Polymorphs and salts of N-desmethyllboxystaurin

Abstract

Novel compositions of N-desmethyllboxystaurin L-lactate are provided. The use of N-desmethyllboxystaurin L-lactate compositions for modulating GSK-3 signaling is disclosed, as is the use of N-desmethyllboxystaurin composition L-lactate for inhibiting protein kinase C. Furthermore, N-desmethyllboxystaurin composition L-lactate is disclosed in the treatment of subjects with neurological and / or psychiatric disorders, including Alzheimer's disease, bipolar disorder, depression, schizophrenia, Parkinson's disease, or neuroinflammation, as well as in the treatment of diabetes or its complications, or symptoms associated with ischemia, inflammation, pulmonary hypertension, congestive heart failure, cardiovascular disease, skin disease, or cancer. Furthermore, N-desmethyllboxystaurin composition L-lactate administered in combination with lithium or other therapeutic agents for bipolar disorder is disclosed.

Description

[Technical Field]

[0001] Aspects of the present invention relate to novel compositions of N-desmethyllboxystaurin, a kinase inhibitor, for treating neurological or psychiatric disorders including Alzheimer's disease, bipolar disorder, depression, schizophrenia, Parkinson's disease, or neuroinflammation, and for treating diabetes and its complications, or ischemia, inflammation, pulmonary hypertension, congestive heart failure, cardiovascular disease, skin disease, cancer, or GM2 gangliosidosis, or other conditions for which N-desmethyllboxystaurin is clinically useful. [Background technology]

[0002] N-desmethylruboxystaurin has been shown to modulate GSK-3 signaling and inhibit protein kinase C.

[0003] Ruboxystaurin has been studied in several clinical trials for the treatment of diabetes and its complications (including diabetic retinopathy, diabetic neuropathy, and diabetic nephropathy). See A. Girach's U.S. Patent Publication 2008 / 0096923, which is incorporated herein by reference in its entirety. However, ruboxystaurin has several limitations, including the possibility of prolonging the QT interval on electrocardiograms in human subjects, a short half-life, a high plasma peak / trough ratio when administered daily, and the possibility of interaction with concomitant medications that inhibit CYP3A4 due to its metabolism by CYP3A4.

[0004] Recently, N-desmethyllboxystaurin has been found to be more effective, more stable, and to have superior pharmacokinetics compared to ruboxystaurin in terms of GSK-3 inhibition, and its metabolism is less affected by CYP3A4 inhibition. Therefore, N-desmethyllboxystaurin, either alone or in combination with other drugs, is a desirable alternative to ruboxystaurin, which is preferred for its desired inhibition of GSK-3 or protein kinase C (or both).

[0005] As a GSK-3 inhibitor, N-desmethyllboxystaurin has been proposed as a treatment for subjects with neurological and / or psychiatric disorders, including Alzheimer's disease, frontotemporal dementia, behavioral complications of dementia, bipolar disorder, depression, schizophrenia, Parkinson's disease, or neuroinflammation. Because GSK-3 inhibitors are known to increase WNT protein expression, they enhance the regenerative medicine pathways widely proposed for the treatment of neurological and psychiatric disorders. Inhibition of GSK-3 or enhancement of WNT signaling is associated with potential therapeutic approaches for type 2 diabetes, renal impairment (including diabetic nephropathy, chronic kidney disease, polycystic kidney disease, and focal segmental glomerulosclerosis), atherosclerosis, alopecia, bone and joint disorders (including osteoarthritis and osteoporosis), inflammatory diseases (including alcoholic hepatitis, inflammatory bowel disease, and septic shock), and eye disorders (including exudative age-related macular degeneration, atrophic age-related macular degeneration, diabetic macular edema, Fuchs corneal endothelial dystrophy, corneal epithelial cell deficiency, dry eye, glaucoma, familial exudative vitreoretinopathy (FEVR), Nolier's disease, Coats' disease, retinopathy of prematurity, macular telangiectasia, retinal vein occlusion, and Sjögren's syndrome). Inhibition of GSK-3 or enhancement of WNT signaling is associated with potential therapeutic approaches for ear disorders (including sensorineural and conductive hearing loss), lung disorders (including chronic obstructive pulmonary disease (COPD) and idiopathic pulmonary fibrosis), short bowel syndrome, and cancers (including melanoma, pancreatic cancer, prostate cancer, colon cancer, and leukemia). As a GSK-3 inhibitor, the use of N-desmethyllboxystaurine has been proposed for the treatment of bipolar disorder as monotherapy, in combination with lithium, or in combination with other bipolar disorder medications.

[0006] As a protein kinase C inhibitor, N-desmethyllboxystaurin has been proposed for the treatment of symptoms associated with diabetes, diabetic nephropathy, diabetic neuropathy, diabetic retinopathy, ischemia, inflammation, pulmonary hypertension, congestive heart failure, cardiovascular disease, skin diseases, cancer, and GM2 gangliosidosis. Protein kinase C inhibition has also been proposed for the treatment of bipolar disorder.

[0007] To develop N-desmethyllboxystaurin as a therapeutic agent, it needs to possess favorable physical and pharmacological properties that allow for delivery via a suitable route of administration. One such property is solubility, which can be enhanced by identifying the appropriate salt form and its crystalline form. Therefore, to develop it as a therapeutic agent, it is necessary to identify the appropriate salt form of N-desmethyllboxystaurin and the appropriate crystalline form of the salt form. [Overview of the Initiative]

[0008] Aspects of the present invention relate to novel compositions of N-desmethyllboxystaurin L-lactate, identified as N-desmethyllboxystaurin L-lactate type 1, N-desmethyllboxystaurin L-lactate type 2, and N-desmethyllboxystaurin L-lactate type 3.

[0009] For general reference, N-desmethyllboxystaurine hydrochloride was prepared according to the scheme shown in Figure 1. It was then converted to N-desmethyllboxystaurine L-lactate according to the scheme shown in Figure 2. (Figure 3) 1 The isolation of the desired salt form was confirmed by 1H NMR spectroscopy.

[0010] In one embodiment, the present invention is characterized by a crystalline form of N-desmethyllboxystaurin L-lactate, characterized by a powder X-ray diffractogram (XRPD) shown in Figure 4, which, when irradiated with CuKa X-rays, has peaks represented by 2θ at at least about 9.4°, 14.6°, 19.3°, 20.8°, and 23.5°. The above N-desmethyllboxystaurin L-lactate is named type 1 and is further characterized by DSC and TGA scans shown in Figure 5.

[0011] In one embodiment, the present invention is characterized by a crystalline form of N-desmethyllboxystaurin L-lactate, characterized by the XRPD shown in Figure 6, which, when irradiated with CuKa X-rays, has peaks represented by 2θ at at least about 10.6°, 11.0°, 14.7°, 17.3°, and 21.7°. The above N-desmethyllboxystaurin L-lactate is named type 2, and since this type is judged to be metastable, it is not further characterized by DSC and TGA scans.

[0012] In one embodiment, the present invention features a crystalline form of N-desmethyllboxystaurin L-lactate, characterized by the XRPD shown in Figure 7, which, when irradiated with CuKa X-rays, has peaks represented by 2θ at at least about 5.5° and 11.1°. The above N-desmethyllboxystaurin L-lactate is named type 3 and is further characterized by the DSC and TGA scans shown in Figure 8.

[0013] The relationships between types 1, 2, and 3 of N-desmethyllboxystaurine L-lactate are shown in Figure 9, with type 2 designated as metastable and type 3 designated as hydrate and lowest energy form.

[0014] Furthermore, aspects of the present invention relate to the therapeutic use of L-lactic acid compositions of N-desmethyllboxystaurin listed. [Brief explanation of the drawing]

[0015] [Figure 1] The synthesis scheme for N-desmethyllboxystaurine hydrochloride is shown. [Figure 2] This document presents a synthetic scheme for converting N-desmethyllboxystaurine hydrochloride to N-desmethyllboxystaurine L-lactate. [Figure 3] The 1H NMR spectrum of N-desmethyllboxystaurin L-lactate is shown. [Figure 4] This shows the XRPD pattern of N-desmethyllboxystaurin L-lactic acid type 1. [Figure 5] The TGA / DSC curve of N - desmethyl ruboxistaurin L - lactate form 1 is shown. [Figure 6] The XRPD pattern of N - desmethyl ruboxistaurin L - lactate form 2 is shown. [Figure 7] The XRPD pattern of N - desmethyl ruboxistaurin L - lactate form 3 is shown. [Figure 8] The TGA / DSC curve of N - desmethyl ruboxistaurin L - lactate form 3 is shown. [Figure 9] The transition map of forms 1, 2, and 3 of N - desmethyl ruboxistaurin L - lactate is shown. [Figure 10] The dynamic solubility of N - desmethyl ruboxistaurin L - lactate in water, SGF, FaSSIF, and FeSSIF is shown.

Mode for Carrying Out the Invention

[0016] The present invention relates to crystalline forms of N-desmethyllboxystaurin L-lactate. In one embodiment, the present invention is characterized by crystalline form 1 of N-desmethyllboxystaurin L-lactate, characterized by a powder X-ray diffractogram (XRPD) having peaks represented by 2θ at at least about 9.4°, 14.6°, 19.3°, 20.8° and 23.5° when irradiated with CuKa X-rays. In another embodiment of the present invention, form 1 of N-desmethyllboxystaurin L-lactate is characterized by a powder X-ray diffractogram (XRPD) having peaks represented by 2θ at at least about 9.4°, 14.6° and 19.3° when irradiated with CuKa X-rays. In another aspect of the present invention, type 1 of N-desmethyllboxystaurin L-lactate is characterized by a powder X-ray diffractogram (XRPD) having peaks represented by 2θ at at least about 14.6°, 19.3°, and 20.8° when irradiated with CuKa X-rays. In another aspect of the present invention, type 1 of N-desmethyllboxystaurin L-lactate is characterized by a powder X-ray diffractogram (XRPD) having peaks represented by 2θ at at least about 19.3°, 20.8°, and 23.5° when irradiated with CuKa X-rays. In another aspect of the present invention, type 1 of N-desmethyllboxystaurin L-lactate is characterized by a powder X-ray diffractogram (XRPD) having peaks represented by 2θ at at least about 9.4°, 20.8°, and 23.5° when irradiated with CuKa X-rays. In another aspect of the present invention, type 1 of N-desmethyllboxystaurin L-lactate is characterized by a powder X-ray diffractogram (XRPD) having peaks represented by 2θ at at least about 9.4°, 14.6°, and 23.5° when irradiated with CuKa X-rays.In another aspect of the present invention, type 1 of N-desmethyllboxystaurin L-lactate is characterized by a powder X-ray diffractogram (XRPD) having peaks represented by 2θ at at least about 9.4°, 19.3°, and 23.5° when irradiated with CuKa X-rays. In another aspect of the present invention, type 1 of N-desmethyllboxystaurin L-lactate is characterized by a powder X-ray diffractogram (XRPD) having peaks represented by 2θ at at least about 9.4°, 14.6°, and 20.8° when irradiated with CuKa X-rays. In another aspect of the present invention, type 1 of N-desmethyllboxystaurin L-lactate is characterized by a powder X-ray diffractogram (XRPD) having peaks represented by 2θ at at least about 14.6°, 19.3°, and 23.5° when irradiated with CuKa X-rays. In another aspect of the present invention, type 1 of N-desmethyllboxystaurin L-lactate is characterized by a powder X-ray diffractogram (XRPD) having peaks represented by 2θ at at least about 9.4°, 19.3°, and 20.8° when irradiated with CuKa X-rays. In yet another aspect of the present invention, type 1 of N-desmethyllboxystaurin L-lactate is characterized by a powder X-ray diffractogram (XRPD) having any number and / or combination of peaks represented by 2θ at at least about 9.4°, 14.6°, 19.3°, 20.8°, and 23.5° when irradiated with CuKa X-rays.

[0017] In one embodiment, the present invention features a crystalline form 2 of N-desmethyllboxystaurin L-lactate, characterized by a powder X-ray diffractogram (XRPD) having peaks represented by 2θ at at least about 10.6°, 11.0°, 14.7°, 17.3°, and 21.7° when irradiated with CuKa X-rays. In another embodiment of the present invention, the above-mentioned form 2 of N-desmethyllboxystaurin L-lactate is characterized by a powder X-ray diffractogram (XRPD) having peaks represented by 2θ at at least about 10.6°, 11.0°, and 14.7° when irradiated with CuKa X-rays. In yet another embodiment of the present invention, the above-mentioned form 2 of N-desmethyllboxystaurin L-lactate is characterized by a powder X-ray diffractogram (XRPD) having peaks represented by 2θ at at least about 11.0°, 14.7°, and 17.3° when irradiated with CuKa X-rays. In another aspect of the present invention, type 2 of N-desmethyllboxystaurin L-lactate is characterized by a powder X-ray diffractogram (XRPD) having peaks represented by 2θ at at least about 14.7°, 17.3°, and 21.7° when irradiated with CuKa X-rays. In another aspect of the present invention, type 2 of N-desmethyllboxystaurin L-lactate is characterized by a powder X-ray diffractogram (XRPD) having peaks represented by 2θ at at least about 10.6°, 17.3°, and 21.7° when irradiated with CuKa X-rays. In another aspect of the present invention, type 2 of N-desmethyllboxystaurin L-lactate is characterized by a powder X-ray diffractogram (XRPD) having peaks represented by 2θ at at least about 10.6°, 11.0°, and 21.7° when irradiated with CuKa X-rays. In another aspect of the present invention, type 2 of the above N-desmethyllboxystaurin L-lactate is characterized by a powder X-ray diffractogram (XRPD) having peaks represented by 2θ at at least about 11.0°, 17.3°, and 21.7° when irradiated with CuKa X-rays.In another aspect of the present invention, the type 2 of the above N-desmethylruboxistaurin L-lactate is characterized by a powder X-ray diffractogram (XRPD) having peaks represented by 2θ at at least about 10.6°, 14.7° and 21.7° when irradiated with Cu Ka X-rays. In another aspect of the present invention, the type 2 of the above N-desmethylruboxistaurin L-lactate is characterized by a powder X-ray diffractogram (XRPD) having peaks represented by 2θ at at least about 10.6°, 11.0° and 17.3° when irradiated with Cu Ka X-rays. In another aspect of the present invention, the type 2 of the above N-desmethylruboxistaurin L-lactate is characterized by a powder X-ray diffractogram (XRPD) having peaks represented by 2θ at at least about 11.0°, 14.7° and 21.7° when irradiated with Cu Ka X-rays. In another aspect of the present invention, the type 2 of the above N-desmethylruboxistaurin L-lactate is characterized by a powder X-ray diffractogram (XRPD) having peaks represented by 2θ at at least about 10.6°, 14.7° and 17.3° when irradiated with Cu Ka X-rays. In yet another aspect of the present invention, the type 2 of the above N-desmethylruboxistaurin L-lactate is characterized by a powder X-ray diffractogram (XRPD) having peaks represented by 2θ at at least about 10.6°, 11.0°, 14.7°, 17.3° and 21.7° in any number and / or combination when irradiated with Cu Ka X-rays.

[0018] In one embodiment, the present invention features a crystalline form 3 of N-desmethyllboxystaurin L-lactate, characterized by a powder X-ray diffractogram (XRPD) having peaks represented by 2θ at at least about 5.5° and 11.1° when irradiated with CuKa X-rays. In another embodiment of the present invention, type 2 of the above N-desmethyllboxystaurin L-lactate is characterized by a powder X-ray diffractogram (XRPD) having peaks represented by 2θ at at least about 5.5° when irradiated with CuKa X-rays. In yet another embodiment of the present invention, type 2 of the above N-desmethyllboxystaurin L-lactate is characterized by a powder X-ray diffractogram (XRPD) having peaks represented by 2θ at at least about 11.1° when irradiated with CuKa X-rays. In yet another aspect of the present invention, type 2 of the above N-desmethyllboxystaurin L-lactate is characterized by a powder X-ray diffractogram (XRPD) having any number and / or combination of peaks represented by 2θ at at least about 5.5° and 11.1° when irradiated with CuKa X-rays.

[0019] Unless otherwise defined, all terms used in the disclosure of this invention, including technical and scientific terms, have the meanings generally understood by those skilled in the art to which this invention pertains.

[0020] As disclosed herein, several ranges of values ​​are provided. Unless otherwise indicated in the context, each intermediate value between the upper and lower limits of that range is also specifically disclosed to the tenth of a unit of the lower limit. Smaller ranges between any stated value or intermediate value within a stated range and any other stated value or intermediate value within that stated range are included in the present invention. The upper and lower limits of these smaller ranges may be included in or excluded from the range independently, and each range that includes either the limitation, does not include the limitation, or includes both limitations is included in the present invention, subject to the specifically excluded limitation within the stated range. If a stated range includes one or both limitations, a range that excludes either or both of those included limitations is also included in the present invention. The term “about” generally includes ±10% of the stated value. For example, “about 10%” may refer to a range of 9% to 11%, and “about 20” may mean 18 to 22. Preferably, “about” includes ±6% of the stated value. Alternatively, "approximately" can include ±5% of the stated value. Other meanings of "approximately" may become clear from contexts such as rounding; for example, "approximately 1" could mean between 0.5 and 1.4.

[0021] The term "pharmaceutically acceptable salt" of a compound means a salt that is pharmaceutically acceptable and possesses the desired pharmacological activity of the parent compound. Acids that combine with a base to form a pharmaceutically acceptable salt are generally understood to be safe for pharmaceutical use. Such acids include inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, and phosphoric acid, or formic acid, acetic acid, propionic acid, hexanoic acid, cyclopentanepropionic acid, glycolic acid, pyruvic acid, lactic acid, malonic acid, succinic acid, malic acid, maleic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, 3-(4-hydroxybenzoyl)benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, 1,2-ethanedisulfonic acid, and 2-hydroxyethane. This includes organic acids such as sulfonic acid, benzenesulfonic acid, 4-chlorobenzenesulfonic acid, 2-naphthalenesulfonic acid, 4-toluenesulfonic acid, camphorsulfonic acid, glucoheptonic acid, 4,4'-methylenebis-(3-hydroxy-2-ene-1-carboxylic acid), 3-phenylpropionic acid, trimethylacetic acid, tertiary butylacetic acid, lauryl sulfate, gluconic acid, glutamic acid, hydroxynaphthoic acid, salicylic acid, stearic acid, and muconic acid. Additional information regarding appropriate pharmaceutically acceptable salts is found in "Remington's Pharmaceutical Sciences, 17th edition, Mack Publishing Company, Easton, PA, 1985," which is incorporated herein by reference.

[0022] As used herein, the term “therapeutic dose” means the amount of the compound of the present invention that can alleviate the symptoms of the various pathological conditions described herein. Naturally, the specific dose of the compound administered in accordance with the present invention will be determined by the specific circumstances surrounding the case, including, for example, the compound administered, the route of administration, the patient’s condition, and the pathological condition being treated. The administration may be once daily or divided into multiple doses per day (for example, two, three, or more times a day).

[0023] The effective dose of N-desmethyllboxystaurine or its pharmaceutically acceptable salts, solvates, or polymorphs is approximately 32 to 320 mg once daily or approximately 16 to 160 mg twice daily when used as monotherapy. A pharmaceutical composition of N-desmethyllboxystaurine or its pharmaceutically acceptable salts, solvates, or polymorphs further comprises at least one pharmaceutically acceptable adjuvant or excipient. In combination therapy, a dose less than the effective dose of N-desmethyllboxystaurine or its pharmaceutically acceptable salts, solvates, or polymorphs is approximately 8 to 32 mg once daily or approximately 4 to 16 mg twice daily. When N-desmethyllboxystaurine is used in combination with lithium, a dose less than the effective dose of lithium is approximately 60 to 600 mg once daily or approximately 30 to 300 mg twice daily. This dose of lithium is sufficient to prevent the kidney damage typically caused by lithium therapy. The effective once-daily dose of N-desmethyllboxystaurine or its pharmaceutically acceptable salt, solvate, or polymorph may be about 32, about 64, about 96, about 128, about 160, about 192, about 224, about 256, about 288, or about 320 mg. The effective twice-daily dose of N-desmethyllboxystaurine or its pharmaceutically acceptable salt, solvate, or polymorph may be about 16, about 32, about 48, about 64, about 80, about 96, about 112, about 128, about 144, or about 160 mg. Less than the effective once-daily dose of N-desmethyllboxystaurine or its pharmaceutically acceptable salt, solvate, or polymorph may be about 8, about 16, about 24, or about 32 mg. Less than two effective doses of N-desmethyllboxystaurine or its pharmaceutically acceptable salts, solvates, or polymorphs may be about 4, about 8, about 12, or about 16 mg per day.

[0024] For therapeutic use, N-desmethyllboxystaurin needs to be identified and characterized in terms of its salt and crystalline forms. Aspects of the present invention include the discovery, characterization, and production methods of novel crystalline forms of N-desmethyllboxystaurin L-lactic acid. These crystalline forms exhibit superior solubility compared to N-desmethyllboxystaurin hydrochloride.

[0025] The preparation of N-desmethyllboxystaurine hydrochloride (compound 1) generally follows the method shown in Figure 1. As illustrated, starting material 1 reacts with a vinyl Grignard reagent in the presence of copper iodide to produce alcohol intermediate 2. Those skilled in the art will recognize that substitutes for vinyl Grignard are useful in producing a similar conversion. Such substitutes include, but are not limited to, vinyl zinc reagents, vinyl copperate reagents, and vinyl lithium reagents. Those skilled in the art will recognize that substitutes for copper iodide are useful in facilitating the conversion from intermediate 1 to intermediate 2. Such substitutes include, but are not limited to, alternative Lewis acid reagents and chelating agents such as crown ethers.

[0026] Following the isolation of intermediate 2, Figure 1 shows its conversion to intermediate 3 upon reaction with allyl bromide. Those skilled in the art will recognize that alternative allylation agents are useful for the allylation from intermediate 2 to intermediate 3. Such allylation agents generally use substitutes for the bromide leaving group and include, but are not limited to, allyl chloride, allyl iodide, and allyl mesylate. Those skilled in the art will also recognize that substitutes for the illustrated potassium tert-butoxide base are useful for causing the reaction between intermediate 2 and the allylation agent. Such bases include, but are not limited to, hydride reagents, carbonate reagents, bicarbonate reagents, lithium diisopropylamide, and sodium hexamethyldisilazide.

[0027] Figure 1 further illustrates the two-step transformation from intermediate 3 to intermediate 4. As shown, the first step is an ozonolysis reaction, which causes the cleavage of the bisolefin to a bisaldehyde, and the second step is the reduction of the bisaldehyde to a bisalcohol with sodium borohydride. Those skilled in the art will recognize that ozonolysis is only one of several reactions or combinations of reactions suitable for the cleavage of olefins to aldehydes. Such transformations include, but are not limited to, the dihydroxylation of olefins and the subsequent cleavage of the resulting diol to an aldehyde. Suitable reagents for the dihydroxylation of olefins include, but are not limited to, osmium tetroxide. Suitable reagents for the cleavage of diols to aldehydes include, but are not limited to, sodium periodate and lead tetraacetate. Those skilled in the art will also recognize that alternatives to the sodium borohydride reducing agent are suitable for the reduction of aldehydes to alcohols. Such reagents include, but are not limited to, lithium aluminum hydride, diisopropylaluminum hydride, lithium borohydride, and borane. Those skilled in the art will recognize that alternatives to boron and aluminum-based reducing agents are also useful for the reduction of aldehydes to alcohols. Such alternatives include, but are not limited to, samarium iodide and triethylsilane.

[0028] As shown in Figure 1, the diol of intermediate 4 is converted to bismesylate intermediate 5 upon reaction with methanesulfonyl chloride and triethylamine. Those skilled in the art will recognize that general alternative leaving groups, including but not limited to chlorides, bromides, iodides, and tosylates, are also generally useful as leaving groups. Those skilled in the art will also recognize that alternatives to triethylamine are useful for converting alcohols to mesylates. Such alternatives include, but are not limited to, diisopropylethylamine, pyridine, carbonate reagents, and bicarbonate reagents.

[0029] The reaction between bismesilate intermediate 5 and bisindolylmaleimide intermediate 6, which forms intermediate 7, is shown in Figure 6, using cesium carbonate as the base. Those skilled in the art will recognize that alternative bases are useful for causing the reaction to intermediate 7 shown. Such bases include, but are not limited to, hydrides, alkoxides, carbonates, and bicarbonates.

[0030] The process of converting methylmaleimide to its demethylated version involves initial hydrolysis of intermediate 7 to maleic anhydride intermediate 8. As shown in Figure 1, this conversion is carried out using potassium hydroxide in ethanol. Those skilled in the art will recognize that the conversion from intermediate 7 to intermediate 8 can be performed using substitutes for potassium hydroxide, including but not limited to sodium hydroxide and lithium hydroxide. Furthermore, those skilled in the art will recognize that ethanol can be replaced with any protic solvent, including but not limited to methanol and water.

[0031] As shown in Figure 1, the conversion of maleic anhydride intermediate 8 to the corresponding maleimide intermediate 9 is completed during the reaction of intermediate 8 with hexamethyldisilazane. Those skilled in the art will recognize that maleic anhydride can be converted to maleimide using alternative reagents, including but not limited to ammonia and sodium amide.

[0032] With the maleimide established, Figure 1 shows the cleavage of the trityl protecting group from intermediate 9 to alcohol intermediate 10. In Figure 6, hydrochloric acid is highlighted as the reagent that causes trityl cleavage, but those skilled in the art will recognize that alternative acids can be used. These alternatives include, but are not limited to, hydrobromic acid, trifluoroacetic acid, and acetic acid.

[0033] As shown in Figure 1, the alcohol of intermediate 10 is converted to mesylate intermediate 11 upon reaction with methanesulfonyl chloride and pyridine. Those skilled in the art will recognize that general alternative leaving groups, including but not limited to chlorides, bromides, iodides, and tosylates, are also generally useful as leaving groups. Those skilled in the art will also recognize that alternatives to triethylamine are useful for converting alcohols to mesylates. Such alternatives include, but are not limited to, diisopropylethylamine, pyridine, carbonate reagents, and bicarbonate reagents.

[0034] In the final stage of synthesis, Figure 1 shows the conversion of intermediate 11 to compound 1 upon reaction with methylamine. Although not shown in Figure 6, methylamine is further converted to its corresponding hydrochloride salt upon treatment with hydrochloric acid. Those skilled in the art will understand that alternative strategies exist for converting compounds such as intermediate 11 to structures such as compound 1. Such strategies are generally recognizable to those skilled in the art and are generally supported by resources such as Comprehensive Organic Transformations (Larock, Wiley). Those skilled in the art will also recognize that, instead of forming a hydrochloride salt at this stage, any alternative salt form can be produced by substituting hydrochloric acid with another acid. Such alternative acids include, but are not limited to, L-lactic acid.

[0035] Those skilled in the art will recognize that, in addition to those described above and shown in Figure 1, there are many additional reactions useful for the production of N-desmethylruboxystaurin (compound 1). Suitable reactions are readily identifiable by those skilled in the art and are available in resources such as Comprehensive Organic Transformations (Larock, Wiley). Strategies for introducing and cleaving protecting groups are readily identifiable by those skilled in the art and are available in resources such as Protective Groups in Organic Synthesis (Greene and Wutz, Wiley). Those skilled in the art will also recognize that, generally applicable to all alternative and modified routes of the pathways described in Figure 1, similar chemical results enabling the production of compound 1 can be obtained by substituting different combinations of reagents, solvents, temperature conditions, and reaction times.

[0036] The production of N-desmethyllboxystaurine L-lactate from N-desmethyllboxystaurine hydrochloride generally follows the method shown in Figure 2.

[0037] As shown in Figure 2, compound 1 of Figure 1 is treated with an aqueous sodium bicarbonate solution to liberate N-desmethyllboxystaurine as its free base. Those skilled in the art will recognize that substitutes for sodium bicarbonate yield suitable results for the conversion of N-desmethyllboxystaurine hydrochloride to N-desmethyllboxystaurine free base. Suitable substitutes for sodium bicarbonate include, but are not limited to, inorganic bases such as sodium carbonate, potassium carbonate, potassium bicarbonate, and cesium carbonate. Suitable substitutes for sodium bicarbonate also include, but are not limited to, organic bases such as pyridine, triethylamine, and diisopropylethylamine.

[0038] As further shown in Figure 2, free N-desmethyllboxystaurin base is treated with L-lactic acid to produce N-desmethyllboxystaurin L-lactate. The produced N-desmethyllboxystaurin L-lactate was subjected to polymorph screening experiments to identify and characterize its various crystalline forms.

[0039] After manufacturing, the solubility of N-desmethyllboxystaurin L-lactate was investigated in comparison to the solubility of available hydrochloride and free base (both manufactured according to Figure 1). Dynamic solubility tests were evaluated over 24 hours in water, artificial gastric fluid (SGF), fasting artificial intestinal fluid (FaSSIF), and feeding artificial intestinal fluid (FeSSIF). As shown in Table 1, L-lactate showed approximately 10 to 20 times improvement in solubility in water after 4 hours compared to hydrochloride. The dynamic solubility of L-lactate is shown in Figure 10.

[0040] Table 1 Dynamic solubility of N-desmethyllboxystaurin L-lactate compared to free base and hydrochloride [Table 1]

[0041] N-desmethyllboxystaurin L-lactate exhibited superior solubility compared to available hydrochlorides and free bases, and its crystalline form was evaluated using a standard polymorph screening method. Those skilled in the art will recognize that the above polymorph screening method includes, but is not limited to, poor solvent addition temperature cycling, slurrying at room temperature, slurrying at high temperatures (e.g., 50°C), slow evaporation, solid-gas diffusion, liquid-gas diffusion, slow cooling, grinding, and polymer-induced crystallization.

[0042] After subjecting N-desmethyllboxystaurin L-lactate to standard polymorph screening experiments, three different unknown crystalline forms were isolated and characterized by XRPD, DSC, and TGA. Type 1, characterized by the XRPD pattern shown in Figure 4 and the DSC and TGA data shown in Figure 5, was shown to be an anhydride, as shown in Figure 9. Type 2, characterized by the XRPD pattern shown in Figure 6, was shown to be a metastable anhydride, as shown in Figure 9. Type 3, characterized by the XRPD pattern shown in Figure 7 and the DSC and TGA data shown in Figure 8, was shown to be an anhydride, as shown in Figure 9, and was the hydrate and lowest energy form among the three identified crystalline forms. These evaluations were performed by competitive transition studies, as summarized in the legend in Figure 9. Table 2 summarizes the properties of the identified crystalline forms.

[0043] Table 2 Summary of the characteristics of the crystalline form of N-desmethyllboxystaurin L-lactate [Table 2] * : Fever, --: Insufficient test volume

[0044] In one embodiment, the listed crystalline forms of N-desmethylruboxystaurin can be used as potential therapeutic agents for subjects with neurological disorders and / or mental disorders.

[0045] The compositions and therapeutic methods currently disclosed are suitable for all cases in which ruboxystaurin may be clinically useful as a GSK-3 inhibitor, including psychiatric and neurological disorders such as bipolar disorder, depression, Alzheimer's disease, autism spectrum disorder, fragile X syndrome, Pitt-Hopkins syndrome, traumatic brain injury, stroke, acute spinal cord injury, schizophrenia, Parkinson's disease, amyotrophic lateral sclerosis (ALS), and multiple sclerosis (MS). These compositions and therapeutic methods are suitable for conditions in which ruboxystaurin is applied as a protein kinase C inhibitor, including diabetes mellitus, diabetic nephropathy, diabetic neuropathy, diabetic retinopathy, ischemia, inflammation, cardiovascular disease, pulmonary hypertension, congestive heart failure, skin diseases, cancer, and GM2 gangliosidosis. These compositions and therapeutic methods are also suitable for conditions in which inhibition of GSK-3 and / or enhancement of WNT signaling have been proposed, including alopecia, osteoarthritis, osteoporosis, alcoholic hepatitis, inflammatory bowel disease, exudative age-related macular degeneration, atrophic age-related macular degeneration, diabetic macular edema, Fuchs corneal endothelial dystrophy, corneal epithelial cell defects, dry eye, glaucoma, familial exudative vitreoretinopathy (FEVR), Nolier's disease, Coats' disease, retinopathy of prematurity, macular telangiectasia, retinal vein occlusion, Sjögren's syndrome, sensorineural hearing loss, conductive hearing loss, schizophrenia, Parkinson's disease, polycystic kidney disease, focal segmental glomerulosclerosis, chronic obstructive pulmonary disease, idiopathic pulmonary fibrosis, short bowel syndrome, melanoma, pancreatic cancer, prostate cancer, colon cancer, leukemia, septic shock, and ischemia / reperfusion injury. Such compositions and methods are also suitable for GM2 gangliosidosis, for which the use of ruboxystaurin has been proposed.

[0046] In one embodiment, clinical responses to N-desmethyllboxystaurine or N-desmethyllboxystaurine in combination with lithium can help establish the diagnosis of bipolar disorder and other conditions for which GSK-3 inhibition is clinically useful. In Alzheimer's disease, positron emission tomography (PET) of GSK-3 beta activity has been developed as a diagnostic method. In some embodiments, N-desmethyllboxystaurine can be administered alone or in combination with lithium to subjects showing excessive GSK-3 beta activity on PET to treat Alzheimer's disease, and the decrease in GSK-3 beta activity on PET after administration of N-desmethyllboxystaurine can support the use of N-desmethyllboxystaurine (alone or in combination with lithium) as an appropriate treatment when administered at an appropriate dose.

[0047] In one embodiment, the compositions and therapeutic methods currently disclosed may also be used in veterinary applications to improve the health and well-being of livestock and pets by treating any of the aforementioned symptoms occurring in animals. Ruboxystaurin and N-desmethylruboxystaurin have equivalent efficacy in inhibiting protein kinase C.

[0048] Those skilled in the art will understand that numerous variations are possible without departing from the spirit of the invention. Therefore, it should be clearly understood that the various embodiments of the invention described herein are illustrative and do not limit the scope of the invention. Examples

[0049] instrumentation Powder X-ray diffraction (XRPD) XRPD analysis was performed using a PANalytical Empyrean and an X' Pert3 X-ray powder diffractometer. The XRPD parameters used are shown in Table 3.

[0050] Table 3 Parameters of the XRPD trial [Table 3]

[0051] Thermogravimetric analysis and differential scanning calorimetry (TGA and DSC)

[0052] TGA data was collected using TA Instruments' Discovery TGA 5500. DSC was performed using TA Instruments' Discovery DSC 2500. The detailed parameters used are shown in Table 4.

[0053] Table 4 Parameters of TGA and DSC tests [Table 4]

[0054] Solution nuclear magnetic resonance (NMR) spectroscopy Solution NMR was collected using a Bruker 400M NMR spectrometer with DMSO-d6 as the solvent.

[0055] Ion chromatography (IC)

[0056] Thermo Scientific with conductivity detector TM Dionex TM Aquion TM An ion chromatography (IC) system 1100 was used, and the detailed chromatography conditions are listed in Table 5.

[0057] Table 5 Chromatographic conditions and parameters for ion content testing [Table 5]

[0058] High-performance liquid chromatography (HPLC)

[0059] We used Waters H-Class chromatography, and the detailed chromatography conditions are listed in Table 6.

[0060] Table 6 Chromatographic conditions and parameters for purity and solubility tests [Table 6]

[0061] Example 1. Regarding the synthesis of the compound, N-desmethylruboxystaurine hydrochloride was synthesized according to the following procedure.

[0062] Step 1: Synthesis of (S)-1-(trityloxy)penten-4-en-2-ol (2) TIFF2026514867000007.tif56159 To a solution of magnesium vinyl bromide (1 M, 840 mL, 0.84 mol in THF), copper iodide (4.5 g, 23.62 mmol) was added at -40°C under a nitrogen environment. After stirring at -40°C for 20 min, compound 1 (150 g, 0.46 mmol) dissolved in dry THF (750 mL) was added dropwise to the reaction mixture, and the resulting reaction mixture was stirred at -40°C for 2 hours. After the reaction was complete (monitored by TLC), saturated ammonium chloride (1000 mL) was added. The reaction mixture was warmed to room temperature and extracted with ethyl acetate (1000 mL) while stirring. The organic layer was separated and washed with aqueous ammonia solution (250 mL). The organic layer was separated, dried over sodium sulfate, filtered, and evaporated under vacuum to obtain compound 2 (166 g, crude yield 100%) as a dark brown viscous substance. 1 H NMR (400 MHz, CDCl3): δ 7.45-7.42 (m, 6H), 7.32-7.28 (m, 6H), 7.26-7.22 (m, 3H), 5.74-5.71 (m, 1H), 5.09-5.02 (m, 2H), 3.85-3.82 (m, 1H), 3.18 (dd, J=9.6 Hz, J=4.0 Hz, 1H), 3.09 (dd, J=9.2 Hz, J=6.8 Hz, 1H), 2.27-2.22 (m, 3H).

[0063] Step 2 Synthesis of (S)-(((2-(allyloxy)penten-4-en-1-yl)oxy)methanetrityl)tribenzene (3) TIFF2026514867000008.tif46159 To a stirred solution of compound 2 (165 g, 0.48 mol) in dry THF (1500 mL), potassium tert-butoxide (70.0 g, 0.62 mmol) was gradually added under a nitrogen environment. The resulting reaction mixture was heated to 45°C, stirred for 2 hours, cooled to room temperature, and then allyl bromide (145.5 g, 1.22 mol) was added under room temperature conditions, and stirring continued for 1 hour under room temperature conditions. After the reaction was complete (monitored by TLC), saturated ammonium chloride (1500 mL) was added to the reaction mixture and extracted with ethyl acetate (1500 mL). The organic layer was separated, dried over sodium sulfate, filtered, and evaporated under vacuum to obtain crude compound 3. Crude compound 3 was further purified by silica gel column chromatography (100-200 mesh) and eluted with 0.5-1% ethyl acetate in hexane. The pure fraction was collected and evaporated under reduced pressure to obtain the desired compound 3 (106 g, 58% yield) as a pale yellow semi-solid. 1 H NMR (400 MHz, CDCl3): δ 7.48-7.44 (m, 6H), 7.31-7.26 (m, 6H), 7.25-7.20 (m, 3H), 5.95-5.88 (m, 1H), 5.75-5.70 (m, 1H), 5.27 (dd, J=17.2 Hz, J=2.0 Hz, 1H), 5.15 (dd, J=10.4 Hz, J=2.0 Hz, 1H), 5.03 (dd, J=17.2 Hz, J=2.0 Hz, 1H), 4.96 (dt, J=10.4 Hz, J=1.2 Hz, 1H), 4.12-4.10 (m, 1H), 4.04-4.02 (m, 1H), 3.52-3.49 (m, 1H), 3.17-3.09 (m, 2H), 2.35-2.31 (m, 2H)

[0064] Step 3 Synthesis of (S)-3-(2-hydroxyethoxy)-4-(trityloxy)butan-1-ol (4) TIFF2026514867000009.tif46159 Compound 3 (100 g, 0.26 mol) in MeOH:DCM (1:1) (800 mL) was bubbled with ozone gas at -45 °C for 18 hours. After the reaction was complete (monitored by TLC), the mixture was poured into a solution of sodium borohydride (21.5 g, 0.57 mol) in 0.5 N NaOH solution (370 mL) at 0 °C. The resulting reaction mixture was stirred at room temperature for 16 hours. After the reaction was complete (monitored by TLC), the reaction was quenched with 1 N HCl solution until the pH was 6-7. The resulting solution was then extracted with ethyl acetate (750 mL). The organic layer was separated, dried over sodium sulfate, filtered, and evaporated under vacuum to obtain the crude compound. The crude compound was further purified by silica gel column chromatography (100-200 mesh) and eluted with 20-25% ethyl acetate in hexane. The pure fraction was collected and evaporated to obtain the desired compound 4 (58 g, 57% yield) as a yellow viscous liquid. 1 H NMR (400 MHz, DMSO-d6): δ 7.42-7.40 (m, 6H), 7.34 (t, J=7.6 Hz, 6H), 7.28-7.24 (m, 3H), 4.60 (t, J=5.6 Hz 1H), 4.36 (t, J=5.6 Hz, 1H), 3.60-3.56 (m, 2H), 3.52-3.48 (m, 2H), 3.44-3.41 (m, 3H), 2.99-2.97 (m, 2H), 1.61-1.56 (m, 2H).

[0065] Step 4 Synthesis of (S)-2-((4-((methylsulfonyl)oxy)-1-(trityloxy)butan-2-yl)oxy)ethylmethanesulfonate (5) To a stirred solution of Compound 4 (60 g, 0.15 mol) in 35159 DCM (1000 mL) at 0 °C, triethylamine (66 mL, 0.47 mmol) was added and stirred for 15 min, then methanesulfonyl chloride (32.0 mL, 0.41 mmol) was added. The resulting reaction mixture was stirred at 0 °C for 2 h (the reaction was monitored by TLC), and the reaction was quenched with saturated ammonium chloride solution (600 mL). The organic layer was separated, dried over sodium sulfate, filtered, and evaporated under vacuum (≤25 °C) to obtain a crude compound. The crude compound was suspended in a 1:1 mixture of ethyl acetate and heptane (600 mL) and evaporated under vacuum to obtain a solid. The resulting solid compound was suspended in a 1:1 mixture of ethyl acetate and heptane (600 mL), stirred for 30 min, filtered, the solid was washed with heptane (80 mL), and dried under vacuum to obtain Compound 5 (88 g, crude yield 100%) as a cream-colored solid. 1 H NMR (400 MHz, DMSO-d6): δ 7.42-7.39 (m, 5H), 7.37-7.31 (m, 5H), 7.29-7.23 (m, 3H), 7.22-7.18 (m, 2H), 4.34-4.22 (m, 4H), 3.84-3.83 (m, 1H), 3.69-3.64 (m, 2H), 3.17 (s, 3H), 3.13 (s, 3H), 3.09-3.06 (m, 1H), 3.04-3.02 (m, 1H), 1.88-1.85 (m, 2H).

[0066] Step 5 Synthesis of (12E,32E,7S)-21-Methyl-7-((trityloxy)methyl)-22,25-dihydro-11H,21H,31H-6-oxa-1,3(3,1)-diindolo-2(3,4)-pyrrolasacyclononaphan-22,25-dione (7) To a stirred solution of compound 6 (41.5 g, 0.12 mol) in DMF (850 mL), cesium carbonate (86.0 g, 0.26 mol) was added. The reaction mixture was heated to 100 °C, and compound 5 (85.0 g (crude), 0.15 mol) was added dropwise at the same temperature. The resulting reaction mixture was stirred at 100 °C for 24 hours. After the reaction was complete (monitored by TLC), it was cooled to 50 °C, and Celite (25 g) was added and stirred for 15 minutes. The reaction product was filtered through Celite, and the filtrate was partitioned into ethyl acetate (800 mL) and water (400 mL). The organic layer was separated, dried over sodium sulfate, filtered, and evaporated under vacuum to obtain the crude compound. The crude compound was further purified by silica gel column chromatography (100-200 mesh) and eluted with 25-30% ethyl acetate in hexane. The pure fraction was collected and evaporated to obtain the desired compound 7 (55 g, 51% yield) as a reddish-brick-colored solid. 1 H NMR (400 MHz, DMSO-d6): δ 7.83 (d, J=7.6 Hz 1H), 7.75 (d, J=8.0 Hz 1H), 7.49 (d, J=8.4 Hz 1H), 7.45 (s, 1H), 7.41 (s, 1H), 7.34-7.26 (m, 10H), 7.25-7.22 (m, 6H), 7.18-7.15 (m, 2H), 7.12-7.06 (m, 2H), 4.25-4.24 (m, 1H), 4.17-4.04 (m, 3H), 3.71-3.67 (m, 1H), 3.55-3.50 (m, 1H), 3.31-3.30 (m, 1H), 3.07 (s, 3H), 3.05-3.00 (m, 2H), 2.10-2.07 (m, 1H), 2.02 (m, 1H).

[0067] Step 6: Synthesis of (12E,32E,7S)-7-((trityloxy)methyl)-22,25-dihydro-11H,31H-6-oxa-1,3(3,1)-diindra-2(3,4)-flanacyclonononaphane-22,25-dione (8) TIFF2026514867000012.tif65159 A stirred solution of compound 7 (85.0 g, 0.12 mol) in ethanol (850 mL) was mixed with potassium hydroxide (68.0 g, 1.22 mol) and heated to 80 °C. The resulting reaction mixture was stirred for 24 hours. After the reaction was complete (monitored by TLC), the reaction mixture was evaporated under vacuum to obtain the residue, which was then partitioned into DCM (850 mL) and a 20% citric acid solution (450 mL). The organic layer was separated, dried over sodium sulfate, filtered, and evaporated under vacuum to obtain crude compound 8 (62 g, yield 74%) as a dark brown solid. 1 H NMR (400 MHz, DMSO-d6): δ 7.88 (d, J=7.6 Hz 1H), 7.82 (d, J=7.6 Hz 1H), 7.65 (d, J=2.0 Hz 2H), 7.55 (d, J=8.0 Hz 1H), 7.41 (d, J=7.6 Hz 1H), 7.34-7.26 (m, 12H), 7.25-7.20 (m, 5H), 7.19-7.13 (m, 2H), 4.33-4.28 (m, 1H), 4.20-4.06 (m, 3H), 3.73-3.69 (m, 1H), 3.58-3.54 (m, 1H), 3.09-3.07 (m, 2H), 2.17–2.12 (m, 1H), 2.01–1.97 (m, 1H) (extra protons in the aromatic region are not included).

[0068] Step 7 Synthesis of (12E,32E,7S)-7-((trityloxy)methyl)-22,25-dihydro-11H,21H,31H-6-oxa-1,3(3,1)-diindra-2(3,4)-pyrrolacyclonononaphane-22,25-dione (9) TIFF2026514867000013.tif93127 Compound 8 (95.0 g, 0.25 mol) in DMF (950 mL) was stirred, to which HMDS (294.0 mL, 2.47 mol) and methanol (6.0 mL) were added, and the mixture was heated to 80°C. The reaction mixture was stirred at 80°C for 5 hours. After the reaction was complete (monitored by TLC), it was cooled to room temperature, the reaction was quenched with 1 N HCl solution (950 mL), and extracted with DCM (1500 mL). The organic layer was separated, dried over sodium sulfate, filtered, and evaporated under vacuum to obtain the crude compound (84 g). The crude compound was further purified by silica gel column chromatography (100-200 mesh) and eluted with 20-25% ethyl acetate in hexane. The pure fraction was collected and evaporated under vacuum to obtain the desired compound 9 (70 g, yield 74%) as a purple solid. 1 H NMR (400 MHz, DMSO-d6): δ 10.91 (s, 1H), 7.81 (d, J=8.0 Hz 1H), 7.73 (d, J=8.0 Hz 1H), 7.48 (d, J=8.4 Hz 2H), 7.43 (s, 1H), 7.39 (s, 1H), 7.33-7.23 (m, 12H), 7.23-7.21 (m, 3H), 7.18-7.14 (m, 2H), 7.11-7.06 (m, 2H), 4.27-4.23 (m, 1H), 4.13-4.00 (m, 3H), 3.70-3.67 (m, 1H), 3.55-3.47 (m, 1H), 3.33-3.26 (m, 1H), 3.02-2.99 (m, 2H), 2.13-2.08 (m, 1H), 2.01-1.98 (m, 1H).

[0069] Step 8 Synthesis of (12E,32E,7S)-7-(hydroxymethyl)-22,25-dihydro-11H,21H,31H-6-oxa-1,3(3,1)-diindra-2(3,4)-pyrrolacyclonononaphane-22,25-dione (10) TIFF2026514867000014.tif87127 Compound 9 (70.0 g, 0.18 mol) in ethanol (700 mL) was stirred, and 6N HCl (700 mL) was added at room temperature. The resulting reaction mixture was heated at 80 °C for 3 hours. After the reaction was complete (monitored by TLC), it was cooled to room temperature, stirred for 1 hour, filtered, washed with water (350 mL), and dried at 45 °C under vacuum to obtain compound 10 (40 g, crude yield 88%) as a purple solid. 1 H NMR (400 MHz, DMSO-d6): δ 10.92 (s, 1H), 7.82 (d, J=7.6 Hz 1H), 7.78 (d, J=7.6 Hz 1H), 7.53 (d, J=8.0 Hz, 1H), 7.51 (s, 1H), 7.46 (d, J=8.4 Hz, 1H), 7.45 (s, 1H), 7.25-7.22 (m, 2H), 7.13-7.10 (m, 2H), 4.69 (t, J=5.2 Hz 1H), 4.35-4.33 (m, 1H), 4.24-4.15 (m, 3H), 3.91-3.87 (m, 1H), 3.65-3.60 (m, 1H), 3.53-3.49 (m, 1H), 3.43-3.39 (m, 1H), 2.09-2.07 (m, 1H), 1.98-1.97 (m, 1H).

[0070] Step 9: Synthesis of ((12E,32E,7S)-22,25-dioxo-22,25-dihydro-11H,21H,31H-6-oxa-1,3(3,1)-diindra-2(3,4)-pyrrolacyclonononaphane-7-yl)methylmethanesulfonate (11) To a stirred solution of compound 10 (39.0 g, 0.09 mol) in TIFF2026514867000015.tif73127THF (400 mL), pyridine (33.2 mL, 0.39 mol) was added at room temperature, and the mixture was stirred for 20 minutes. Then, methanesulfonic anhydride (46.0 g, 0.26 mol) was added to the reaction mixture at room temperature. The resulting reaction mixture was stirred for 4 hours. After the reaction was complete (monitored by TLC), the reaction mixture was partitioned into ethyl acetate (100 mL) and water (50 mL). The organic layer was separated, dried over sodium sulfate, filtered, and evaporated under vacuum to obtain the crude compound (37.0 g). The crude compound was further purified by silica gel column chromatography (100-200 mesh) and eluted by DCM. The pure fraction was collected and evaporated under reduced pressure to obtain the desired compound 11 (30 g, yield 65%) as a purple solid. 1 H NMR (400 MHz, DMSO-d6): δ 10.92 (s, 1H), 7.83 (d, J=7.6 Hz 1H), 7.78 (d, J=7.6 Hz 1H), 7.54 (d, J=8.4 Hz, 1H), 7.52 (s, 1H), 7.48 (d, J=8.4 Hz, 1H), 7.46 (s, 1H), 7.22-7.17 (m, 2H), 7.14-7.10 (m, 2H), 4.44-4.38 (m, 2H), 4.22-4.14 (m, 4H), 3.93-3.90 (m, 1H), 3.66-3.61 (m, 1H), 3.17 (s, 3H), 2.19-2.14 (m, 1H), 2.03-1.98 (m, 1H).

[0071] Step 10: Synthesis of (12E,32E,7S)-7-((methylamino)methyl)-22,25-dihydro-11H,21H,31H-6-oxa-1,3(3,1)-diindra-2(3,4)-pyrrolacyclonononaphane-22,25-dione hydrochloride (compound 1) In an autoclave, 2M methylamine in THF (400 mL) was added at -40°C to a stirred solution of compound 11 (10.0 g, 0.019 mol) in THF (400 mL). The reaction mixture was gradually heated to 70°C and stirred for 24 hours. After the reaction was complete (monitored by TLC), the mixture was evaporated under vacuum to obtain the crude compound (12.0 g). This batch was combined with four additional batches of the same size to obtain 60.0 g of crude product. 60 g of the crude product was purified by silica gel column chromatography (230-400 mesh, 2% MeOH / DCM). The pure fraction was collected and concentrated to obtain the free base of the desired compound 1 (22.0 g) as a red solid. The free base was suspended in diethyl ether (220 mL) and cooled to 0°C. Ethanol-based HCl (33 mL) was added at 0°C. The resulting suspension was stirred at 0°C for 30 min, filtered, washed with diethyl ether (50 mL), and dried under vacuum at 40°C for 1 hour to obtain compound 1 (16.9 g, yield 36%) as a reddish-brick-colored solid. 1 H NMR (400 MHz, DMSO-d6): δ 10.93 (s, 1H, exchanged in D2O), 8.72-8.71 (m, 2H, exchanged in D2O), 7.81 (t, J=8.0 Hz, 2H), 7.55 (d, J=8.0 Hz 1H), 7.49 (s, 2H), 7.47 (d, J=8.4 Hz, 1H), 7.23 (t, J=7.2 Hz, 2H), 7.14 (t, J=7.2 Hz, 2H), 4.46-4.41 (m, 1H), 4.33-4.25 (m, 2H), 4.15-4.10 (m, 1H), 3.86-3.84 (m, 1H), 3.73-3.71 (m, 1H), 3.62 (t, J=9.2 Hz, 1H), 3.27-3.24 (m, 1H), 3.01-2.98 (m, 1H), 2.53 (t, J=5.6 Hz, 3H), 2.22-2.20 (m, 1H), 2.06-2.03 (m, 1H).

[0072] Example 2. Regarding the synthesis of the compound, N-desmethyllboxystaurin L-lactate was prepared from N-desmethyllboxystaurin hydrochloride according to the following procedure.

[0073] Step 1: Synthesis of desmethyllboxystaurin (free base) To a 10% methanol suspension of (desmethyllboxystaurin)HCl salt (10 g, 20.36 mmol) in DCM (3 L), aqueous sodium bicarbonate solution was added at 0-5°C until the pH of the reaction mixture reached 7.5-8.0. After stirring the reaction mixture for 1 hour (while constantly monitoring the pH), the organic layer was separated. The separated organic layer was washed with water (500 mL x 3), separated, dried over sodium sulfate, and evaporated under vacuum to obtain desmethyllboxystaurin (free base) (8.7 g, yield 94%) as a red solid.

[0074] Step 2: Synthesis of (desmethyllboxystaurine)-L-lactate TIFF2026514867000018.tif76127 A solution of desmethyllboxystaurin (free base) (8.7 g, 19.14 mmol) in acetone (240 mL) was stirred, to which a solution of L-lactic acid (1.84 g, 20.42 mmol) in acetone (10 mL) was added dropwise over 15-20 min, followed by the addition of 50 mg of (desmethyllboxystaurin)-L-lactate and seeding. The resulting reaction contents were stirred in the dark (RB wrapped in aluminum foil) for 7 days, the precipitated solid was filtered, washed with acetone (40 mL) to obtain a wet solid, and then freeze-dried for 16 hours (by freezing in CAN / water 3:1) to obtain lactate (7.8 g, yield 75%) as a red brick-colored solid compound. 1H NMR (400 MHz, DMSO-d6): δ 10.99 (br s, 1H), 7.83 (t, J=8.4 Hz, 2H), 7.56 (d, J=8.0 Hz 1H), 7.52 (s, 1H), 7.49-7.47 (m, 2H), 7.24-7.19 (m, 2H), 7.15-7.11 (m, 2H), 4.43-4.38 (m, 1H), 4.27-4.20 (m, 2H), 4.16-4.14 (m, 1H), 3.92 (q, J=6.8 Hz, 1H), 3.87-3.84 (m, 1H), 3.60 (t, J=8.8 Hz, 1H), 3.50 (br s, 1H), 2.79-2.76 (m, 1H), 2.69-2.64 (m, 1H), 2.33 (s, 3H), 2.17-2.16 (m, 1H), 2.04-2.02 (m, 1H), 1.21 (d, J=6.8 Hz, 1H).

[0075] Example 3. Preparation and Characterization of N-Desmethyllboxystaurin L-Lactate Type 1

[0076] Type 1 was obtained by slurring 50.0 mg of N-desmethyllboxystaltin free base and 11.2 mg of L-lactic acid (1.0 equivalent) in 1.0 mL of ethyl acetate at room temperature for 5 days (N2, dark environment). The solid was isolated by centrifugation and characterized after RT vacuum drying at room temperature. The XRPD (Figure 4) showed the peaks shown in the table below. The TGA / DSC curve (Figure 5) showed a weight loss of 4.19% up to 120.0 °C, with one endothermic peak at 87.7 °C and one exothermic peak at 112.7 °C (peak temperature).

[0077] [Table 7]

[0078] Example 4. Preparation and Characterization of N-Desmethyllboxystaurin L-Lactate Type 2

[0079] Type 2 was obtained by slurring the L-lactate type in acetone / H2O (857:143, v:v) at room temperature for 6 days (N2, dark environment). After centrifugation, the sample was air-dried overnight at room temperature to convert it to L-lactate type 3.

[0080] Three subsequent batches of L-lactic acid type 2 were reproduced by slurring L-lactic acid type 1 in acetone / H2O (857:143, v:v) at room temperature for 3 days (N2, dark environment). XRPD (Figure 6) recorded before conversion to type 3 showed the peaks shown in the table below. Type 2 is considered metastable because it tends to convert rapidly to type 3.

[0081] [Table 8]

[0082] Example 5. Preparation and Characterization of N-Desmethyllboxystaurin L-Lactate Type 3

[0083] Type 3 was obtained by slurring Type 1 in H2O under room temperature conditions for 6 days (N2, dark environment). After centrifugation, it was air-dried overnight at room temperature, and XRPD data showing the peaks shown in the table below (Figure 7) were collected.

[0084] Type 3 was reproduced by slurring Type 1 in H2O at room temperature for 3 days (N2, dark environment) and air-drying at room temperature for 4 hours. The TGA / DSC curve (Figure 8) showed a 3.95% weight loss up to 110.0°C, with four endothermic peaks at 98.2, 126.6, 150.8, and 221.5°C, and one exothermic peak at 165.0°C (peak temperature).

[0085] [Table 9]

[0086] Example 6. Evaluation of the solubility of N-methyl luboxystaurin L-lactate

[0087] The dynamic solubility of N-methylruboxystaurin L-lactate, hydrochloride, and free base was evaluated in H2O and three bio-culture media: SGF, FaSSIF, and FeSSIF. In the experiment, samples were mixed by rotation at 37°C and dissolved in H2O, SGF, FaSSIF, and FeSSIF with a solid load of approximately 10 mg / mL (calculated as free base). Solubility tests were performed at different time points (1 hour, 4 hours, and 24 hours). Samples from each time point were centrifuged and filtered (through a 0.45 μm PTFE membrane) to isolate the liquid phase for free base concentration and pH testing. N-methylruboxystaurin L-lactate showed the highest solubility in water at 7–10 mg / mL compared to its corresponding hydrochloride and free base.

[0088] [Table 10]

Claims

1. A crystalline form of N-desmethylruboxystaurin L-lactate having at least one peak at diffraction angles 2θ(°) of at least approximately 9.4°, 14.6°, or 19.3°, as measured by or calculated from X-ray diffraction using Cu Ka X-ray irradiation.

2. The crystalline form of N-desmethylruboxystaurin L-lactate according to claim 1, having at least one peak at a diffraction angle 2θ(°) of at least about 9.4°, 14.6°, or 19.3°, as measured by or calculated from X-ray diffraction by Cu Ka X-ray irradiation.

3. The crystalline form of N-desmethylruboxystaurin L-lactate according to claim 1, having at least one peak at diffraction angles 2θ(°) of at least about 9.4°, 14.6°, 19.3°, 20.8°, or 23.5°, as measured by or calculated from X-ray diffraction by Cu Ka X-ray irradiation.

4. A crystalline form of N-desmethyllboxystaurin L-lactate according to any one of claims 1 to 3, having the powder X-ray diffraction spectrum shown in Figure 4.

5. A crystalline form of N-desmethylruboxystaurin L-lactate having at least one peak at a diffraction angle 2θ(°) of at least about 10.6°, 11.0°, or 14.7°, as measured by or calculated from X-ray diffraction by Cu Ka X-ray irradiation.

6. A crystalline form of N-desmethylruboxystaurin L-lactate having at least one peak at a diffraction angle 2θ(°) of at least about 10.6°, 11.0°, or 14.7°, as measured by or calculated from X-ray diffraction by Cu Ka X-ray irradiation.

7. A crystalline form of N-desmethylruboxystaurin L-lactate having at least one peak at diffraction angles 2θ(°) of at least approximately 10.6°, 11.0°, 14.7°, 17.3°, or 21.7°, as measured by or calculated from X-ray diffraction by Cu Ka X-ray irradiation.

8. A crystalline form of N-desmethyllboxystaurin L-lactate according to any one of claims 5 to 7, having the powder X-ray diffraction spectrum shown in Figure 6.

9. A crystalline form of N-desmethylruboxystaurin L-lactate, which, when measured by or calculated from X-ray diffraction using Cu Ka X-ray irradiation, has at least one peak at a diffraction angle 2θ(°) of at least approximately 5.5° or 11.1°.

10. A crystalline form of N-desmethylruboxystaurin L-lactate, which, when measured by or calculated from X-ray diffraction using Cu Ka X-ray irradiation, has at least one peak at a diffraction angle 2θ(°) of at least approximately 5.5° or 11.1°.

11. A crystalline form of N-desmethyllboxystaurin L-lactate according to claim 9 or 10, having the powder X-ray diffraction spectrum shown in Figure 7.

12. A method for treating a disorder involving abnormal signaling of GSK-3 or protein kinase C, comprising administering a therapeutically effective dose of the crystalline form of N-desmethyllboxystaurin according to any one of claims 1 to 11 to a subject in need thereof.

13. The method according to claim 12, wherein the crystalline form of N-desmethylruboxystaurin is administered to a subject who 1) has never taken ruboxystaurin, 2) has taken ruboxystaurin and experienced side effects, 3) exhibits an extended QT interval, 4) shows a high plasma concentration of ruboxystaurin, 5) may be administered a drug that may interfere with the metabolism of ruboxystaurin, or 6) may require a high dose of ruboxystaurin and is of concern for side effects, QT prolongation, or adverse drug interactions.

14. The method according to claim 12 or 13, wherein the subject has a neurological disorder and / or mental disorder.

15. The method according to claim 14, wherein the disease / disorder is selected from Alzheimer's disease, frontotemporal dementia, behavioral complications of dementia, bipolar disorder, depression, schizophrenia, Parkinson's disease, neuroinflammation, autism spectrum disorder, fragile X syndrome, Pitt-Hopkins syndrome, Rett syndrome, traumatic brain injury, stroke, acute spinal cord injury, amyotrophic lateral sclerosis (ALS), multiple sclerosis (MS), neurofibromatosis type 1, neuronal ceroid lipofuscinosis, chronic pain, neuropathic pain, chemotherapy-induced neuropathy and / or chemotherapy-induced cognitive impairment.

16. The method according to claim 12 or 13, wherein the disease / disorder is selected from type 2 diabetes, diabetic retinopathy, diabetic neuropathy, diabetic macular edema, diabetic nephropathy, chronic kidney disease, polycystic kidney disease and / or focal segmental glomerulosclerosis.

17. The method according to claim 12 or 13, wherein the disease / disorder is selected from bone and joint disorders including atherosclerosis, alopecia, osteoarthritis and osteoporosis, inflammatory disorders including alcoholic hepatitis and inflammatory bowel disease, and septic shock.

18. The method according to claim 12 or 13, wherein the disease / disorder is selected from eye disorders including exudative age-related macular degeneration, atrophic age-related macular degeneration, Fuchs corneal endothelial dystrophy, corneal epithelial cell defects, dry eye, glaucoma, familial exudative vitreoretinopathy (FEVR), Nolier's disease, Coats' disease, retinopathy of prematurity, macular telangiectasia, retinal vein occlusion, and Sjögren's syndrome, and / or ear disorders including sensorineural hearing loss and conductive hearing loss.

19. The method according to claim 12 or 13, wherein the disease / disorder is selected from lung disorders including chronic obstructive pulmonary disease (COPD), idiopathic pulmonary fibrosis, and pulmonary hypertension, and / or cancers including melanoma, pancreatic cancer, prostate cancer, colon cancer, and leukemia, and / or short bowel syndrome, ischemia, inflammation, cardiovascular disease, congestive heart failure, skin disease, inflammation, or GM2 gangliosidosis.

20. The method according to any one of claims 12 to 19, wherein the crystalline form of N-desmethyllboxystaurin is administered once daily in an amount of about 32 to about 320 mg of N-desmethyllboxystaurin, or twice daily in an amount of about 16 to about 160 mg of N-desmethyllboxystaurin.

21. The method according to any one of claims 12 to 20, wherein the crystalline form of N-desmethyllboxystaurin is administered in combination with lithium.

22. The method according to any one of claims 12 to 20, wherein the subject does not react to lithium.

23. The method according to claim 21, wherein the subject reacts to lithium.

24. The method according to claim 21, wherein lithium is administered in a dose less than the effective dose used in monotherapy, and the crystalline form of N-desmethylruboxystaurin is administered in a dose less than the effective dose used in monotherapy.

25. The method according to claim 24, wherein a dose of lithium less than an effective dose is administered once daily at a dose of approximately 60 mg to approximately 600 mg, or twice daily at a dose of approximately 30 mg to approximately 300 mg.

26. The method according to claim 24, wherein less than an effective amount of the crystalline form of N-desmethyllboxystaurin is administered once daily at a dose of about 8 to about 32 mg, or twice daily at a dose of about 4 to about 16 mg.

27. A method for establishing a diagnosis of bipolar disorder or other conditions for which GSK-3 inhibition is clinically useful, comprising administering a therapeutically effective dose of N-desmethyllboxystaurin crystalline form to a subject to be evaluated, and evaluating the clinical response of the subject.

28. A method for establishing an appropriate therapeutic dose of N-desmethyllboxystaurin in a subject, comprising administering an increasing dose of the crystalline form of N-desmethyllboxystaurin and evaluating the response using GSK-3 imaging or GSK-3 serology.

29. A method for treating a subject having Alzheimer's disease, bipolar disorder, or depression who shows evidence of elevated GSK-3, comprising administering to the subject a therapeutically effective dose of the crystalline form of N-desmethyllboxystaurin and evaluating and monitoring the subject using positron emission tomography (PET) or serology.

30. A method for establishing a diagnosis of bipolar disorder or other conditions for which GSK-3 inhibition is clinically useful, comprising administering a therapeutically effective dose of N-desmethyllboxystaurin crystalline form together with a therapeutically effective dose of lithium to a subject to be evaluated, and evaluating the clinical response of the subject.

31. The method according to claim 30, wherein the doses of both N-desmethylruboxystaurin and lithium are less than the effective dose in monotherapy.

32. A method for treating a subject with Alzheimer's disease having elevated GSK-3 beta activity, comprising administering to the subject a therapeutically effective dose of N-desmethyllboxystaurine or a pharmaceutically acceptable salt, solvate, or polymorph thereof, and a therapeutically effective dose of lithium, and monitoring the subject using positron emission tomography (PET).

33. The method according to claim 32, wherein the doses of both the crystalline form and lithium of N-desmethyllboxystaurin are less than the effective dose in the case of monotherapy.

34. A method for establishing an appropriate therapeutic dose of the crystalline form of N-desmethyllboxystaurin in a subject, comprising administering the subject an increasing dose of the crystalline form of N-desmethyllboxystaurin and lithium, and evaluating the response using positron emission tomography (PET).

35. The crystalline form of N-desmethyllboxystaurin L-lactate.

36. The crystalline form of N-desmethyllboxystaurin L-lactate according to claim 35, having the X-ray powder diffraction pattern shown in Figure 4, and produced by slurring N-desmethyllboxystaurin free base with L-lactic acid in ethyl acetate and recovering the lactate.

37. The crystalline form of N-desmethyllboxystaurin L-lactate according to claim 35, having the X-ray powder diffraction pattern shown in Figure 6, and produced by slurring N-desmethyllboxystaurin free base with L-lactic acid in a mixture of acetone and water to recover the lactate.

38. The crystalline form of N-desmethyllboxystaurin L-lactate according to claim 35, having the X-ray powder diffraction pattern shown in Figure 7, and produced by slurring N-desmethyllboxystaurin free base with L-lactic acid in water and recovering the lactate.

Images