram. After standing overnight at room temperature the material reverted to pattern 1. Pattern 5 is metastable and converts to pattern 1 under ambient laboratory conditions after standing overnight on the XRPD disc. A summary of the initial and further characterization of DAM is shown in Table 2. [Table 2]

[0039] A portion of DAM (8 mg) was completely dissolved in t-butanol (0.5 mL) and then frozen on dry ice. The frozen sample was lyophilized in a freeze dryer until all solvent was removed. The resulting solid material (DAM, Lot: ED21356-005-001-01) was analyzed by XRPD. It was not amorphous and showed a different pattern from the input material. This was designated Pattern 2. Figure 3 shows the overlaid diffractograms of Pattern 1 (black) and Pattern 2 (red) DAM. A sample of Pattern 2 was reanalyzed by XRPD after 7 weeks of storage in a closed vial and showed a diffractogram consistent with Pattern 1.

[0040] A portion of DAM (10.9 mg) was dried under vacuum at room temperature overnight to give solid ED21356-004-001-00. It was analyzed by XRPD and found to be non-amorphous. It showed diffraction peaks consistent with crystalline material. It showed a pattern very similar to Pattern 1, but with an additional peak at 18° 2θ, and some other minor differences, as shown in the overlay in Figure 4. This was designated Pattern 5.

[0041] Attempts to produce amorphous material under these test conditions by vacuum drying, rapid evaporation from DCM, antisolvent precipitation, or lyophilization from water, 1,4-dioxane / water (1:1), MeCN / water (1:1), or t-butanol did not yield amorphous material. Crystalline material consistent with pattern 1 was obtained by rapid evaporation from DCM or lyophilization from water, 1,4-dioxane / water (1:1), or MeCN / water (1:1). Rapid addition of a concentrated solution of DAM in DCM to heptane resulted in precipitation. Analysis of the solid by XRPD showed diffraction peaks for DAM pattern 1. In the case of lyophilization from t-BuOH, a new pattern, designated pattern 2, was isolated. It was found to have converted to pattern 1 after 7 weeks of storage in a closed vial.

[0042] Vacuum drying of the DAM sample at room temperature gave pattern 5, which was found to have converted to pattern 1 after storage in a closed vial for 6 weeks.

[0043] Attempts have been made to crystallize the API (DAM) from a wide range of solvent systems to generate new crystal forms. DAM was used as the input material for these screening studies and the solvents selected were primarily ICH class II and III with a variety of properties as shown in Table 3 below. [Table 3]

[0044] Portions of the DAM crystals (approximately 10 mg each) were treated with the solvent until a solution was formed or 1 mL was added. The resulting samples were either frozen for several days (solutions) or shaken at room temperature (suspensions). Samples that initially formed solutions were examined for signs of solid formation after refrigeration. If solid material formed, it was analyzed by XRPD. If no solid formed, the vial was uncapped, the solution was allowed to evaporate, and the solid residue was analyzed by XRPD. For samples that initially formed suspensions, if solids were still present, they were analyzed by XRPD. The supernatant of the suspension was filtered and the filtrate treated as the solution above. In some experiments, further solids were obtained from evaporation of the filtrate and analyzed by XRPD if the amount of material permitted.

[0045] The results are summarized in Tables 4 and 5 below. [Table 4]

[0046] Most of the experiments using the first 24 solvents yielded solid materials with XRPD diffractograms consistent with Pattern 1. Ethyl formate gave a new pattern (designated Pattern 3), and formic acid gave a syrup after evaporation of the experiment. Further data collected on Pattern 3 suggested that it was a mixture of desodium formate and acetate.

[0047] In runs 25-48, nearly all solvents yielded solid material with XRPD diffractograms consistent with Pattern 1. Three samples remained in solution after six weeks of evaporation. A new pattern, designated Pattern 4, was obtained from 2-methoxyethanol. [Table 5]

[0048] Pattern 2 was obtained from an attempt to prepare amorphous material by lyophilization of a solution of DAM in t-butanol to obtain DAM (Lot: ED21356-005-001-01). Characterization data is summarized in Table 6 below. [Table 6]

[0049] Pattern 3 was obtained from a polymorph screening by evaporation of a solution of DAM in ethyl formate. 1 H NMR analysis shows a reduction in the acetate peak and the addition of a formate peak, consistent with a mixture of desodium formate and acetate. The characterization data is summarized in Table 7 below. [Table 7]

[0050] Pattern 4 was obtained from a polymorph screening by evaporation of a solution of DAM in 2-methoxyethanol. Samples reanalyzed by XRPD after storage in closed vials for 6 weeks show conversion to pattern 1. Characterization data is summarized in Table 8 below. [Table 8]

[0051] Pattern 5 was observed by storing DAM (Pattern 1) at 0% RH, drying under vacuum at room temperature, or heating to 120°C and rapid cooling in the DSC. After 4 hours under ambient laboratory conditions on an XRPD disc, Pattern 5 converted to Pattern 1. Samples left overnight under ambient laboratory conditions on an XRPD disc were also found to convert to Pattern 1. Samples stored for 6 weeks in a sealed vial also converted to Pattern 1. A summary of the characterization data for the Pattern 5 samples is shown in Table 9. [Table 9]

[0052] DAM was highly crystalline and had an XRPD pattern designated Pattern 1. During investigations into DAM, four additional XRPD patterns were found. One of these (Pattern 3) was: 1 It was found to be a mixture of denatonium acetate and formate salts based on H NMR and is therefore not a pattern representative of denatonium acetate alone. Pattern 2 was obtained from an attempt to produce amorphous material by freeze-drying a solution of DAM in t-butanol. After 7 weeks of storage in a closed vial, it was observed to convert to pattern 1. Pattern 4 was obtained from a polymorph screening by evaporation of a solution of DAM in 2-methoxyethanol. A sample reanalyzed by XRPD after 6 weeks of storage in a closed vial showed conversion to pattern 1. Pattern 5 was observed by storage of DAM (pattern 1) at 0% RH, drying in vacuum at room temperature, or heating to 120 °C and rapid cooling in a DSC. When left on an XRPD disc under ambient laboratory conditions, it was observed to convert readily to pattern 1 after 4 hours.

Claims

1. Structural formula 1: 【Chemical 1】 1. A denatonium acetate monohydrate salt having the formula:

2. 10. The salt of claim 1, characterized by an X-ray powder diffraction (XRPD) spectrum substantially as shown in Figure 5A or Figure 5B.

3. 10. A pharmaceutical composition comprising a therapeutically effective amount of the salt of claim 1 or 2 in combination with one or more pharmaceutically acceptable carriers.

4. 4. The pharmaceutical composition of claim 3, wherein the one or more pharmaceutically acceptable carriers comprise a biocompatible polymer.

5. 5. The pharmaceutical composition of claim 4, wherein the biocompatible polymer is cellulose.

6. 4. The pharmaceutical composition of claim 3, wherein the one or more pharmaceutically acceptable carriers comprises a saccharide.

7. The pharmaceutical composition of claim 6, wherein the saccharide is a sugar alcohol.

8. 8. The pharmaceutical composition of claim 7, wherein the sugar alcohol is mannitol.

9. 4. The pharmaceutical composition of claim 3, wherein the one or more pharmaceutically acceptable carriers comprises talc.

10. 4. The pharmaceutical composition of claim 3, wherein the one or more pharmaceutically acceptable carriers comprise an organic acid.

11. 11. The pharmaceutical composition of claim 10, wherein the organic acid is acetic acid.

12. 4. The pharmaceutical composition of claim 3, wherein the pharmaceutical composition is formulated for oral administration.

13. 4. The pharmaceutical composition of claim 3, wherein the pharmaceutical composition comprises solid granules.

14. Structural formula 1: 【Chemistry 2】 1. A method for producing denatonium acetate monohydrate salt represented by the formula:

15. 15. A method for producing the salt of claim 14, comprising the steps of: (1) contacting an equivalent amount of DAA (denatonium acetate anhydrous) with methyl isobutyl ketone and water such that the concentration of water is greater than 10% by weight; (2) recovering the resulting solid phase; and (3) removing the solid therefrom.

16. 15. The method according to claim 14, wherein the lower alkyl isobutyl ketone is methyl isobutyl ketone or ethyl isobutyl ketone.

17. 15. The process of claim 14, wherein contacting denatonium acetate anhydrous with a lower alkyl isobutyl ketone and water forms a composition having a water concentration greater than 10% by weight.

18. The method according to any one of claims 14 to 17, wherein the denatonium acetate monohydrate is a monohydrate crystal.

19. 19. The method of claim 18, wherein the monohydrate crystal is characterized by an X-ray powder diffraction (XRPD) spectrum substantially as shown in Figure 5A or Figure 5B.

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