Crystalline forms of pyridazinone TRPC inhibitors

Abstract

The present invention relates to crystalline forms of pyridazinone TRPC inhibitors, in particular to crystalline forms of Compound (100), pharmaceutical compositions comprising the same and methods of treatment using the same. Compounds (100) are inhibitors of TRPC1, TRPC4 and TRPC5 ion channels for use in the treatment of kidney diseases. # imgabs0 #

Description

[0001] This application is a divisional application of an invention patent application with an application date of April 10, 2020, application number 202080042302.9, and name “Crystalline form of pyridazinone TRPC inhibitor”.

[0002] Related Applications

[0003] This application claims priority to international application PCT / CN19 / 81985 filed on April 10, 2019. Technical Field

[0004] The present invention relates to a crystalline polymorph of Compound 100, a pharmaceutical composition comprising the same, and a method for preparing a pharmaceutical composition using the same. Background Art

[0005] Proteinuria is a condition in which excess protein in the blood leaks into the urine. Proteinuria can develop from 30 mg of protein loss in the urine in a 24-hour period (called microalbuminuria) to >300 mg / day (called macroalbuminuria), and then reach a level of 3.5 grams of protein or more in a 24-hour period, or 25 times the normal amount. Proteinuria occurs when the glomeruli of the kidney malfunction, causing fluid to accumulate in the body (edema). Long-term protein leakage has been shown to lead to kidney failure. Nephrotic syndrome (NS) disease accounts for about 12% of common end-stage renal disease cases and costs more than $3 billion per year in the United States. About 5 in every 100,000 children are diagnosed with NS each year, and now 15 in every 100,000 children suffer from this disease. For patients who respond positively to treatment, the recurrence frequency is also extremely high. 90% of children with nephrotic syndrome will respond to treatment, however, it is estimated that 75% will relapse. There is a need for more effective methods of treating kidney diseases such as proteinuria or reducing the risk of their development.

[0006] Mammalian TRP channel proteins form six transmembrane cation-permeable channels, which can be divided into six subfamilies (TRPC, TRPV, TRPM, TRPA, TRPP and TRPML) based on amino acid sequence homology. Recent studies of TRP channels have shown that they are involved in many basic cellular functions and are thought to play an important role in the pathophysiology of many diseases. Many TRPs are expressed in the kidney along different parts of the nephron, and increasing evidence indicates that these channels are involved in inherited as well as acquired renal disorders. TRPC6, TRPM6 and TRPP2 are associated with hereditary focal segmental glomerulosclerosis (FSGS), hypomagnesemia with secondary hypocalcemia (HSH) and polycystic kidney disease (PKD), respectively. TRPC5 is also associated with FSGS and diabetic nephropathy. (J Clin Invest. 2013 Dec 2; 123(12): 5298–5309.10.1172 / JCI71165; Zhou et al., Science 358, 1332–1336 (2017)

[0007] TRPC5 has also been reported to contribute to mechanisms underlying the regulation of innate fear responses (J Neurosci. 2014 Mar 5;34(10):3653–3667).

[0008] Therefore, additional inhibitors of TRPC5 are needed. Summary of the invention

[0009] In some aspects, the present invention relates to a crystalline form of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one selected from the group consisting of:

[0010] Form A, which is an anhydrate and is characterized by X-ray powder diffraction peaks at 2θ angles of 4.43±0.2°, 11.69±0.2°, 17.75±0.2°, and 27.58±0.2°;

[0011] Form H, which is an anhydrate and is characterized by X-ray powder diffraction peaks at 2θ angles of 13.79±0.2°, 23.61±0.2°, and 27.10±0.2°;

[0012] Form E, which is a hydrate and is characterized by X-ray powder diffraction peaks at 2θ angles of 11.71±0.2°, 15.24±0.2°, 24.79±0.2°, and 26.15±0.2°; or

[0013] Form G, which is a hydrate and is characterized by X-ray powder diffraction peaks at 2θ angles of 15.34±0.2°, 24.58±0.2°, and 25.86±0.2°.

[0014] In some aspects, the present invention relates to a crystalline form of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one, the crystalline form being further selected from:

[0015] Form B, which is an anhydrate and is characterized by X-ray powder diffraction peaks at 2θ angles of 4.40±0.2°, 17.48±0.2°, 17.72±0.2°, and 27.49±0.2°; or

[0016] Form C, which is a hydrate and is characterized by X-ray powder diffraction peaks at 2θ angles of 4.42±0.2°, 8.83±0.2°, 13.27±0.2°, and 17.72±0.2°.

[0017] In some aspects, the invention relates to a pharmaceutical composition comprising crystalline Form A.

[0018] In some aspects, the invention relates to a pharmaceutical composition comprising crystalline Form H.

[0019] In some aspects, the invention relates to a pharmaceutical composition comprising crystalline Form E.

[0020] In some aspects, the invention relates to a pharmaceutical composition comprising crystalline Form G.

[0021] In some aspects, the invention relates to a pharmaceutical composition comprising crystalline Form B.

[0022] In some aspects, the invention relates to a pharmaceutical composition comprising crystalline Form C.

[0023] In some aspects, the present invention is directed to a method of preparing a pharmaceutical composition of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one, the method comprising combining a sample of crystalline Form A with a pharmaceutically acceptable carrier.

[0024] In some aspects, the present invention relates to a method for preparing a pharmaceutical composition of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one, the method comprising the following steps:

[0025] a. dissolving crystalline Form A in a solvent to form a solution; and

[0026] b. preparing a pharmaceutical composition from the solution.

[0027] In some aspects, the invention relates to a method of preparing a pharmaceutical composition of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one, the method comprising combining a sample of crystalline Form H with a pharmaceutically acceptable carrier.

[0028] In some aspects, the present invention relates to a method for preparing a pharmaceutical composition of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one, the method comprising the following steps

[0029] a. dissolving the crystalline Form H in a solvent to form a solution; and

[0030] b. preparing a pharmaceutical composition from the solution.

[0031] In some aspects, the invention relates to a method of preparing a pharmaceutical composition of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one, the method comprising combining a sample of crystalline Form E with a pharmaceutically acceptable carrier.

[0032] In some aspects, the present invention relates to a method for preparing a pharmaceutical composition of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one, the method comprising the following steps

[0033] a. dissolving the crystalline Form E in a solvent to form a solution; and

[0034] b. preparing a pharmaceutical composition from the solution.

[0035] In some aspects, the invention relates to a method of preparing a pharmaceutical composition of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one, the method comprising combining a sample of crystalline Form G with a pharmaceutically acceptable carrier.

[0036] In some aspects, the present invention relates to a method for preparing a pharmaceutical composition of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one, the method comprising the following steps

[0037] a. dissolving the crystalline Form G in a solvent to form a solution; and

[0038] b. preparing a pharmaceutical composition from the solution.

[0039] In some aspects, the invention relates to a method of preparing a pharmaceutical composition of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one, the method comprising combining a sample of crystalline Form B with a pharmaceutically acceptable carrier.

[0040] In some aspects, the present invention relates to a method for preparing a pharmaceutical composition of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one, the method comprising the following steps

[0041] a. dissolving the crystalline Form B in a solvent to form a solution; and

[0042] b. preparing a pharmaceutical composition from the solution.

[0043] In some aspects, the invention relates to a method of preparing a pharmaceutical composition of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one, the method comprising combining a sample of crystalline Form C with a pharmaceutically acceptable carrier.

[0044] In some aspects, the present invention relates to a method for preparing a pharmaceutical composition of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one, comprising the following steps: a. dissolving crystalline form C in a solvent to form a solution; and b. preparing a pharmaceutical composition from the solution.

[0045] In some embodiments, the present invention relates to a method of inhibiting one or more of TRPC1, TRPC4 and TRPC5 ion channels, or ion channels comprising a tetrameric combination of any one of TRPC1, TRPC4 and TRPC5 in a subject in need thereof, the method comprising administering to the subject an effective amount of a pharmaceutical composition comprising a crystalline form of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one. In some aspects of these embodiments, the ion channel is a heterotetrameric form comprising a combination of one or more TRPC1 ion channels and one or more TRPC4 and / or TRPC5 ion channels. In some aspects of these embodiments, the ion channel is a heterotetrameric form comprising one or more TRPC4 ion channels and one or more TRPC5 ion channels.

[0046] In some aspects, the invention relates to a method of treating kidney disease or renal disease associated with a disorder or disease, the method comprising administering to a subject in need thereof a therapeutically effective amount of a pharmaceutical composition comprising a crystalline form of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one.

[0047] In some aspects, the present invention relates to a method for preparing crystalline form A, the method comprising the steps of:

[0048] a. dissolving a certain amount of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one in a certain amount of DMSO:ethanol at a ratio of 2:1 (v / v) at room temperature to form a supersaturated solution;

[0049] b. adding to the solution in step a. an amount sufficient to precipitate 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one in a 1:1 (v / v) ratio of ethanol:H2O; and

[0050] c. Isolating the precipitated material from step b. to produce crystalline Form A.

[0051] In some aspects, the present invention relates to a method for preparing crystalline form A, the method comprising the steps of:

[0052] a. dissolving a certain amount of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one in a certain amount of DMSO:ethanol at a ratio of 2:1 (v / v) at room temperature to form a supersaturated solution;

[0053] b. adding to the solution in step a. an amount of ethanol:H2O in a ratio of 1:1 (v / v) and crystalline Form A seeds to precipitate 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one; and

[0054] c. Isolating the precipitated material from step b. to produce crystalline Form A.

[0055] In some aspects, the present invention relates to a method for preparing crystalline Form H, the method comprising the steps of:

[0056] a. 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one was suspended in a 1:1 (v / v) ratio of isopropanol:isopropyl acetate;

[0057] b. heating the suspension of step a. to a temperature between 45°C and 55°C under stirring for at least 24 hours; and

[0058] c. Separating the insoluble material from step b. to produce crystalline Form H.

[0059] In some aspects, the present invention relates to a method for preparing crystalline Form H, the method comprising the steps of:

[0060] a. Dissolve 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one in a 2:1 (v / v) ratio of DMSO:isopropanol at a temperature between 45°C and 55°C;

[0061] b. The solution from step a. is filtered through a 0.45 micron PTFE membrane;

[0062] c. adding to the filtrate from step b. (i) isopropanol in an amount of 30% to 50% of the amount of isopropanol used in step a.; and (ii) crystalline Form H, and stirring at a temperature between 45°C and 55°C for at least 5 minutes;

[0063] d. adding a 1:1 (v / v) ratio of isopropanol:HO to the solution from step c. with stirring over a period of at least 4 hours while maintaining the temperature between 45°C-55°C to produce a suspension of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one, wherein the amount of isopropanol added in steps c. and d. is about the amount of isopropanol added in step a.;

[0064] e. maintaining the suspension from step d. at a temperature between 45°C and 55°C for at least 2 hours without stirring; and

[0065] f. Isolating the precipitated material from step e. to produce crystalline Form H.

[0066] In some aspects, the present invention relates to a method for preparing crystalline Form H, the method comprising the steps of:

[0067] a. Dissolve 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one in DMSO at a temperature between 65°C and 75°C;

[0068] b. The solution from step a. is filtered through a 0.45 micron PTFE membrane;

[0069] c. adding to the filtrate from step b. (i) isopropanol:H2O in a 1:1 (v / v) ratio in an amount of about 10% of the volume of DMSO used in step A; and (ii) crystalline Form H;

[0070] d. adding an additional amount of isopropanol:HO in a 1:1 (v / v) ratio under stirring to the filtrate from step c. over a period of at least 5 hours while maintaining the temperature between 65°C and 75°C to produce a suspension of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one, wherein the total amount of isopropanol added in steps c. and d. is about half the volume of DMSO used in step a.;

[0071] e. The suspension of step d. was cooled to room temperature with stirring and maintained for at least 2 hours;

[0072] f. keeping the suspension of step e. at room temperature without stirring for at least another 45 minutes; and

[0073] g. Isolating the precipitated material from step f. to produce crystalline Form H.

[0074] In some aspects, the invention relates to a method of forming crystalline Form E, the method comprising the steps of:

[0075] a. 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one was suspended in DMF / H2O (1:9, v / v) at room temperature to form a slurry; and

[0076] b. The suspension was dried under vacuum.

[0077] In some aspects, the invention relates to a method of forming crystalline Form G, the method comprising the steps of:

[0078] a. 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one was suspended in a w 0.8 or more in a solvent to form a slurry; and

[0079] b. The suspension was dried under vacuum. BRIEF DESCRIPTION OF THE DRAWINGS

[0080] The drawings are provided for purposes of illustration and not limitation.

[0081] Figure 1A An experimental XRPD pattern of crystalline Form A of Compound 100 is shown.

[0082] Figure 1B Experimental thermogravimetric analysis (TGA) and differential scanning calorimetry (DSC) data of crystalline Form A of Compound 100 are shown.

[0083] Figure 2A An experimental XRPD pattern of crystalline Form H of Compound 100 is shown.

[0084] Figure 2B Shown are experimental differential scanning calorimetry (DSC) data of crystalline Form H of Compound 100 obtained by crystallization in DMSO / IPA / H2O system.

[0085] Figure 2C Shown are experimental TGA and DSC data of crystalline Form H of Compound 100 obtained via a slurry of Compound 100 in IPA / IPAc (1:1, v / v) at 50 °C.

[0086] Figure 3AAn experimental XRPD pattern of crystalline Form E of Compound 100 is shown.

[0087] Figure 3B Experimental TGA and DSC data of crystalline Form E of Compound 100 are shown.

[0088] Figure 4A An experimental XRPD pattern of crystalline Form G of Compound 100 is shown.

[0089] Figure 4B Experimental TGA and DSC data of crystalline Form G of Compound 100 are shown.

[0090] Figure 5A An experimental XRPD pattern of crystalline Form B of Compound 100 is shown.

[0091] Figure 5B Experimental TGA and DSC data of crystalline Form B of Compound 100 are shown.

[0092] Fig. 6A An experimental XRPD pattern of crystalline Form C of Compound 100 is shown.

[0093] Figure 6B Experimental TGA and DSC data of crystalline Form C of Compound 100 are shown. DETAILED DESCRIPTION

[0094] The invention features crystalline polymorphs of Compound 100, Compound 100 is an inhibitor of TRPC1, TRPC4 and TRPC5 as described in WO20 / 061162, US2020 / 0102301, U.S. Patent Application No. 16 / 575,161 filed on September 18, 2019, No. 62 / 732,728 filed on September 18, 2018, and No. 62 / 780,553 filed on December 17, 201; all of which are incorporated herein by reference.

[0095] The crystalline form of Compound 100 can be used to modulate / improve the physicochemical properties of the compound, including but not limited to solid state properties (e.g., crystallinity, hygroscopicity, melting point or hydration), pharmaceutical properties (e.g., solubility / dissolution rate, stability or compatibility), and crystal characteristics (e.g., purity, yield or morphology).

[0096] In one aspect, the invention features a crystalline Form A of Compound 100 having an X-ray powder diffraction (XRPD) pattern substantially similar to Figure 1A characteristic peaks.

[0097] In another aspect, the invention features a crystalline Form A of Compound 100 having characteristic peaks at 2θ values ​​(°2θ) in an X-ray powder diffraction (XRPD) pattern as shown in Table 1.

[0098] Table 1, Table 2, Table 3 and Table 4 and Figure 1A , Figure 2A , Figure 3A and Figure 4A The relative intensity of each peak in the XRPD data and the 2θ values ​​may change or shift under certain conditions, even though the crystalline form is the same. A person of ordinary skill in the art should be able to easily determine whether a given crystalline form is the same as the crystalline form by comparing its XRPD data. Figure 1A , Figure 2A , Figure 3A and Figure 4A or the same crystalline form as described in one of Table 1, Table 2, Table 3, and Table 4. As used herein, an XRPD data set is "substantially similar" to another XRPD data set if one or more of the peaks in one data set is within ±0.2° 2θ of the corresponding peak in the other data set.

[0099] In yet another aspect, the invention features a crystalline Form A of Compound 100 having characteristic peaks in an X-ray powder diffraction (XRPD) pattern at 2θ values ​​(°2θ) of 4.43±0.2°, 11.69±0.2°, 17.75±0.2°, and 27.58±0.2°. In some embodiments, the indicated characteristic peak is the highest peak in the XRPD pattern.

[0100] In yet another aspect, the invention features a crystalline Form A of Compound 100 having an X-ray powder diffraction (XRPD) pattern at 4.43±0.2°, 8.80±0.2°, 9.17±0.2°, 11.69±0.2°, 12.27±0.2°, 13.28±0.2°, 14.24±0.2°, 14.67±0.2°, 15.96±0. There are characteristic peaks at 2θ values ​​(°2θ) of 2°, 16.93±0.2°, 17.75±0.2°, 19.64±0.2°, 20.98±0.2°, 22.23±0.2°, 22.71±0.2°, 24.19±0.2°, 25.55±0.2°, 27.58±0.2°, 27.95±0.2° and 30.32±0.2°.

[0101] In yet another aspect, the invention features a crystalline Form A of Compound 100 characterized by a differential scanning calorimetry pattern having an onset between 237±2° C. and 256±2° C. In some embodiments, the invention features a crystalline Form A of Compound 100 characterized by a differential scanning calorimetry pattern having onsets at 236.4±1° C., 243.5±1° C., and 256.3±1° C.

[0102] In yet another aspect, the invention features a crystalline Form H of Compound 100 having an X-ray powder diffraction (XRPD) pattern substantially similar to Figure 2A characteristic peaks.

[0103] In yet another aspect, the invention features a crystalline Form H of Compound 100 having characteristic peaks at 2θ values ​​(°2θ) in an X-ray powder diffraction (XRPD) pattern as shown in Table 2.

[0104] In yet another aspect, the invention features a crystalline Form H of Compound 100 having characteristic peaks in an X-ray powder diffraction (XRPD) pattern at 2θ values ​​(°2θ) of 13.79±0.2°, 23.61±0.2°, and 27.10±0.2°. In some embodiments, the indicated characteristic peak is the highest peak in the XRPD pattern.

[0105] In yet another aspect, the invention features a crystalline Form H of Compound 100 having characteristic peaks in an X-ray powder diffraction (XRPD) pattern at 2θ values ​​(°2θ) of 13.79±0.2°, 23.61±0.2°, 27.10±0.2°, and 27.49±0.2°. In some embodiments, the indicated characteristic peak is the highest peak in the XRPD pattern.

[0106] In yet another aspect, the invention features a crystalline Form H of Compound 100 having an X-ray powder diffraction (XRPD) pattern at 4.59±0.2°, 11.92±0.2°, 12.27±0.2°, 13.47±0.2°, 13.79±0.2°, 14.82±0.2°, 15.27±0.2°, 15.89±0.2°, 16.28±0. There are characteristic peaks at 2θ values ​​(°2θ) of 2°, 18.08±0.2°, 19.24±0.2°, 20.77±0.2°, 23.61±0.2°, 24.47±0.2°, 25.11±0.2°, 26.13±0.2°, 27.10±0.2°, 27.49±0.2°, 28.42±0.2° and 30.49±0.2°.

[0107] In yet another aspect, the invention features a crystalline Form H of Compound 100 characterized by a differential scanning calorimetry pattern having an onset at 258° C.±2° C. More specifically, the invention features a crystalline Form H of Compound 100 characterized by a differential scanning calorimetry pattern having onsets at 74.1° C.±1° C., 241.4° C.±1° C., and 257.0° C.±1° C.

[0108] In yet another aspect, the invention features a crystalline Form E of Compound 100 having an X-ray powder diffraction (XRPD) pattern substantially similar to Figure 3A characteristic peaks.

[0109] In yet another aspect, the invention features a crystalline Form E of Compound 100 having characteristic peaks at 2θ values ​​(°2θ) in an X-ray powder diffraction (XRPD) pattern as shown in Table 3.

[0110] In yet another aspect, the invention features a crystalline Form E of Compound 100 having characteristic peaks in an X-ray powder diffraction (XRPD) pattern at 2θ values ​​(°2θ) of 11.71±0.2°, 15.24±0.2°, 24.79±0.2°, and 26.15±0.2°. In some embodiments, the indicated characteristic peak is the highest peak in the XRPD pattern.

[0111] In yet another aspect, the invention features a crystalline Form E of compound 100, having characteristic peaks in an X-ray powder diffraction (XRPD) pattern at 2θ values ​​(°2θ) of 11.71±0.2°, 14.39±0.2°, 15.24±0.2°, 15.63±0.2°, 17.02±0.2°, 17.84±0.2°, 18.30±0.2°, 19.56±0.2°, 20.22±0.2°, 20.65±0.2°, 23.96±0.2°, 24.79±0.2°, and 26.15±0.2°.

[0112] In yet another aspect, the invention features crystalline Form E of Compound 100, characterized by a differential scanning calorimetry pattern having onsets at 78.5°C ± 2°C and 256.7°C ± 2°C. In some embodiments, there is an additional onset at 257.9°C ± 2°C.

[0113] In yet another aspect, the invention features a crystalline Form G of Compound 100 having an X-ray powder diffraction (XRPD) pattern substantially similar to Figure 4A characteristic peaks.

[0114] In yet another aspect, the invention features a crystalline Form G of Compound 100 having characteristic peaks at 2θ values ​​(°2θ) in an X-ray powder diffraction (XRPD) pattern as shown in Table 4.

[0115] In yet another aspect, the invention features a crystalline Form G of Compound 100 having characteristic peaks in an X-ray powder diffraction (XRPD) pattern at 2θ (°2θ) values ​​of 15.34±0.2°, 24.58±0.2°, and 25.86±0.2°. In some embodiments, the indicated characteristic peak is the highest peak in the XRPD pattern.

[0116] In yet another aspect, the invention features a crystalline Form G of Compound 100 having an X-ray powder diffraction (XRPD) pattern at 7.88±0.2°, 11.82±0.2°, 12.85±0.2°, 14.39±0.2°, 14.96±0.2°, 15.34±0.2°, 15.81±0.2°, 16.70±0.2°, 17.40±0. There are characteristic peaks at 2θ values ​​(°2θ) of 2°, 19.51±0.2°, 19.72±0.2°, 20.17±0.2°, 20.63±0.2°, 23.18±0.2°, 23.90±0.2°, 24.58±0.2°, 25.33±0.2°, 25.86±0.2°, 26.26±0.2° and 28.61±0.2°.

[0117] In yet another aspect, the invention features crystalline Form G of Compound 100, characterized by a differential scanning calorimetry pattern having onsets at 80.5°C ± 2°C and 257.2°C ± 2°C. In some embodiments, there is an additional onset at 258.3°C ± 2°C.

[0118] In yet another aspect, the invention features a crystalline Form B of Compound 100 having an X-ray powder diffraction (XRPD) pattern substantially similar to Figure 5A characteristic peaks.

[0119] In yet another aspect, the invention features a crystalline Form B of Compound 100 having a characteristic peak at a 2θ value (° 2θ) in an X-ray powder diffraction (XRPD) pattern as shown in Table 5

[0120] In yet another aspect, the invention features a crystalline Form B of Compound 100 having characteristic peaks in an X-ray powder diffraction (XRPD) pattern at 2θ values ​​(°2θ) of 4.40±0.2°, 17.48±0.2°, 17.72±0.2°, 18.46±0.2°, and 27.49±0.2°. In some embodiments, the indicated characteristic peak is the highest peak in the XRPD pattern.

[0121] In yet another aspect, the invention features a crystalline Form B of Compound 100 having an X-ray powder diffraction (XRPD) pattern at 4.40 ± 0.2°, 8.74 ± 0.2°, 9.13 ± 0.2°, 11.67 ± 0.2°, 12.51 ± 0.2°, 13.10 ± 0.2°, 13.64 ± 0.2°, 14.03 ± 0.2°, 16.26 ± 0.2°, It has characteristic peaks at 2θ values ​​(°2θ) of 16.93±0.2°, 17.48±0.2°, 17.72±0.2°, 18.46±0.2°, 20.51±0.2°, 21.89±0.2°, 23.97±0.2°, 24.79±0.2°, 27.49±0.2°, 27.86±0.2° and 31.76±0.2°.

[0122] In yet another aspect, the invention features crystalline Form B of Compound 100 characterized by a differential scanning calorimetry pattern having an onset at 254.5°±2° C. In some embodiments, there is an additional onset at 256.3±2° C.

[0123] In yet another aspect, the invention features a crystalline Form C of Compound 100 having an X-ray powder diffraction (XRPD) pattern substantially similar to Fig. 6A characteristic peaks.

[0124] In yet another aspect, the invention features a crystalline Form C of Compound 100 having characteristic peaks at 2θ values ​​(°2θ) in an X-ray powder diffraction (XRPD) pattern as shown in Table 6.

[0125] In yet another aspect, the invention features a crystalline Form C of Compound 100 having characteristic peaks in an X-ray powder diffraction (XRPD) pattern at 2θ values ​​(°2θ) of 4.42±0.2°, 8.83±0.2°, 13.27±0.2°, and 17.72±0.2°. In some embodiments, the indicated characteristic peak is the highest peak in the XRPD pattern.

[0126] In yet another aspect, the invention features a crystalline Form C of compound 100 having characteristic peaks in an X-ray powder diffraction (XRPD) pattern at 2θ values ​​(°2θ) of 4.42±0.2°, 8.83±0.2°, 10.52±0.2°, 13.27±0.2°, 16.58±0.2°, 16.97±0.2°, 17.72±0.2°, 21.18±0.2°, 22.21±0.2°, and 24.49±0.2°.

[0127] In yet another aspect, the invention features crystalline Form C of Compound 100, characterized by a differential scanning calorimetry pattern having onsets at 49.6°C ± 2°C and 257.9°C ± 2°C. In some embodiments, there is an additional onset at 258.9°C ± 2°C.

[0128] As used herein, XRPD data can be collected using a D8ADVANCE X-ray diffractometer (Bruker) equipped with a LynxEye detector. In the XRPD analysis, the sample was scanned from 3 ° to 40 ° (2θ) with a step time of 0.3 s. The tube voltage and current were 40 KV and 40 mA, respectively. The XRPD peak position measurement error is typically ± 0.2 degrees 2θ (° 2θ). Alternatively, PANalytical Empyrean and X'Pert3 X-ray powder diffractometers can be used to collect XRPD data using the following parameters:

[0129]

[0130]

[0131] As used herein, differential scanning calorimetry (DSC) data can be collected using Discovery DSC 250 (TAInstruments, US). A weighed sample is placed in a DSC pinhole pan and the weight is accurately recorded. The sample is heated to the final temperature at 10°C / min. As used herein, a DSC data set is "substantially similar" to another DSC data set if one or more of the features in one data set are within ±3°C of the corresponding features in the other data set.

[0132] As used herein, thermogravimetric analysis (TGA) data can be collected using a Discovery TGA 55 (TA Instruments, US). The sample can be placed in an open tarred aluminum pan, automatically weighed, and inserted into a TGA furnace. The sample can be heated to the final temperature at 10°C / min.

[0133] Alternatively, TGA and DSC data can be collected using a TA Q500 / Q5000 / 5500 TGA from TA Instruments and a TA Q200 / Q2000 / 2500 DSC from TA Instruments, respectively, using the following parameters:

[0134] parameter TGA DSC method Slope Slope Sample tray Aluminum, Open Aluminum, curled temperature RT-350℃ 25-270℃ Heating rate 10℃ / min 10℃ / min Purge gas <![CDATA[N2]]> <![CDATA[N2]]>

[0135] In another aspect, the present invention features any one of the embodiments of the above-mentioned crystalline form, and it is substantially pure. As used herein, the term "substantially pure", when used to refer to a given crystalline form, refers to a crystalline form that is at least about 90% pure. This means that the crystalline form does not contain more than about 10% of any other form of compound 100. More preferably, the term "substantially pure" refers to a crystalline form of compound 100 that is at least about 95% pure. This means that the crystalline form of compound 100 does not contain more than about 5% of any other form of compound 100. Even more preferably, the term "substantially pure" refers to a crystalline form of compound 100 that is at least about 97% pure. This means that the crystalline form of compound 100 does not contain more than about 3% of any other form of compound 100. As used herein, the term "about" is defined as close to what is understood by a person of ordinary skill in the art. In a non-limiting embodiment, when used with respect to the amount or volume of a reagent or solvent, the term "about" is defined as within 10%, preferably within 5%, more preferably within 1%, and most preferably within 0.5%.

[0136] In another aspect, the invention features methods of using the crystalline forms of the invention to prepare compositions, such as pharmaceutical compositions, comprising Compound 100.

[0137] The compositions and methods of the present invention can be used to treat subjects in need. In certain embodiments, the subject is a mammal, such as a human or non-human mammal. When administered to a subject (e.g., a human), the composition or compound is preferably administered as a pharmaceutical composition comprising, for example, a compound of the present invention and a pharmaceutically acceptable carrier. Pharmaceutically acceptable carriers are well known in the art and include, for example, aqueous solutions, such as water or physiologically buffered saline; or other solvents or vehicles, such as glycols, glycerol, oils such as olive oil; or injectable organic esters. In a preferred embodiment, when such pharmaceutical compositions are used for human administration, particularly for invasive administration routes (i.e., routes of transport or diffusion through epithelial barriers, such as injection or implantation), aqueous solutions are pyrogen-free or substantially pyrogen-free. Excipients may be selected, for example, to achieve delayed release of the agent. The pharmaceutical composition may be in dosage unit form, such as tablets, capsules (including spray capsules and gelatin capsules), granules, lyophilized agents for reconstitution, powders, solutions or syrups. In some aspects, the composition is a tablet or capsule.

[0138] A pharmaceutically acceptable carrier may contain a physiologically acceptable agent, which acts, for example, to stabilize, increase the solubility of, or increase the absorption of a compound such as a compound of the invention. Such physiologically acceptable agents include, for example, carbohydrates such as glucose, sucrose, or dextran; antioxidants such as ascorbic acid or glutathione; chelating agents; low molecular weight proteins or other stabilizers or excipients. The choice of a pharmaceutically acceptable carrier, including a physiologically acceptable agent, depends, for example, on the route of administration of the composition.

[0139] As used herein, the phrase "pharmaceutically acceptable" refers to those compounds, materials, compositions, and / or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of subjects without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit / risk ratio.

[0140] As used herein, the phrase "pharmaceutically acceptable carrier" means a pharmaceutically acceptable material, composition, or vehicle, such as a liquid or solid filler, diluent, excipient, solvent, or encapsulating material. Each carrier must be "acceptable" in the sense of being compatible with the other ingredients of the formulation and not harmful to the subject. Some examples of materials that can be used as pharmaceutically acceptable carriers include: (1) sugars such as lactose, glucose, and sucrose; (2) starches such as corn starch and potato starch; (3) cellulose and its derivatives such as sodium carboxymethylcellulose, ethyl cellulose, and cellulose acetate; (4) powdered tragacanth; (5) malt; (6) gelatin; (7) talc; (8) excipients such as cocoa butter and suppository waxes; (9) oils such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn starch, and cellulose acetate. oil and soybean oil; (10) glycols such as propylene glycol; (11) polyols such as glycerol, sorbitol, mannitol and polyethylene glycol; (12) esters such as ethyl oleate and ethyl laurate; (13) agar; (14) buffers such as magnesium hydroxide and aluminum hydroxide; (15) alginic acid; (16) pyrogen-free water; (17) isotonic saline; (18) Ringer's solution; (19) ethanol; (20) phosphate buffered saline; and (21) other nontoxic compatible substances used in pharmaceutical preparations.

[0141] The pharmaceutical composition (preparation) of the present invention is preferably administered orally to a subject.

[0142] The preparation can be conveniently present in unit dosage form and can be prepared by any method well known in the pharmaceutical field. The amount of active ingredient that can be combined with a carrier material to produce a single dosage form will vary according to the subject being treated, the specific mode of administration. The amount of active ingredient that can be combined with a carrier material to produce a single dosage form will generally be the amount of the compound that produces the therapeutic effect. Typically, out of 100%, the amount ranges from about 0.5% to about 99% of the active ingredient, preferably from about 0.75% to about 40%, and most preferably from about 0.8% to about 12.5%.

[0143] The method of preparing these preparations or compositions comprises the step of bringing into association the crystalline form of the invention with a carrier and optionally one or more accessory ingredients. In general, the preparations are prepared by uniformly and intimately bringing into association a compound of the invention with liquid carriers or finely divided solid carriers or both, and then, if necessary, shaping the product.

[0144] To prepare solid dosage forms for oral administration (capsules (including spray capsules and gelatin capsules), tablets, pills, dragees, powders, granules, etc.), the active ingredient is mixed with one or more pharmaceutically acceptable carriers (such as sodium citrate or dicalcium phosphate) or solvents and / or any of the following: (1) fillers or extenders, such as starch, lactose, sucrose, glucose, mannitol, microcrystalline cellulose and / or silicic acid; (2) binders, for example, carboxymethyl cellulose, alginates, gelatin, polyvinyl pyrrolidone, sucrose and / or acacia; (3) wetting agents, such as glycerol; (4) disintegrants, such as agar-agar, Calcium carbonate, potato starch or tapioca starch, alginic acid, certain silicates, sodium carbonate and cross-linked carboxymethyl cellulose salts; (5) solution retardants such as paraffin; (6) absorption promoters such as quaternary ammonium compounds; (7) wetting agents, for example, cetyl alcohol and glyceryl monostearate; (8) absorbents such as kaolin and bentonite; (9) lubricants such as talc, calcium stearate, magnesium stearate, solid polyethylene glycol, sodium lauryl sulfate, sodium stearyl fumarate and mixtures thereof; (10) complexing agents such as modified and unmodified cyclodextrins; (11) coloring agents; (12) glidants such as colloidal silicon dioxide; and (12) hydrophilic polymers. In the case of capsules (including spray capsules and gelatin capsules), tablets and pills, the pharmaceutical composition may also contain a buffer. Solid compositions of a similar type may also be used as fillers in soft and hard-filled gelatin capsules using excipients such as lactose or milk sugar and high molecular weight polyethylene glycols and the like.

[0145] Tablets can be prepared by compression or molding, optionally with one or more auxiliary ingredients. Compressed tablets can be prepared using a binder (e.g., gelatin or hydroxypropyl methylcellulose), a lubricant, an inert diluent, a preservative, a disintegrant (e.g., sodium starch glycolate or cross-linked sodium carboxymethyl cellulose), a surfactant or a dispersant. Molded tablets can be prepared by molding a mixture of powdered compounds moistened with an inert liquid diluent in a suitable machine.

[0146] The selected dosage level will depend upon a variety of factors including the activity of the specific compound or combination of compounds employed, or their esters, salts or amides, the route of administration, the time of administration, the rate of excretion of the specific compound employed, the duration of the treatment, other drugs, compounds and / or materials used in combination with the specific compound employed, the age, sex, weight, condition, general health and prior medical history of the subject being treated, and like factors well known in the medical arts.

[0147] A physician or veterinarian with ordinary skills in the art can easily determine and prescribe the therapeutically effective amount of the desired pharmaceutical composition. For example, a physician or veterinarian can start the dosage of the pharmaceutical composition or compound at a level lower than that required to achieve the desired therapeutic effect, and gradually increase the dosage until the desired effect is achieved. "Therapeutically effective amount" refers to the concentration of the compound sufficient to cause the desired therapeutic effect. It is generally understood that the effective amount of the compound will vary according to the subject's weight, sex, age and medical history. Other factors affecting the effective amount may include, but are not limited to, the severity of the subject's condition, the condition being treated, the stability of the compound, and, if necessary, another type of therapeutic agent administered with the compound of the present invention. Larger total doses can be delivered by multiple administrations of the agent. Methods for determining efficacy and dosage are known to those skilled in the art (Isselbacher et al., (1996) Harrison's Principles of Internal Medicine, 13th edition, 1814-1882, incorporated herein by reference).

[0148] In general, a suitable daily dose of an active compound for use in the compositions and methods of the invention will be that amount of the compound which is the lowest dose effective to produce a therapeutic effect. Such an effective dose will generally depend upon the factors described above.

[0149] If desired, the effective daily dose of the active compound may be administered as one, two, three, four, five, six or more sub-doses divided at appropriate intervals throughout the day, optionally in unit dosage form. In certain embodiments of the invention, the active compound may be administered twice or three times a day. In a preferred embodiment, the active compound will be administered once a day.

[0150] In certain embodiments, the compounds of the present invention can be used alone or in combination with another type of therapeutic agent. As used herein, the phrase "combined administration" refers to any form of administration of two or more different therapeutic compounds, so that the second compound is administered when the previously administered therapeutic compound is still effective in vivo (for example, the two compounds are effective in the subject at the same time, which may include the synergistic effect of the two compounds). For example, different therapeutic compounds can be administered simultaneously or continuously with the same preparation or with a separate preparation. In certain embodiments, different therapeutic compounds can be administered within one hour, 12 hours, 24 hours, 36 hours, 48 ​​hours, 72 hours or a week from each other. Therefore, the subject receiving such treatment can benefit from the combined effect of different therapeutic compounds.

[0151] In certain embodiments, the combined administration of a compound of the invention and one or more additional therapeutic agents provides improved efficacy relative to each separate administration of a compound of the invention or the one or more additional therapeutic agents. In certain such embodiments, the combined administration provides an additive effect, wherein the additive effect refers to the sum of each effect of the separate administration of a compound of the invention and the one or more additional therapeutic agents.

[0152] In certain embodiments, the method for forming a pharmaceutical composition of the present invention comprises: a. dissolving a crystalline form of the present invention (such as crystalline form A, crystalline form H, crystalline form E, crystalline form G, crystalline form B or crystalline form C) in a solvent to form a solution; and b. preparing a pharmaceutical composition from the solution. In some aspects, preparing a pharmaceutical composition from a solution comprises spray drying the solution and formulating the spray-dried solution into a solid dosage form. In some embodiments, the present invention relates to a pharmaceutical composition prepared by the method. In some embodiments, the crystalline form is form A. In some embodiments, the crystalline form is form H. In some embodiments, the crystalline form is form E. In some embodiments, the crystalline form is form G. In some embodiments, the crystalline form is form B. In some embodiments, the crystalline form is form C.

[0153] Any crystalline form described herein, including any crystalline form described in any aspect, embodiment or example of the present application, can be used in any method of the invention described herein.

[0154] Treatment

[0155] Non-selective Ca 2+-Permeable transient receptor potential (TRP) channels act as sensors to transduce extracellular cues to the intracellular environment in different cellular processes including actin remodeling and cell migration (Greka et al., Nat Neurosci 6, 837-845, 2003; Ramsey et al., Annu Rev Physiol 68, 619-647, 2006; Montell, Pflugers Arch 451, 19-28, 2005; Clapham, Nature 426, 517-524, 2003). The dynamic rearrangement of the actin cytoskeleton depends on the spatiotemporally regulated Ca 2+ Influx (Zheng and Poo, Annu Rev Cell Dev Biol 23, 375-404, 2007); Brandman and Meyer, Science 322, 390-395, 2008); Collins and Meyer, Dev Cell 16, 160-161, 2009) and the small GTPases RhoA and Rac1 serve as key regulators of these changes (Etienne-Manneville and Hall, Nature 420, 629-635, 2002); Raftopoulou and Hall, Dev Biol 265, 23-32, 2004). RhoA induces the formation of stress fibers and adhesion plaques, while Rac1 mediates the formation of lamellipodia (Etienne-Manneville and Hall, Nature 420, 629-635, 2002). Transient receptor potential cation channel subfamily C member 5 (TRPC5) works in concert with TRPC6 to regulate Ca2+ influx, actin remodeling, and cell motility in renal podocytes and fibroblasts. TRPC5-mediated Ca 2+ Influx increases Rac1 activity, whereas TRPC6-mediated Ca2+ influx promotes RhoA activity. Genetic silencing of TRPC6 channels abolishes stress fibers and reduces focal contacts, presenting a motile, migratory cell phenotype. In contrast, genetic silencing of TRPC5 channels rescues stress fiber formation, presenting a contracted cell phenotype. The results described here reveal a conserved signaling mechanism whereby TRPC5 and TRPC6 channels control the balance of tightly regulated cytoskeletal dynamics through differential coupling with Rac1 and RhoA.

[0156] Ca 2+-dependent remodeling of the actin cytoskeleton is a dynamic process that drives cell migration (Wei et al., Nature 457, 901-905, 2009). RhoA and Racl act as switches responsible for cytoskeletal rearrangement in migrating cells (Etienne-Manneville and Hall, Nature 420, 629-635, 2002); Raftopoulou and Hall, Dev Biol 265, 23-32, 2004). Activation of Rac1 mediates a motile cell phenotype, while RhoA activity promotes a contractile phenotype (Etienne-Manneville and Hall, Nature 420, 629-635, 2002). 2+ It plays an important role in the regulation of small GTPases (Aspenstrom et al., Biochem J 377, 327-337, 2004). 2+ Spatially and temporally restricted flashes of Ca2+ are enriched near the leading edge of migrating cells (Wei et al., Nature 457, 901-905, 2009). Therefore, Ca2+ microdomains engage local bursts of Rac1 activity (Gardiner et al., Curr Biol 12, 2029-2034, 2002; Machacek et al., Nature 461, 99-103, 2009) as key events at the leading edge. To date, the source of Ca2+ influx responsible for GTPase regulation remains largely elusive. TRP (transient receptor potential) channels generate temporally and spatially restricted Ca2+ influxes associated with cell migration in fibroblasts and neuronal growth cones. 2+ Signal 0. Specifically, TRPC5 channels are known regulators of neuronal growth cone guidance1, and their activity in neurons depends on the activity of PI3K and Rac1 (Bezzerides et al., Nat Cell Biol 6, 709-720, 2004).

[0157] Podocytes are neuron-like cells that originate from the metanephric mesenchyme of the renal glomeruli and are essential for the formation of the renal filter (Somlo and Mundel, Nat Genet. 24, 333-335, 2000; Fukasawa et al., J Am Soc Nephrol 20, 1491-1503, 2009). Podocytes display a series of extremely precise cytoskeletal adaptations to environmental cues (Somlo and Mundel, Nat Genet 24, 333-335, 2000; Garg et al., Mol Cell Biol 27, 8698-8712, 2007; Verma et al., J Clin Invest 116, 1346-1359, 2006; Verma et al., J Biol Chem 278, 20716-20723, 2003; Barletta et al., J Biol Chem 278, 19266-19271, 2003; Holzman et al., Kidney Int 56, 1481-1491, 1999; Ahola et al., Am J Pathol 155, 907-913, 1999; Tryggvason and Wartiovaara, N Engl J Med 354, 1387-1401, 2006; Schnabel and Farquhar, J Cell Biol 111, 1255-1263, 1990; Kurihara et al., Proc Natl Acad Sci USA 89, 7075-7079, 1992). Early events in podocyte injury are characterized by dysregulation of the actin cytoskeleton (Faul et al., Trends Cell Biol 17, 428-437, 2007; Takeda et al., J Clin Invest 108, 289-301, 2001; Asanuma et al., Nat Cell Biol 8, 485-491, 2006) and Ca2+ homeostasis (Hunt et al., J Am Soc Nephrol 16, 1593-1602, 2005; Faul et al., Nat Med 14, 931-938, 2008). These changes are associated with the onset of proteinuria, loss of albumin into the renal cystic space, and eventual renal failure (Tryggvason and Wartiovaara, N Engl J Med 354, 1387-1401, 2006). The vasoactive hormone angiotensin II induces Ca2+ in podocytes. 2+Influx of Ca2+ and long-term treatment leads to stress fiber loss (Hsu et al., J Mol Med 86, 1379-1394, 2008). Although there is a recognized association between Ca2+ influx and cytoskeletal reorganization, the mechanism by which podocytes sense and transduce extracellular cues that regulate cell shape and activity remains elusive. Mutations in the TRP canonical 6 (TRPC6) channel have been associated with podocyte injury (Winn et al., Science 308, 1801-1804, 2005; Reiser et al., Nat Genet 37, 739-744, 2005; Moller et al., J Am Soc Nephrol 18, 29-36, 2007; Hsu et al., Biochim Biophys Acta 1772, 928-936, 2007), but little is known about the specific pathways that regulate this process. In addition, TRPC6 shares close homology with the other six members of the TRPC channel family (Ramsey et al., Annu Rev Physiol 68, 619-647, 2006; Clapham, Nature 426, 517-524, 2003). TRPC5 channels have an antagonistic effect on TRPC6 channel activity to control the tightly regulated cytoskeletal dynamics balance through differential coupling with different small GTPases.

[0158] Proteinuria

[0159] Proteinuria is a pathological condition in which protein is present in urine. Albuminuria is a type of proteinuria. Microalbuminuria occurs when the kidneys leak a small amount of albumin into the urine. In a normally functioning body, albumin is usually absent from the urine because it is held in the bloodstream by the kidneys. Microalbuminuria can be diagnosed from 24-hour urine collections (20 μg / min to 200 μg / min) or more commonly from concentrations (30 mg / L to 300 mg / L) that rise at at least two moments. Microalbuminuria may be a precursor to diabetic nephropathy. Albumin levels exceeding these values ​​are referred to as macroalbuminuria. Subjects suffering from certain conditions such as diabetic nephropathy can develop from microalbuminuria to macroalbuminuria, and reach the nephrotic range (>3.5 g / 24 hours) when kidney disease reaches the late stage.

[0160] Causes of proteinuria

[0161] Proteinuria may be associated with a variety of diseases, including focal segmental glomerulosclerosis, IgA nephropathy, diabetic nephropathy, lupus nephritis, membranoproliferative glomerulonephritis, progressive (crescentic) glomerulonephritis, and membranous glomerulonephritis.

[0162] A. Focal segmental glomerulosclerosis (FSGS)

[0163] Focal segmental glomerulosclerosis (FSGS) is a disease that attacks the kidney's filtration system (glomeruli), causing severe scarring. FSGS is one of many causes of a condition called nephrotic syndrome, which occurs when protein in the blood leaks into the urine (proteinuria).

[0164] There are few treatments available for patients with FSGS. Many patients are treated with steroid regimens, most of which have very severe side effects. Some patients have been shown to respond positively to immunosuppressive drugs, as well as blood pressure medications that have been shown to reduce protein levels in the urine. To date, there is no generally accepted effective treatment or cure, and there are no FDA-approved drugs for the treatment of FSGS. Therefore, more effective methods of reducing or inhibiting proteinuria are needed.

[0165] B. IgA nephropathy

[0166] IgA nephropathy (also known as IgA nephritis, IgAN, Bergey's disease and glomerulonephritis) is a form of glomerulonephritis (inflammation of the glomeruli of the kidney). IgA nephropathy is the most common glomerulonephritis worldwide. Primary IgA nephropathy is characterized by the deposition of IgA antibodies in the glomeruli. There are other diseases associated with glomerular IgA deposition, the most common of which is Henlein-Schönlein purpura (HSP), which is considered by many to be a systemic form of IgA nephropathy. Henlein-Schönlein purpura presents with characteristic purpuric rash, arthritis and abdominal pain and occurs more often in young people (16-35 years old). HSP is associated with a better prognosis than IgA nephropathy. In IgA nephropathy, 25%-30% of cases slowly progress to chronic renal failure over a period of 20 years.

[0167] C. Diabetic nephropathy

[0168] Diabetic nephropathy, also known as Kimmelstiel-Wilson syndrome and intercapillary glomerulonephritis, is a progressive kidney disease caused by vascular changes in the capillaries in the glomeruli of the kidney. It is characterized by nephrotic syndrome and diffuse glomerulosclerosis. It is caused by long-standing diabetes and is the main reason for dialysis. The earliest detectable change in the course of diabetic nephropathy is thickening of the glomeruli. At this stage, the kidneys may begin to allow higher than normal levels of serum albumin in the urine. As diabetic nephropathy progresses, more and more glomeruli are destroyed by nodular glomerulosclerosis and the amount of albumin excreted in the urine increases.

[0169] D. Lupus nephritis

[0170] Lupus nephritis is a kidney disorder that is a complication of systemic lupus erythematosus. Lupus nephritis occurs when antibodies and complement build up in the kidneys, causing inflammation. It often causes proteinuria and can quickly progress to kidney failure. Nitrogen waste products build up in the blood. Systemic lupus erythematosus causes various diseases of the internal structures of the kidneys, including interstitial nephritis. About 3 in every 10,000 people are affected by lupus nephritis.

[0171] E. Membranoproliferative glomerulonephritis I / II / III

[0172] Membranoproliferative glomerulonephritis is a type of glomerulonephritis caused by deposits in the mesangium of the kidney and thickening of the basement membrane, activated complement, and damage to the glomeruli. There are three types of membranoproliferative glomerulonephritis. Type I is caused by the deposition of immune complexes in the kidney and is thought to be related to the classical complement pathway. Type II is similar to type I but is believed to be related to the alternative complement pathway. Type III is very rare and is characterized by subepithelial deposits mixed with the typical pathological findings of type I disease.

[0173] F. Progressive (crescentic) glomerulonephritis

[0174] Progressive (crescentic) glomerulonephritis (PG) is a renal syndrome that, if left untreated, rapidly progresses to acute renal failure and death within a few months. In 50% of cases, PG is associated with an underlying disease such as Goodpasture's syndrome, systemic lupus erythematosus, or Wegener granulomatosis; the remaining cases are idiopathic. Regardless of the underlying cause, PG involves severe damage to the glomeruli of the kidney, many of which contain characteristic crescent-shaped scars. Patients with PG have hematuria, proteinuria, and occasionally hypertension and edema. Although the degree of proteinuria may occasionally exceed 3 g / 24 hours, a range associated with nephrotic syndrome, the clinical picture is consistent with renal syndrome. Untreated disease may progress to a decrease in urine volume (oliguria), which is associated with poor renal function.

[0175] G. Membranous glomerulonephritis

[0176] Membranous glomerulonephritis (MGN) is a slowly progressive kidney disease that primarily affects patients aged 30 to 50 years, usually Caucasians. It can progress to nephrotic syndrome. MGN is caused by circulating immune complexes. Current studies have shown that most immune complexes are formed by antibodies binding to antigens in situ on the glomerular basement membrane. The antigens can be endogenous to the basement membrane or deposited from the systemic circulation.

[0177] H.Alport syndrome

[0178] Alport syndrome is a genetic disorder that affects 1 in 5,000-10,000 children and is characterized by glomerulonephritis, end-stage renal disease, and hearing loss. Alport syndrome can also affect the eyes, but these changes do not usually affect vision unless changes to the lens occur later in life. Blood in the urine is common. Proteinuria is characteristic of progressive kidney disease.

[0179] I. Hypertensive kidney disease

[0180] Hypertensive kidney disease (hypertensive nephrosclerosis (HN or HNS) or hypertensive nephropathy (HN)) is a medical condition involving damage to the kidneys caused by chronic high blood pressure. HN can be divided into two types: benign and malignant. Benign nephrosclerosis is common in individuals over the age of 60, while malignant nephrosclerosis is uncommon and affects 1%-5% of individuals with hypertension, who have a diastolic blood pressure of more than 130 mm Hg. Signs and symptoms of chronic kidney disease may occur, including loss of appetite, nausea, vomiting, itching, drowsiness or confusion, weight loss, and bad taste in the mouth. Chronic high blood pressure causes damage to the tissue of the kidneys; this includes the small blood vessels, glomeruli, tubules, and interstitial tissue. The tissue becomes hardened and thickened, which is called nephrosclerosis. The narrowing of the blood vessels means that less blood enters the tissue, and therefore less oxygen reaches the tissue, which leads to tissue death (local ischemia).

[0181] J. Nephrotic syndrome

[0182] Nephrotic syndrome is a series of symptoms caused by kidney damage. This includes protein in the urine, low blood albumin levels, high blood fats, and significant swelling. Other symptoms may include weight gain, feeling tired, and foamy urine. Complications may include blood clotting, infection, and high blood pressure. Causes include many kidney diseases, such as focal segmental glomerulosclerosis, membranous nephropathy, and minimal change disease. It may also occur as a complication of diabetes or lupus. The underlying mechanism usually involves damage to the glomeruli of the kidney. Diagnosis is usually based on urine tests and sometimes on kidney biopsies. It differs from nephritic syndrome in that there are no red blood cells in the urine. Nephrotic syndrome is characterized by heavy proteinuria (>3.5g / 1.73m2 body surface area / day, or children>40mg / square meter body surface area / hour), hypoalbuminemia (<2,5g / dl), hyperlipidemia, and edema starting from the face. Lipiduria (lipids in the urine) may also occur, but is not necessary for the diagnosis of nephrotic syndrome. Hyponatremia can also occur with a low sodium excretion rate. Inherited forms of nephrotic syndrome are often resistant to steroids and other immunosuppressive treatments. The goals of therapy are to control urinary protein loss and swelling, provide good nutrition to allow the child to grow, and prevent complications. Early and aggressive treatment is used to control the condition.

[0183] K. Minimal change disease

[0184] Minimal change disease (also known as MCD, minimal change glomerulopathy, and no disease, among others) is a disease that affects the kidneys, causing nephrotic syndrome. The clinical signs of minimal change disease are proteinuria (abnormal excretion of protein, primarily albumin, into the urine), edema (soft tissue swelling due to water retention), weight gain, and hypoalbuminemia (low serum albumin). These signs are collectively referred to as nephrotic syndrome. The first clinical sign of minimal change disease is usually edema, with associated weight gain. The swelling can be mild, but patients may present with edema of the lower half of the body, periorbital edema, swelling of the scrotum / labial region, and, in more severe cases, generalized edema. In the elderly, patients may also develop acute kidney injury (20%-25% of affected adults) and hypertension. Due to the disease process, patients with minimal change disease are also at risk for blood clotting and infection.

[0185] L. Membranous nephropathy

[0186] Membranous nephropathy is the deposition of immune complexes on the glomerular basement membrane (GBM), with GBM thickening. The cause is usually unknown (idiopathic), but secondary causes include drugs, infection, autoimmune disease, and cancer. Manifestations include insidious onset of edema and heavy proteinuria with benign urine sediment, normal renal function, and normal or elevated blood pressure. Membranous nephropathy is diagnosed by renal biopsy. Spontaneous remission is common. Treatment of patients at high risk of progression is usually with corticosteroids and cyclophosphamide or chlorambucil.

[0187] M. Postinfectious glomerulonephritis

[0188] Acute proliferative glomerulonephritis is a disorder of the glomeruli (glomerulonephritis) or small blood vessels in the kidneys. It is a common complication of bacterial infection, typically a skin infection caused by type 12, 4 and 1 streptococcal bacteria (impetigo), but also occurs after streptococcal pharyngitis, so it is also called post-infection or post-streptococcal glomerulonephritis. It may be a risk factor for future albuminuria. In adults, signs and symptoms of infection may still be present when kidney problems occur, and terms such as infection-related glomerulonephritis or bacterial infection-related glomerulonephritis are also used. Acute glomerulonephritis caused 19,000 deaths in 2013, down from 24,000 deaths worldwide in 1990. Acute proliferative glomerulonephritis (post-streptococcal glomerulonephritis) is caused by a streptococcal bacterial infection, usually three weeks after infection, usually of the pharynx or skin, given the time required to produce antibodies and complement proteins. The infection causes inflammation of the blood vessels in the kidneys, which impedes the ability of the kidney organ to filter urine. [citation needed] Acute proliferative glomerulonephritis most commonly occurs in children.

[0189] N. Thin basement membrane disease

[0190] Thin basement membrane disease (TBMD, also called benign familial hematuria and thin basement membrane nephropathy or TBMN), along with IgA nephropathy, is the most common cause of hematuria without other symptoms. The only abnormal finding in this disease is thinning of the glomerular basement membrane in the kidney. Its importance lies in the fact that it has a benign prognosis and patients maintain normal kidney function throughout their lives. Most patients with thin basement membrane disease have microscopic hematuria found incidentally on urinalysis. Blood pressure, renal function, and urinary protein excretion are usually normal. Mild proteinuria (less than 1.5 g / day) and hypertension may be seen in a few patients. Frank hematuria and flank pain should prompt a search for other causes, such as kidney stones or flank pain-hematuria syndrome. In addition, there are no systemic manifestations, so the presence of hearing impairment or visual impairment should prompt a search for hereditary nephritis, such as Alport syndrome. Some TBMD patients are thought to be carriers of the gene that causes Alport syndrome.

[0191] O. Mesangial proliferative glomerulonephritis

[0192] Mesangial proliferative glomerulonephritis is a type of glomerulonephritis that is primarily associated with the mesangium. There is some evidence that interleukin-10 may inhibit it in animal models. [2] The World Health Organization (WHO) classifies it as type II lupus nephritis. Mesangial cells in the glomeruli take up and degrade circulating immunoglobulins by endocytosis. This normal process stimulates mesangial cell proliferation and matrix deposition. Therefore, during periods of elevated circulating immunoglobulins (i.e., lupus and IgA nephropathy), one would expect to see an increase in the number of mesangial cells and matrix in the glomeruli. This is characteristic of the nephritic syndrome.

[0193] P. Amyloidosis (primary)

[0194] Amyloidosis is a group of diseases in which abnormal proteins called amyloid fibrils accumulate in tissues. [4] Symptoms depend on the type and are often variable. [2] They can include diarrhea, weight loss, feeling tired, a swollen tongue, bleeding, numbness, feeling weak when standing, leg swelling, or an enlarged spleen. [2] There are about 30 different types of amyloidosis, each caused by a specific protein misfolding. [5] Some are inherited, while others are acquired. [3] They are divided into localized and systemic forms. [2] The four most common systemic types are light chain (AL), inflammatory (AA), dialysis (Aβ2M), and inherited age-related (ATTR). Primary amyloidosis refers to amyloidosis in which no associated clinical condition is identified.

[0195] Q.c1q nephropathy

[0196] C1q nephropathy is a rare glomerular disease with characteristic mesangial C1q deposits observed on immunofluorescence microscopy. It is histologically defined and poorly understood. Light microscopic features are heterogeneous and include minimal change disease (MCD), focal segmental glomerulosclerosis (FSGS), and proliferative glomerulonephritis. Clinical presentation is also varied, ranging from asymptomatic hematuria or proteinuria in children and adults to Frank nephritis or nephrotic syndrome. Hypertension and renal insufficiency at diagnosis are common findings. Optimal treatment is unclear and is often guided by the underlying light microscopic lesions. Corticosteroids are the mainstay of treatment, with immunosuppressants reserved for steroid-resistant cases. The presence of nephrotic syndrome and FSGS appears to predict a poor outcome, in contrast to the favorable outcome seen in patients with MCD. (Devasahayam et al., “C1qNephropathy: The Unique Underrecognized Pathological Entity,” Analytical Cellular Pathology, volume 2015, article ID 490413, page 5, 2015. https: / / doi.org / 10.1155 / 2015 / 490413.)

[0197] R. Anti-GBM disease

[0198] Anti-glomerular basement membrane (GBM) disease, also known as Goodpasture's disease, is a rare condition that causes inflammation of small blood vessels in the kidneys and lungs. Anti-glomerular basement membrane (GBM) antibodies primarily attack the kidneys and lungs, but systemic symptoms such as malaise, weight loss, fatigue, fever and chills are also common, as are joint aches and pains. 60% to 80% of patients experience both lung and kidney involvement; 20%-40% affect only the kidneys, and less than 10% affect only the lungs. Pulmonary symptoms usually precede kidney symptoms and often include: coughing up blood, chest pain (less than 50% of total cases), cough and shortness of breath. Kidney symptoms often include blood in the urine, protein in the urine, unexplained swelling of the limbs or face, high urea levels in the blood, and high blood pressure. GPS causes abnormal production of anti-GBM antibodies by the plasma cells of the blood. Anti-GBM antibodies attack the alveoli and glomerular basement membranes. These antibodies bind their reactive epitopes to the basement membrane and activate the complement cascade, leading to marked cell death. T cells are also involved. It is generally considered a type II hypersensitivity reaction.

[0199] Measurement of urine protein levels

[0200] Protein levels in urine can be measured using methods known in the art. Until recently, accurate protein measurement required a 24-hour urine collection. During the 24-hour collection, the patient urinates into a container that is kept refrigerated between bathroom visits. The patient is instructed to begin collecting urine after the first bathroom visit in the morning. Every drop of urine for the rest of the day should be collected in the container. The next morning, the patient adds the first urination after waking up and the collection is complete.

[0201] Recently, researchers have discovered that a single urine sample can provide the needed information. In the newer technique, the amount of albumin in the urine sample is compared to the amount of creatinine, a waste product of normal muscle breakdown. The measurement is called the urine albumin-to-creatinine ratio (UACR). A urine sample that contains more than 30 milligrams of albumin per gram of creatinine (30 mg / g) warns that there may be a problem. If the lab test is over 30 mg / g, another UACR test should be done 1 to 2 weeks later. If the second test also shows high levels of protein, the person has persistent proteinuria, a sign of decreased kidney function, and additional tests to evaluate kidney function should be done.

[0202] A test that measures the amount of creatinine in the blood will also show whether the subject's kidneys are effectively removing waste. Too much creatinine in the blood is a sign that a person has kidney damage. Doctors can use creatinine measurements to estimate how effectively the kidneys filter the blood. This calculation is called the estimated glomerular filtration rate, or eGFR. Chronic kidney disease is present when the eGFR is less than 60 milliliters per minute (mL / min).

[0203] TRPC5

[0204] TRPC is a family of transient receptor potential cation channels in animals. TRPC5 is a subtype of the mammalian transient receptor potential ion channel TRPC family. Three examples of TRPC5 are people (Gene Bank Accession Numbers NM_012471.2 and NP_036603.1; Gene ID 7224); mice (Gene Bank Accession Numbers NM_009428.2 and NP_033454.1; Gene ID 22067); and rats (Gene Bank Accession Numbers NM_080898.2 and NP_543174.1; Gene ID 140933).

[0205] TRPC1

[0206] TRPC1 is an ion channel located on the plasma membrane of many human and animal cell types. It is a nonspecific cation channel, meaning that both sodium and calcium ions can pass through it. TRPC1 is thought to mediate calcium entry in response to depletion of endosomal calcium stores or activation of receptors coupled to the phospholipase C system. In HEK293 cells, the overall current-voltage relationship of endogenous TRPC1 channels is nearly linear, with a slope conductance of approximately 17 pS. The extrapolated reversal potential of TRPC1 channels is +30 mV. TRPC1 protein is widely expressed throughout the mammalian brain and has a corticolimbic expression pattern similar to that of TRPC4 and TRPC5. The highest density of TRPC1 protein is found in the lateral septum, an area with dense expression of TRPC4, and in the hippocampus and prefrontal cortex, areas with dense expression of TRPC5.

[0207] TRPC4

[0208] TRPC4 is a member of the transient receptor potential cation channels. This protein forms a non-selective calcium-permeable cation channel activated by Gαi-coupled receptors, Gαq-coupled receptors, and tyrosine kinases, and plays a role in a variety of processes including endothelial permeability, vasodilation, neurotransmitter release, and cell proliferation. The non-selective cation channel TrpC4 has been shown to be present in high abundance in the corticolimbic region of the brain. In addition, TRPC4 mRNA is present in the ventral tegmental area and substantia nigra of midbrain dopaminergic neurons. Deletion of the trpc4 gene reduces the level of sociability in a social exploration task. These results suggest that TRPC4 may play a role in regulating social anxiety in many different disorders. TRPC4 has been shown to interact with TRPC1 and TRPC5.

[0209] Therefore, the present invention provides a method of inhibiting one or more of TRPC1, TRPC4 and TRPC5 ion channels of a subject in need, or an ion channel comprising a tetramer combination of any one of TRPC1, TRPC4 and TRPC5, the method comprising administering to the subject an effective amount of a pharmaceutical composition of compound 100 as described herein. "Ion channels comprising a tetramer combination of any one of TRPC1, TRPC4 and TRPC5" ion channels may contain any combination of TRPC1, TRPC4 and TRPC5 ion channels. In some embodiments, the ion channel to be inhibited is a heterotetrameric form comprising a combination of one or more TRPC1 ion channels and one or more TRPC4 and / or TRPC5 ion channels. In some embodiments, the ion channel to be inhibited is a heterotetrameric form comprising a combination of one or more TRPC1 ion channels and one or more TRPC4 and / or TRPC5 ion channels. In some embodiments, the ion channel to be inhibited is a heterotetrameric form comprising one or more TRPC4 ion channels and one or more TRPC5 ion channels. The heterotetrameric form may comprise any combination of TRPC1, TRPC4 and TRPC5 ion channels. In some embodiments, the heterotetrameric form is TRPC1:TRPC4:TRPC4:TRPC5, TRPC1:TRPC1:TRPC5:TRPC5, TRPC4:TRPC4:TRPC5:TRPC5 or TRPC4:TRPC5:TRPC5:TRPC5:TRPC5.

[0210] In some embodiments, the subject in need of inhibiting an ion channel comprising a tetramer combination of any one of TRPC1, TRPC4, and TRPC5 suffers from kidney disease, nephropathy associated with a disease or disorder, pain, anxiety, or depression. In some embodiments, the pain is selected from neuropathic pain and visceral pain. In embodiments, the cancer is selected from chemoresistant breast cancer, adriamycin-resistant breast cancer, chemoresistant colorectal cancer, medulloblastoma, and tumor angiogenesis.

[0211] In certain embodiments, the present invention provides a method for treating a disease or condition selected from kidney disease, kidney disease associated with the disease or condition, pain, anxiety, or depression, or reducing the risk of developing the severity thereof, the method comprising administering to a subject in need thereof a therapeutically effective amount of a pharmaceutical composition comprising a crystalline form of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one described herein.

[0212] In some embodiments, the disease or disorder is a kidney disease or kidney disease associated with a disorder or disease selected from the group consisting of focal segmental glomerulosclerosis (FSGS), diabetic nephropathy, Alport syndrome, hypertensive kidney disease, nephrotic syndrome, steroid-resistant nephrotic syndrome, minimal change disease, membranous nephropathy, idiopathic membranous nephropathy, membranoproliferative glomerulonephritis (MPGN), immune complex-mediated MPGN, complement-mediated MPGN, lupus nephritis, post-infectious glomerulonephritis, thin basement membrane disease, mesangial proliferative glomerulonephritis, amyloidosis (primary), clq nephropathy, rapidly progressive GN, anti-GBM disease, C3 glomerulonephritis, hypertensive nephrosclerosis, IgA nephropathy, IgG4 nephropathy, disease, proteinuric kidney disease, microalbuminuria, macroalbuminuric kidney disease, transplant-associated FSGS, transplant-associated nephrotic syndrome, transplant-associated proteinuria, nodular glomerulonephritis, NASR disease (proliferative glomerulonephritis with monoclonal IgG deposits), polycystic kidney disease, autosomal dominant polycystic kidney disease (ADPKD), autosomal recessive polycystic kidney disease (ARPKD), or kidney disease associated with any of obesity, insulin resistance, type II diabetes, prediabetes, metabolic syndrome, dyslipidemia, Fabry disease, pulmonary hypertension, cholestatic liver disease, nonalcoholic fatty liver disease (NAFLD), nonalcoholic steatohepatitis (NASH), or cancer.

[0213] In some embodiments, the kidney disease or kidney disease associated with the disorder or disease is focal segmental glomerulosclerosis (FSGS), diabetic nephropathy, Alport syndrome, hypertensive kidney disease, obesity-related kidney disease, nephrotic syndrome, steroid-resistant nephrotic syndrome, minimal change disease, membranous nephropathy, membranoproliferative glomerulonephritis (MPGN), lupus nephritis, post-infectious glomerulonephritis, thin basement membrane disease, mesangial proliferative glomerulonephritis, amyloidosis (primary), clq nephropathy, anti-GBM disease, C3 glomerulonephritis, hypertensive nephrosclerosis, IgA nephropathy, IgG4 nephropathy, dyslipidemia associated with chronic kidney disease, nodular glomerulonephritis, NASR disease (proliferative glomerulonephritis with monoclonal IgG deposits), polycystic kidney disease, kidney disease associated with Fabry disease, or kidney disease associated with metabolic syndrome.

[0214] In some embodiments, the kidney disease is a proteinuric kidney disease. In some embodiments, the kidney disease is a microalbuminuric or macroalbuminuric kidney disease.

[0215] In some embodiments, the disease or condition to be treated is renal disease associated with pulmonary hypertension.

[0216] In some embodiments, the disease or condition to be treated is pain selected from neuropathic pain and visceral pain.

[0217] In some embodiments, the disease or disorder is kidney disease associated with a cancer selected from chemoresistant breast cancer, doxorubicin-resistant breast cancer, chemoresistant colorectal cancer, medulloblastoma, and tumor angiogenesis.

[0218] The present invention also provides a method for treating anxiety, depression or cancer or reducing the risk of developing the same, which comprises administering to a subject in need thereof a therapeutically effective amount of a compound of the present invention (eg, a compound of Formula I) or a pharmaceutical composition comprising the compound.

[0219] In some embodiments, the disease or condition to be treated is transplant-related FSGS, transplant-related nephrotic syndrome, transplant-related proteinuria, cholestatic liver disease, polycystic kidney disease, autosomal dominant polycystic kidney disease (ADPKD), autosomal recessive polycystic kidney disease (ARPKD), obesity, insulin resistance, type II diabetes, prediabetes, metabolic syndrome, non-alcoholic fatty liver disease (NAFLD), or non-alcoholic steatohepatitis (NASH).

[0220] In some embodiments, the kidney disease or renal disease associated with the condition or disease to be treated is hypertensive nephropathy, nephropathy associated with metabolic syndrome, nephropathy associated with obesity, nephropathy associated with dyslipidemia, diabetic nephropathy, nephrotic syndrome, FSGS, or minimal change disease.

[0221] In some embodiments, the kidney disease or renal disease associated with the condition or disease to be treated is diabetic nephropathy, FSGS, or minimal change disease.

[0222] The present invention also provides a method for treating anxiety, depression or cancer or reducing the risk of developing the same, which comprises administering a therapeutically effective amount of compound 100 or a pharmaceutical composition comprising the compound to a subject in need thereof.

[0223] The subject to be treated by the method of the present invention is a subject who has been diagnosed with any of the above diseases or symptoms, or is at risk of developing any of the above diseases or symptoms. The method is effective for a variety of subjects, including mammals, such as humans and other animals, such as laboratory animals, such as mice, rats, rabbits or monkeys, or domesticated and farm animals, such as cats, dogs, goats, sheep, pigs, cows or horses. In some embodiments, the subject is a mammal. In some embodiments, the subject is a human.

[0224] Example

[0225] Abbreviations

[0226] <![CDATA[a w ]]> Water activity EtOH Ethanol <![CDATA[H2O]]> water DMSO Dimethyl sulfoxide XRPD Powder X-ray diffraction DSC Differential Scanning Calorimetry TGA Thermogravimetric analysis IPA Isopropyl alcohol

[0227] Example 1. Preparation of Crystalline Form A

[0228] Method A

[0229] In a 100 mL flask equipped with a magnetic stirrer, 1.0 g of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one was suspended in 15 mL of DMSO:EtOH (2:1). The slurry was stirred at RT for 0.5 h to completely dissolve. 15 mL of EtOH:HO (1:1) was gradually added over a period of 2.0 hours. The resulting suspension was filtered and washed with 15 mL of EtOH:HO (1:1) and dried under vacuum (-0.1 MPa) at 50 °C for 3.5 hours to give 0.83 g of a white crystalline solid characterized as Form A.

[0230] Method B

[0231] In a 100 mL flask equipped with a magnetic stirrer, 1.0 g of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one was suspended in 15 mL of DMSO:EtOH (2:1). The slurry was stirred at RT for 0.5 h to completely dissolve. 0.3 mL of EtOH:HO (1:1) was added, followed by 0.03 g of Form A seeds. The resulting mixture was stirred at room temperature for 1.5 hours. Another 15 mL of EtOH:HO (1:1) was then slowly added at room temperature (0.5 mL / h for 1 h, then 1 mL / h for 1 h, then 2 mL / h for 1 h, and the rest was 5 mL / h) while maintaining stirring. The resulting suspension was filtered and washed with 15 mL of EtOH:H2O (1:1) and dried under vacuum (-0.1 MPa) at 50°C for 3.5 hours to afford 0.7 g of a white crystalline solid characterized as Form A.

[0232] The powder X-ray diffraction pattern of the crystalline form thus prepared and the XRPD peaks with relative intensities are shown in Figure 1A and Table 1. The DSC and TGA data are shown in Figure 1B middle.

[0233] Table 1. XRPD peak list of Form A

[0234]

[0235] Example 2. Preparation of Compound 100 Crystalline Form H

[0236] Method A

[0237] In an HPLC vial equipped with a stirring bar, 20 mg of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazine-3(2H)-one was suspended in 0.5 mL of an IPA / IPAc (1:1, v / v) mixture. The resulting slurry was magnetically stirred (~800 rpm) at 50 °C for about 3 days while keeping the vial closed. The slurry was cooled at room temperature, the solid was separated by centrifugation (10000 rpm, 2 min), and dried at RT for 24 hours to give a white crystalline solid.

[0238] Method B

[0239] In a 20 mL glass vial, 1.0 g of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one was dissolved in a mixture containing 10 mL DMSO and 5 mL IPA by stirring at 50° C. for one hour. The resulting solution was filtered through a 0.45 μM PTFE membrane to obtain a clear solution and placed in a 100 mL flask. 2 mL of IPA was added, followed by a small amount of Form H seeds (obtained using Method A), followed by an additional 103.3 mg of Form H seeds (obtained using Method A). After stirring at 50° C. for 10 minutes, 6 mL of IPA / H2O (1:1, v / v) was added over a period of 6 hours while maintaining the temperature at 50° C. Stirring was stopped and the suspension was maintained at 50°C for another 1.5 hours, then cooled at 25°C and maintained at that temperature for 3 hours. The slurry was then filtered and the resulting wet cake was dried under vacuum for 12 hours. 0.77 g of Form H (white crystalline solid) was obtained with a yield of about 67%.

[0240] Method C

[0241] In a 20 mL glass vial, 1.5 g of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one was dissolved in 10 mL DMSO by stirring at 70° C. for one hour. The resulting solution was filtered through a 0.45 μM PTFE membrane to obtain a clear solution and placed in a 100 mL flask equipped with an overhead stirrer and stirred at 70° C. at 300 rpm. 0.9 mL of IPA / HO (1:1, v / v) mixture was added, followed by a small amount of Form H seeds, followed by an additional 75.3 mg of Form H and 9.1 mL of IPA / HO (1:1, v / v) over 6 hours. After stirring at 70° C. for one hour after the IPA / H2O addition was complete, the suspension was cooled to 25° C. while maintaining stirring for 2 hours, and then maintained without stirring for another hour. The suspension was filtered, and the resulting wet cake was rinsed with 20 mL of IPA and then dried in a vacuum oven at 50° C. for 12 hours. 1.28 g of Form H (white crystalline solid) was obtained, with a yield of about 80%.

[0242] The powder X-ray diffraction pattern of the crystalline form thus prepared and the XRPD peaks with relative intensities are shown in Figure 2A and Table 2. The DSC data are shown in Figure 2B middle.

[0243] Table 2. XRPD peak list of Compound 100 Crystalline Form H

[0244]

[0245] Example 3. Preparation of Compound 100 Crystalline Form E

[0246] In an HPLC vial, about 15 mg of compound 100 was suspended in 0.5 mL of DMF:H2O (1:9, v / v). After the suspension was stirred at room temperature for 3 days, the remaining solid was then dried overnight in vacuo at room temperature. Alternatively, about 15 mg of compound 100 was suspended in 0.5 mL of DMSO:H2O (1:9, v / v) or DMF:H2O (1:1, v / v) to produce crystalline form E. Another approach to produce crystalline form E is by crystalline form C as described in Example 7.

[0247] The powder X-ray diffraction pattern of the crystalline form thus prepared and the XRPD peaks with relative intensities are shown in Figure 3A and Table 3. The DSC and TGA data are shown in Figure 3B middle.

[0248] Table 3. XRPD peak list of Compound 100 Crystalline Form E

[0249]

[0250]

[0251] The stability of crystalline Form E at different relative humidities over a period of 3-14 days was also tested.

[0252] The results are shown in Table 3.1 below

[0253] Table 3.1 Stability of crystalline form E at different relative humidity (RH)

[0254]

[0255] The appearance of Form F after storage of Form E at 30% relative humidity for 14 days indicated that Form E was not stable at such low relative humidity.

[0256] Example 4. Preparation of Compound 100 Crystalline Form G

[0257] Crystalline Form G can be obtained by reacting compound 100 in a w For example, Compound 100 was slurried in acetonitrile / water (1:1, v / v) at room temperature to obtain Form G.

[0258] The powder X-ray diffraction pattern of the crystalline form thus prepared and the XRPD peaks with relative intensities are shown in Figure 4A and Table 4. The DSC and TGA data are shown in Figure 4B middle.

[0259] Table 4. XRPD peak list of Compound 100 Crystalline Form G

[0260]

[0261]

[0262] Example 5. Preparation of Compound 100 Crystalline Form B

[0263] Crystalline Form B was obtained by heating compound 100 to 244° C. and then cooling to room temperature under N 2 protection.

[0264] Alternatively, crystalline Form B was obtained by suspending about 30 mg of Compound 100 in 1.5 mL of a 2:1 (v / v) THF:heptane mixture at room temperature to form a slurry. The resulting wet cake was dried overnight to give crystalline Form B.

[0265] The powder X-ray diffraction pattern of the crystalline form thus prepared and the XRPD peaks with relative intensities are shown in Figure 5A and Table 5. The DSC and TGA data are shown in Figure 5B middle.

[0266] Table 5. XRPD peak list of Compound 100 Crystalline Form B

[0267]

[0268] Example 6. Preparation of Compound 100 Crystalline Form C

[0269] Crystalline Form C was prepared by dissolving Compound 100 in 1,4-dioxane and then adding an anti-solvent, H2O, to the solution to precipitate Crystalline Form C.

[0270] Crystalline Form C was also prepared by the following anti-solvent crystallization procedure. About 600 mg of Compound 100 Crystalline Form A was dissolved in 70 mL of THF at room temperature. 30 mL of heptane was added to the solution to precipitate the material. The insoluble material in the form of a wet cake was separated and dried overnight to obtain Crystalline Form C.

[0271] Once the crystals of Form C are isolated, they can be used as seeds in the following process to prepare additional crystalline Form C. 1000 mg of Compound 100 was suspended in 21 mL of a 2:1 (v / v) THF:H2O mixture at room temperature, to which 3% (w / v) Form C crystals were added. The components were mixed together (mechanically or with a magnetic stirring bar) overnight to form a slurry. The resulting wet cake was dried to produce crystalline Form C.

[0272] The powder X-ray diffraction pattern of the crystalline form thus prepared and the XRPD peaks with relative intensities are shown in Fig. 6A and Table 6. The DSC and TGA data are shown in Figure 6B middle.

[0273] Table 6. XRPD peak list of Compound 100 Crystalline Form C

[0274] °2-θ Relative peak height (%) 4.42 100.0％ 8.83 50.3％ 10.52 16.5％ 13.27 53.6％ 16.58 16.8％ 16.97 24.5％ 17.72 49.6％ 21.18 7.6％ 22.21 24.9％ 24.49 14.1％

[0275] Example 7. Preparation of Crystalline Form E Using Crystalline Form C

[0276] Wet crystalline Form C (e.g., wet cake from any of the above Form C preparations prior to drying) was converted to crystalline Form E after 24 hours of magnetic stirring (using a magnetic stirring bar) without any seeding. Similarly, wet crystalline Form C seeded with 1% crystalline Form E was mostly converted to crystalline Form E by mechanical stirring (using a shaker) for 24-72 hours. However, even after 3 days of stirring, 5% of the material remained Form C by XRPD analysis. Magnetic stirring appears to result in better conversion of Form C to Form E compared to mechanical stirring. Without being bound by theory, we believe that magnetic stirring affects the morphology of the material, thereby making the conversion of Form C to Form E more efficient. It is therefore possible that grinding Form C into smaller sized crystals prior to stirring can enhance the conversion of Form C to Form E.

[0277] The present invention thus relates to the following embodiments:

[0278] Scheme 1. A crystalline form of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one selected from the group consisting of:

[0279] a. Form A, characterized by X-ray powder diffraction peaks at 2θ angles of 4.43±0.2°, 11.69±0.2°, 17.75±0.2° and 27.58±0.2°, wherein the characteristic peaks are the highest peaks observed;

[0280] b. Form E, characterized by X-ray powder diffraction peaks at 2θ angles of 11.71±0.2°, 15.24±0.2°, 24.79±0.2° and 26.15±0.2°, wherein the characteristic peaks are the highest peaks observed;

[0281] c. Form G, characterized by X-ray powder diffraction peaks at 2θ angles of 15.34±0.2°, 24.58±0.2°, and 25.86±0.2°, wherein the characteristic peaks are the highest peaks observed;

[0282] d. Form H, characterized by X-ray powder diffraction peaks at 2θ angles of 13.79±0.2°, 23.61±0.2°, 27.10±0.2°, and 27.49±0.2°, wherein the characteristic peaks are the highest peaks observed;

[0283] e. Form B, characterized by X-ray powder diffraction peaks at 2θ angles of 4.40±0.2°, 17.48±0.2°, 17.72±0.2°, 18.46±0.2° and 27.49±0.2°, wherein the characteristic peaks are the highest peaks observed; and

[0284] f. Form C, characterized by X-ray powder diffraction peaks at 2θ angles of 4.42±0.2°, 8.83±0.2°, 13.27±0.2°, and 17.72±0.2°, wherein the characteristic peaks are the highest peaks observed.

[0285] Scheme 2. Crystalline Form A as described in Scheme 1.

[0286] Scheme 3. The crystalline Form A as described in Scheme 2, characterized by X-ray powder diffraction peaks at 2θ angles of 4.43±0.2°, 8.80±0.2°, 9.17±0.2°, 11.69±0.2°, 12.27±0.2°, 13.28±0.2°, 14.24±0.2°, 14.67±0.2°, 15.96±0.2°, 16.93±0.2°, 17.75±0.2°, 19.64±0.2°, 20.98±0.2°, 22.23±0.2°, 22.71±0.2°, 24.19±0.2°, 25.55±0.2°, 27.58±0.2°, 27.95±0.2° and 30.32±0.2°.

[0287] Scheme 4. The crystalline form A as described in Scheme 2, characterized in that the X-ray powder diffraction pattern is substantially similar to Figure 1A .

[0288] Scheme 5. Crystalline Form A as described in any of Schemes 1-4, is further characterized by a differential scanning calorimetry pattern having an onset between 237±2°C and 256°C±2°C.

[0289] Scheme 6. Crystalline Form A as described in Scheme 5, characterized by a differential scanning calorimetry pattern having onsets at 236.4±1°C, 243.5±1°C, and 256.3°C±1°C.

[0290] Scheme 7. Crystalline Form H as described in Scheme 1.

[0291] Scheme 8. The crystalline Form H as described in Scheme 7 is characterized by X-ray powder diffraction peaks at 2θ angles of 4.59±0.2°, 11.92±0.2°, 12.27±0.2°, 13.47±0.2°, 13.79±0.2°, 14.82±0.2°, 15.27±0.2°, 15.89±0.2°, 16.28±0.2°, 18.08±0.2°, 19.24±0.2°, 20.77±0.2°, 23.61±0.2°, 24.47±0.2°, 25.11±0.2°, 26.13±0.2°, 27.10±0.2°, 27.49±0.2°, 28.42±0.2° and 30.49±0.2°.

[0292] Scheme 9. The crystalline form H as described in Scheme 7, characterized in that the X-ray powder diffraction pattern is substantially similar to Figure 2A .

[0293] Scheme 10. Crystalline Form H as described in any of Schemes 7-9, which is further characterized by a differential scanning calorimetry chart having an onset at 258°C ± 2°C.

[0294] Scheme 11. Crystalline Form H as described in Scheme 10, further characterized by additional differential scanning calorimetry patterns with onsets at 74.1°C±1°C, 241.4°C±1°C, and 257.0°C±1°C.

[0295] Scheme 12. Crystalline Form E as described in Scheme 1.

[0296] Scheme 13. The crystalline form E as described in Scheme 12 is characterized by X-ray powder diffraction peaks at 2θ angles of 11.71±0.2°, 14.39±0.2°, 15.24±0.2°, 15.63±0.2°, 17.02±0.2°, 17.84±0.2°, 18.30±0.2°, 19.56±0.2°, 20.22±0.2°, 20.65±0.2°, 23.96±0.2°, 24.79±0.2° and 26.15±0.2°.

[0297] Scheme 14. The crystalline form E as described in Scheme 12, characterized in that the X-ray powder diffraction pattern is substantially similar to Figure 3A .

[0298] Scheme 15. Crystalline Form E as described in any of Schemes 12-14, is further characterized by a differential scanning calorimetry pattern having onsets at 78.5°±2°C and 256.7°±2°C.

[0299] Scheme 16. Crystalline Form E as described in Scheme 15, further characterized by an additional differential scanning calorimetry pattern with an onset at 257.9°C ± 2°C.

[0300] Scheme 17. Crystalline Form G as described in Scheme 1.

[0301] Scheme 18. The crystalline Form G as described in Scheme 17 is characterized by X-ray powder diffraction peaks at 2θ angles of 7.88±0.2°, 11.82±0.2°, 12.85±0.2°, 14.39±0.2°, 14.96±0.2°, 15.34±0.2°, 15.81±0.2°, 16.70±0.2°, 17.40±0.2°, 19.51±0.2°, 19.72±0.2°, 20.17±0.2°, 20.63±0.2°, 23.18±0.2°, 23.90±0.2°, 24.58±0.2°, 25.33±0.2°, 25.86±0.2°, 26.26±0.2° and 28.61±0.2°.

[0302] Scheme 19. The crystalline form G as described in Scheme 18, characterized in that the X-ray powder diffraction pattern is substantially similar to Figure 4A .

[0303] Scheme 20. The crystalline Form G of any one of Schemes 17-19, further characterized by a differential scanning calorimetry pattern having onsets at 80.5°C±2°C and 257.2°C±2°C.

[0304] Scheme 21. The crystalline Form G as described in Scheme 20, further characterized by an additional differential scanning calorimetry pattern with an onset at 258.3°C ± 2°C.

[0305] Scheme 22. A pharmaceutical composition comprising the crystalline form A according to any one of Schemes 2-6; and a pharmaceutically acceptable carrier.

[0306] Scheme 23. A pharmaceutical composition comprising the crystalline form H according to any one of Schemes 7-11; and a pharmaceutically acceptable carrier.

[0307] Scheme 24. A pharmaceutical composition comprising the crystalline form E according to any one of Schemes 12-16; and a pharmaceutically acceptable carrier.

[0308] Scheme 25. A pharmaceutical composition comprising the crystalline form G according to any one of Schemes 17-21; and a pharmaceutically acceptable carrier.

[0309] Scheme 26. A method of inhibiting one or more of TRPC1, TRPC4 and TRPC5 ion channels, or ion channels comprising a tetrameric combination of any one of TRPC1, TRPC4 and TRPC5 in a subject in need thereof, the method comprising administering to the subject an effective amount of a pharmaceutical composition as described in any one of Schemes 22-25.

[0310] Embodiment 27. A method for treating kidney disease or kidney disease associated with a disease or condition, the method comprising administering to a subject in need thereof a pharmaceutical composition as described in any one of Embodiments 22-25.

[0311] Scheme 28. The method of scheme 27, wherein the kidney disease or the kidney disease associated with a disease or condition is focal segmental glomerulosclerosis (FSGS), diabetic nephropathy, Alport syndrome, hypertensive renal disease, nephrotic syndrome, steroid-resistant nephrotic syndrome, minimal change disease, membranous nephropathy, idiopathic membranous nephropathy, membranoproliferative glomerulonephritis (MPGN), immune complex-mediated MPGN, complement-mediated MPGN, lupus nephritis, post-infectious glomerulonephritis, thin basement membrane disease, mesangial proliferative glomerulonephritis, amyloidosis (primary), c1q nephropathy, rapidly progressive GN, anti-GBM disease, C3 glomerulonephritis, hypertensive nephrosclerosis, IgA nephropathy, IgG4 nephropathy. disease, proteinuric kidney disease, microalbuminuria, macroalbuminuric kidney disease, transplant-associated FSGS, transplant-associated nephrotic syndrome, transplant-associated proteinuria, nodular glomerulonephritis, NASR disease (proliferative glomerulonephritis with monoclonal IgG deposits), polycystic kidney disease, autosomal dominant polycystic kidney disease (ADPKD), autosomal recessive polycystic kidney disease (ARPKD), or kidney disease associated with any of obesity, insulin resistance, type 2 diabetes, prediabetes, metabolic syndrome, dyslipidemia, pulmonary hypertension, cancer, cholestatic liver disease, nonalcoholic fatty liver disease (NAFLD), nonalcoholic steatohepatitis (NASH), or Fabry disease

[0312] Embodiment 29. The method of embodiment 28, wherein the kidney disease or the kidney disease associated with a disease or condition is focal segmental glomerulosclerosis (FSGS), diabetic nephropathy, Alport syndrome, hypertensive kidney disease, obesity-related kidney disease, nephrotic syndrome, steroid-resistant nephrotic syndrome, minimal change disease, membranous nephropathy, membranoproliferative glomerulonephritis (MPGN), lupus nephritis, post-infectious glomerulonephritis, thin basement membrane disease, mesangial proliferative glomerulonephritis, amyloidosis (primary), c1q nephropathy, anti-GBM disease, C3 glomerulonephritis, hypertensive nephrosclerosis, IgA nephropathy, IgG4 nephropathy, dyslipidemia-related kidney disease, nodular glomerulonephritis, NASR disease (proliferative glomerulonephritis with monoclonal IgG deposits), polycystic kidney disease, or renal complications caused by Fabry disease.

[0313] Embodiment 30. The method of embodiment 29, wherein the renal disease is hypertensive nephropathy, nephropathy associated with metabolic syndrome, nephropathy associated with obesity, nephropathy associated with dyslipidemia, diabetic nephropathy, nephrotic syndrome, FSGS or minimal change disease.

[0314] Embodiment 31. The method of embodiment 30, wherein the kidney disease is diabetic nephropathy, FSGS or minimal change disease.

[0315] Embodiment 32. A method for treating pain, anxiety or depression, the method comprising administering to a subject in need thereof the pharmaceutical composition of any one of Embodiments 22-25.

[0316] Scheme 33. A method for preparing the crystalline form A as described in any one of Schemes 2-6, the method comprising the following steps:

[0317] a. dissolving a certain amount of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one in a certain amount of DMSO:ethanol at a ratio of 2:1 (v / v) at room temperature to form a supersaturated solution;

[0318] b. adding to the solution in step a. an amount sufficient to precipitate the 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one in a 1:1 (v / v) ratio of ethanol:H2O; and

[0319] c. Isolating the precipitated material from step b. to produce the crystalline Form A.

[0320] Scheme 34. A method for preparing the crystalline form A as described in any one of Schemes 2-6, the method comprising the following steps:

[0321] a. dissolving a certain amount of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one in a certain amount of DMSO:ethanol at a ratio of 2:1 (v / v) at room temperature to form a supersaturated solution;

[0322] b. adding to the solution in step a. an amount of ethanol:H2O in a ratio of 1:1 (v / v) and crystalline Form A seeds to precipitate 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one; and

[0323] c. Isolating the precipitated material from step b. to produce the crystalline Form A.

[0324] Scheme 35. A method for preparing a pharmaceutical composition of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one, the method comprising combining a sample of crystalline Form A with a pharmaceutically acceptable excipient.

[0325] Scheme 36. A method for preparing a pharmaceutical composition of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one, the method comprising the following steps:

[0326] a. dissolving crystalline Form A in a solvent to form a solution; and

[0327] b. preparing the pharmaceutical composition from the solution.

[0328] Embodiment 37. The method of embodiment 36, wherein preparing the pharmaceutical composition from the solution comprises spray-drying the solution, and formulating the spray-dried solution into a solid dosage form.

[0329] Scheme 38. A pharmaceutical composition prepared by the method as described in any one of Schemes 35-37.

[0330] Scheme 39. A method for preparing the crystalline form H as described in any one of Schemes 7-11, the method comprising the following steps:

[0331] a. 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one was suspended in a 1:1 (v / v) ratio of isopropanol:isopropyl acetate;

[0332] b. heating the suspension of step a. to a temperature between 45°C and 55°C under stirring for at least 24 hours; and

[0333] c. separating the insoluble material from step b. to produce the crystalline Form H.

[0334] Scheme 40. A method for preparing the crystalline form H as described in any one of Schemes 7-11, the method comprising the following steps:

[0335] a. Dissolve 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one in a 2:1 (v / v) ratio of DMSO:isopropanol at a temperature between 45°C and 55°C;

[0336] b. The solution from step a. is filtered through a 0.45 micron PTFE membrane;

[0337] c. adding to the filtrate from step b. (i) an amount of 30%-50% of the amount of isopropanol used in step a.; and (ii) crystalline Form H, and stirring at a temperature between 45°C and 55°C for at least 5 minutes;

[0338] d. adding a 1:1 (v / v) ratio of isopropanol:H2O to the solution from step c. with stirring over a period of at least 4 hours while maintaining the temperature between 45°C-55°C to produce a suspension of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one, wherein the amount of isopropanol added in steps c. and d. is approximately equal to the amount of isopropanol added in step a.;

[0339] e. maintaining the suspension from step d. at a temperature between 45°C and 55°C for at least 2 hours without stirring; and

[0340] f. Isolating the precipitated material from step e. to produce the crystalline Form H.

[0341] Scheme 41. A method for preparing the crystalline form H as described in any one of Schemes 7-11, the method comprising the following steps:

[0342] a. Dissolve 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one in DMSO at a temperature between 65°C and 75°C;

[0343] b. The solution from step a. is filtered through a 0.45 micron PTFE membrane;

[0344] c. adding to the filtrate from step b. (i) isopropanol:H2O in a 1:1 (v / v) ratio in an amount of about 10% of the volume of DMSO used in step A; and (ii) crystalline Form H;

[0345] d. adding an additional amount of isopropanol:HO in a 1:1 (v / v) ratio under stirring to the filtrate from step c. over a period of at least 5 hours while maintaining the temperature between 65°C-75°C to produce a suspension of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one, wherein the total amount of isopropanol added in steps c. and d. is about half of the volume of DMSO used in step a.;

[0346] e. The suspension of step d. is cooled to room temperature under stirring and maintained for at least 2 hours;

[0347] f. keeping the suspension of step e. at room temperature without stirring for at least another 45 minutes; and

[0348] g. Isolating the precipitated material from step f. to produce the crystalline Form H.

[0349] Scheme 42. A method for preparing a pharmaceutical composition of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one, the method comprising combining a sample of crystalline Form H with a pharmaceutically acceptable excipient.

[0350] Scheme 43. A method for preparing a pharmaceutical composition of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one, the method comprising the following steps:

[0351] a. dissolving the crystalline Form H in a solvent to form a solution; and

[0352] b. preparing the pharmaceutical composition from the solution.

[0353] Embodiment 44. The method of embodiment 43, wherein preparing the pharmaceutical composition from the solution comprises spray-drying the solution, and formulating the spray-dried solution into a solid dosage form.

[0354] Scheme 45. A pharmaceutical composition prepared by the method as described in any one of Schemes 42-44.

[0355] Scheme 46. A method for forming the crystalline form E as described in any one of Schemes 12-16, the method comprising the following steps:

[0356] a. 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one was suspended in DMF / H2O (1:9, v / v) at room temperature to form a slurry; and

[0357] b. The suspension was dried under vacuum.

[0358] Scheme 47. A method for preparing a pharmaceutical composition of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one, the method comprising combining a sample of crystalline Form E with a pharmaceutically acceptable excipient.

[0359] Scheme 48. A method for preparing a pharmaceutical composition of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one, the method comprising the following steps:

[0360] a. dissolving the crystalline Form E in a solvent to form a solution; and

[0361] b. preparing the pharmaceutical composition from the solution.

[0362] Embodiment 49. The method of Embodiment 48, wherein preparing the pharmaceutical composition from the solution comprises spray-drying the solution, and formulating the spray-dried solution into a solid dosage form.

[0363] Scheme 50. A pharmaceutical composition prepared by the method as described in any one of Schemes 47-49.

[0364] Scheme 51. A method for forming the crystalline form G as described in any one of Schemes 17-21, the method comprising the following steps:

[0365] a. 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one was suspended in a w 0.8 or more in a solvent to form a slurry; and

[0366] b. The suspension was dried under vacuum.

[0367] Scheme 52. The method of Scheme 50, wherein the solvent is a 1:1 (v / v) mixture of acetonitrile and water.

[0368] Scheme 53. A method for preparing a pharmaceutical composition of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one, the method comprising combining a sample of crystalline Form G with a pharmaceutically acceptable excipient.

[0369] Scheme 54. A method for preparing a pharmaceutical composition of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one, the method comprising the following steps:

[0370] a. dissolving the crystalline Form G in a solvent to form a solution; and

[0371] b. preparing the pharmaceutical composition from the solution.

[0372] Embodiment 55. The method of embodiment 54, wherein preparing the pharmaceutical composition from the solution comprises spray-drying the solution, and formulating the spray-dried solution into a solid dosage form.

[0373] Scheme 56. A pharmaceutical composition prepared by the method as described in any one of Schemes 53-55.

[0374] Scheme 57. A method of inhibiting a heterotetrameric form comprising a combination of one or more TRPC1 ion channels and one or more TRPC4 and / or TRPC5 ion channels in a subject in need thereof, the method comprising administering to the subject an effective amount of a pharmaceutical composition as described in any one of Schemes 22-25, 38, 45, 50 or 56.

[0375] Scheme 58. A method of inhibiting a heterotetrameric form comprising one or more TRPC4 ion channels and one or more TRPC5 ion channels in a subject in need thereof, the method comprising administering to the subject an effective amount of a pharmaceutical composition as described in any one of Schemes 22-25, 38, 45, 50 or 56.

[0376] Scheme 59. A method as described in any of schemes 57-58, wherein the subject has a kidney disease (such as focal segmental glomerulosclerosis (FSGS), diabetic nephropathy, Alport syndrome, hypertensive kidney disease, nephrotic syndrome, steroid-resistant nephrotic syndrome, minimal change disease, membranous nephropathy, idiopathic membranous nephropathy, membranoproliferative glomerulonephritis (MPGN), immune complex-mediated MPGN, complement-mediated MPGN, lupus nephritis, post-infectious glomerulonephritis, thin basement membrane disease, mesangial proliferative glomerulonephritis, amyloidosis (primary), c1q nephropathy, rapidly progressive GN, anti-GBM disease, C3 glomerulonephritis, hypertensive nephrosclerosis, IgA nephropathy, IgG4 nephropathy, proteinuria kidney disease, microalbuminuria, proteinuria, macroalbuminuria kidney disease, transplant-associated FSGS, transplant-associated nephrotic syndrome, transplant-associated proteinuria, nodular glomerulonephritis, NASR disease (proliferative glomerulonephritis with monoclonal IgG deposits), polycystic kidney disease, autosomal dominant polycystic kidney disease (ADPKD), autosomal recessive polycystic kidney disease (ARPKD), or kidney disease associated with any of obesity, insulin resistance, type II diabetes, prediabetes, metabolic syndrome, dyslipidemia, pulmonary hypertension, cancer, cholestatic liver disease, nonalcoholic fatty liver disease (NAFLD), nonalcoholic steatohepatitis (NASH), or Fabry disease), pain (such as neuropathic pain or visceral pain), anxiety, or depression.

[0377] The above description of the present invention provides illustration and description, but is not intended to be exhaustive or limit the present invention to the disclosed precise invention. According to the above teachings, modifications and variations are possible, or can be obtained from the practice of the present invention. Therefore, the scope of the present invention is limited by the scheme and its equivalent.

Claims

1. A crystalline form of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one selected from the group consisting of: a. Form A, characterized in that X-ray powder diffraction peaks at 2θ angles of 4.43±0.2°, 11.69±0.2°, 17.75±0.2°, and 27.58±0.2°, wherein the characteristic peak is the highest peak observed; b. Form E, characterized by X-ray powder diffraction peaks at 2θ angles of 11.71±0.2°, 15.24±0.2°, 24.79±0.2° and 26.15±0.2°, wherein the characteristic peaks are the highest peaks observed; c. Form G, characterized by X-ray powder diffraction peaks at 2θ angles of 15.34±0.2°, 24.58±0.2°, and 25.86±0.2°, wherein the characteristic peaks are the highest peaks observed; d. Form H, characterized by X-ray powder diffraction peaks at 2θ angles of 13.79±0.2°, 23.61±0.2°, 27.10±0.2°, and 27.49±0.2°, wherein the characteristic peaks are the highest peaks observed; e. Form B, characterized by X-ray powder diffraction peaks at 2θ angles of 4.40±0.2°, 17.48±0.2°, 17.72±0.2°, 18.46±0.2° and 27.49±0.2°, wherein the characteristic peaks are the highest peaks observed; and f. Form C, characterized by X-ray powder diffraction peaks at 2θ angles of 4.42±0.2°, 8.83±0.2°, 13.27±0.2°, and 17.72±0.2°, wherein the characteristic peaks are the highest peaks observed.

2. A pharmaceutical composition comprising the crystalline form A according to claim 1; and a pharmaceutically acceptable carrier.

3. A pharmaceutical composition comprising the crystalline form H according to claim 1; and a pharmaceutically acceptable carrier.

4. A pharmaceutical composition comprising the crystalline form E according to claim 1; and a pharmaceutically acceptable carrier.

5. A pharmaceutical composition comprising the crystalline form G according to claim 1; and a pharmaceutically acceptable carrier.

6. A method of inhibiting one or more of TRPC1, TRPC4 and TRPC5 ion channels, or ion channels comprising a tetrameric combination of any one of TRPC1, TRPC4 and TRPC5, in a subject in need thereof, the method comprising administering to the subject an effective amount of a pharmaceutical composition as described in any one of claims 2-5.

7. A method of treating kidney disease or kidney disease associated with a disease or condition, the method comprising administering to a subject in need thereof a pharmaceutical composition as described in any one of claims 2 to 5.

8. A method for treating pain, anxiety or depression, the method comprising administering to a subject in need thereof the pharmaceutical composition of any one of claims 2 to 5.

9. A method for preparing the crystalline form A according to claim 1, the method comprising the following steps: a. dissolving a certain amount of 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one in a certain amount of DMSO:ethanol at a ratio of 2:1 (v / v) at room temperature to form a supersaturated solution; b. adding to the solution in step a. an amount sufficient to precipitate the 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one in a 1:1 (v / v) ratio of ethanol:H2O; and c. Isolating the precipitated material from step b. to produce the crystalline Form A.

10. A method for preparing the crystalline form H according to claim 1, comprising the following steps: a. 4-chloro-5-(4-(4-fluoro-2-(trifluoromethyl)phenoxy)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)pyridazin-3(2H)-one was suspended in a 1:1 (v / v) ratio of isopropanol:isopropyl acetate; b. heating the suspension of step a. to a temperature between 45°C and 55°C under stirring for at least 24 hours; and c. separating the insoluble material from step b. to produce the crystalline Form H.

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