Methods for treating cysteamine-sensitive disorders

Abstract

To provide pharmaceuticals for treatment of cystinosis and other cysteamine sensitive disorders in a subject.SOLUTION: The present invention provides a pharmaceutical orally administered to a subject at least once daily, comprising a dose of 50 to 150 milligrams per kilogram of body weight (mg / kg) of N-acetylcysteine-pantetheine disulfide or cysteamine-N-acetylcysteine disulfide, or a pharmaceutically acceptable salt thereof.SELECTED DRAWING: None

Description

[Technical Field]

[0001] The present invention features compositions and methods for treating cysteamine-sensitive conditions, syndromes, and diseases. [Background technology]

[0002] Cysteamine is a naturally occurring aminothiol that is produced in vivo through the catabolism of pantetheine. Preclinical and early clinical studies suggest that cysteamine may be therapeutically active in a variety of diseases, but widespread clinical development has been hindered by the lack of a convenient dosing regimen and poor toxicology.

[0003] Cysteamine has several mechanisms of action, most of which are related to the reducing ability of its thiol moiety. Cysteamine was first clinically investigated in the 1950s as a radioprotectant for cancer patients undergoing radiation therapy and as a treatment for radiation poisoning. The thiol group of cysteamine can reduce free radicals and other oxidized compounds that may be harmful to cells, thereby contributing to redox homeostasis. Cysteamine can also indirectly neutralize harmful oxidants by increasing levels of other antioxidant thiols, such as glutathione and cysteine. For example, cysteamine can participate in thiol-disulfide exchange with cystine (the dimeric oxidized form of cysteine) to form cysteamine-cysteine ​​disulfide and free cysteine. Cysteamine can also form disulfides with cysteine ​​residues in proteins, thereby affecting their structure and function. Cysteamine can inhibit enzymes, including transglutaminases, caspases, matrix metalloproteinases, and glutaminyl cyclase. Cysteamine is a chelating agent with a particular affinity for copper. It also blocks the secretion of certain peptide hormones, including somatostatin.

[0004] Diseases for which there is preclinical or clinical evidence of a cysteamine therapeutic effect include neurodegenerative diseases, including Alzheimer's disease, Huntington's disease, and Parkinson's disease; inflammatory and fibrotic diseases of the kidney, liver, and lung; metabolic diseases, including the spectrum of diabetes, metabolic syndrome, and fatty liver disease; infectious diseases, including viral, bacterial, and parasitic infections; hypercholesterolemia; ischemic diseases, including ischemic heart disease or stroke; sickle cell anemia; genetic mitochondrial disorders; inherited diseases caused by arginine to cysteine ​​mutations; and cancer.

[0005] Unfortunately, cysteamine has very unpleasant sensory properties (malodorous and bitter taste) and can produce body and breath odor when ingested in therapeutically effective amounts (over 1 gram per day in adolescents and adults). Most patients also experience gastrointestinal side effects, including loss of appetite, nausea, vomiting, and / or stomach pain. Bad breath, body odor, and gastrointestinal side effects are all associated with high peak cysteamine blood levels (often more than 50-fold higher than endogenous cysteamine levels in healthy subjects). Furthermore, the elimination half-life of cysteamine is only approximately 25 minutes, necessitating frequent administration. In summary, existing oral formulations of cysteamine have problems with their organoleptic properties (bitter taste, malodorous taste), pharmacology (subtherapeutic blood concentrations for the majority of the dosing interval), toxicology (gastrointestinal and other side effects), and stability (short half-life due to oxidation). Many of these problems are inherent to the drug being a volatile thiol compound. As a result, many patients with cystinosis are not fully compliant with cysteamine therapy and suffer from disease progression as a result.

[0006] Clinical development is hindered by the inability of cysteamine preparations to deliver therapeutic levels of drug for a sustained period with acceptable toxicology.Therefore, there is a need for improved treatment regimens, including improved cysteamine-producing compounds, improved formulations, and improved administration regimens, which can produce sustained elevated blood levels of cysteamine while reducing peak concentration, and increase trough concentration to provide improved efficacy while minimizing side effects.In addition, taking into account the known inter-patient variability of cysteamine pharmacokinetics, there is a need for compositions that allow individualized administration regimens to improve efficacy and reduce toxicity. Summary of the Invention

[0007] In a first aspect, the present invention provides a method for administering Compound 1 at a dose of 50 to 150 milligrams per kilogram of body weight (mg / kg) (e.g., 60±10, 70±10, 80±10, 90±10, 100±25, 110±20, 120±10, 130±10, or 140±10 mg / kg), The present invention features a method for treating a cysteamine-sensitive disorder in a subject, comprising administering TIFF0007794914000001.tif23170, or a pharmaceutically acceptable salt thereof, to the subject one or more times daily (e.g., once, twice, or three times daily). In certain embodiments, a reducing agent is not administered to the subject within two hours (e.g., within 30 minutes, 1 hour, 90 minutes, or 2 hours) of administration of the dose. In certain embodiments, a reducing agent is administered to the subject between two and eight hours after administration of the dose. In certain embodiments, the reducing agent is administered 3±1 hours, 4±1 hours, 5±1 hours, 6±1 hours, 7±1 hours, or 4±2 hours after administration of the dose of Compound 1 or a pharmaceutically acceptable salt thereof. In one embodiment, the reducing agent is selected from glutathione, glutathione diethyl ester, gamma glutamylcysteine, dihydrolipoic acid, N-acetylcysteine, homocysteine, pantetheine, 4-phosphopantetheine, dephospho-coenzyme A, coenzyme A, vitamin E, and ascorbic acid. In certain embodiments, Compound 1 or a pharmaceutically acceptable salt thereof is formulated for immediate release. In certain embodiments, Compound 1 or a pharmaceutically acceptable salt thereof is formulated as a powder and the dosage form is a sachet. In certain embodiments, a pantetheinase inducer selected from the group consisting of a PPAR alpha agonist, a PPAR gamma agonist, or an Nrf2 inducer is administered to the subject. In one embodiment, the pantetheinase inducer is an isothiocyanate present in cruciferous vegetables, sulforaphane, S-allyl cysteine, diallyl trisulfide, oxidized fat, omega-3 fatty acid, or oleylethanolamide. In certain embodiments, 10-50 mg / kg (e.g., 15±5, 20±5, 25±5, 30±5, 35±5, 40±5, or 45±5 mg / kg) of cystamine or a pharmaceutically acceptable salt thereof is administered to a subject within 30 minutes (e.g., 5 minutes, 10 minutes, 15 minutes, 20 minutes, 25 minutes, or 30 minutes) of administration of Compound 1 or a pharmaceutically acceptable salt thereof. In one embodiment, the cystamine or a pharmaceutically acceptable salt thereof is formulated for immediate release. In certain embodiments, the cystamine or a pharmaceutically acceptable salt thereof is administered simultaneously with administration of Compound 1 or a pharmaceutically acceptable salt thereof.

[0008] In certain embodiments, within 30 minutes (e.g., 5 minutes, 10 minutes, 15 minutes, 20 minutes, 20 minutes, or 25 minutes) of administration of Compound 1 or a pharmaceutically acceptable salt thereof, a dose of 10 to 50 mg / kg (e.g., 15±5, 20±5, 25±5, 30±5, 35±5, 40±5, or 45±5 mg / kg) of Compound 3, TIFF0007794914000002.tif23170 or a pharmaceutically acceptable salt thereof is administered to the subject. Optionally, Compound 3 or a pharmaceutically acceptable salt thereof is formulated for immediate release. In certain embodiments, Compound 3 or a pharmaceutically acceptable salt thereof is administered simultaneously with the administration of Compound 1 or a pharmaceutically acceptable salt thereof.

[0009] In any of the above methods, the cysteamine-sensitive disorder may be selected from cystinosis; neurodegenerative diseases; neurodevelopmental diseases; neuropsychiatric diseases; mitochondrial diseases; fibrotic diseases of the kidney, liver, or lungs; parasitic infections; sickle cell anemia; cancer; ischemic diseases, including ischemic heart disease or stroke; chronic obstructive pulmonary disease (COPD); cystic fibrosis (CF); bacterial infections; viral infections; non-alcoholic steatohepatitis (NASH); alcoholic steatohepatitis; and non-alcoholic fatty liver disease (NAFLD), or any other cysteamine-sensitive disorder described herein.

[0010] In a related aspect, the invention features a kit that includes: (i) a first pharmaceutical composition comprising Compound 1 or a pharmaceutically acceptable salt thereof; (ii) a second pharmaceutical composition comprising a reducing agent; and (iii) instructions for administering the second pharmaceutical composition at least two hours after the first pharmaceutical composition is administered to a subject for the treatment of a cysteamine-sensitive disorder.

[0011] In another aspect, the invention features a kit that includes: (i) a first pharmaceutical composition comprising Compound 1 or a pharmaceutically acceptable salt thereof; (ii) a second pharmaceutical composition comprising a pantetheinase inducer; and (iii) instructions for administering the first pharmaceutical composition and the second pharmaceutical composition to a subject for the treatment of a cysteamine-sensitive disorder.

[0012] In another related aspect, the invention features a kit that includes: (i) a first pharmaceutical composition comprising Compound 1 or a pharmaceutically acceptable salt thereof; (ii) a second pharmaceutical composition comprising cystamine or a pharmaceutically acceptable salt thereof; and (iii) instructions for administering the second pharmaceutical composition within 30 minutes of administering the first pharmaceutical composition to a subject for the treatment of a cysteamine-sensitive disorder.

[0013] In another aspect, the invention features a kit that includes: (i) a first pharmaceutical composition comprising Compound 1, or a pharmaceutically acceptable salt thereof; (ii) a second pharmaceutical composition comprising Compound 3, or a pharmaceutically acceptable salt thereof; and (iii) instructions for administering the second pharmaceutical composition within 30 minutes of administering the first pharmaceutical composition to a subject for the treatment of a cysteamine-sensitive disorder.

[0014] In any of the above kits, the instructions can include instructions for practicing the methods of administering Compound 1 described above.

[0015] In another aspect, the present invention provides a method for administering Compound 2 at a dose of 50 to 150 milligrams per kilogram of body weight (mg / kg) (e.g., 60±10, 70±10, 80±10, 90±10, 100±25, 110±20, 120±10, 130±10, or 140±10 mg / kg), The present invention features a method for treating a cysteamine-sensitive disorder in a subject, comprising administering TIFF0007794914000003.tif24170, or a pharmaceutically acceptable salt thereof, to the subject one or more times daily (e.g., once, twice, or three times daily). In certain embodiments, a reducing agent is not administered to the subject within 2 hours (e.g., within 30 minutes, 1 hour, 90 minutes, or 2 hours) of administration of the dose. In certain embodiments, a reducing agent is administered to the subject between 2 and 8 hours after administration of the dose. In certain embodiments, the reducing agent is administered 3±1 hours, 4±1 hours, 5±1 hours, 6±1 hours, 7±1 hours, or 4±2 hours after administration of the dose of Compound 2 or a pharmaceutically acceptable salt thereof. In one embodiment, the reducing agent is selected from glutathione, glutathione diethyl ester, gamma glutamylcysteine, dihydrolipoic acid, N-acetylcysteine, homocysteine, pantetheine, 4-phosphopantetheine, dephospho-coenzyme A, coenzyme A, vitamin E, and ascorbic acid. In certain embodiments, Compound 2 or a pharmaceutically acceptable salt thereof is formulated for immediate release. In certain embodiments, Compound 2 or a pharmaceutically acceptable salt thereof is formulated as a powder and the dosage form is a sachet. In certain embodiments, a pantetheinase inducer selected from the group consisting of a PPAR alpha agonist, a PPAR gamma agonist, or an Nrf2 inducer is administered to the subject. In one embodiment, the pantetheinase inducer is an isothiocyanate present in cruciferous vegetables, sulforaphane, S-allyl cysteine, diallyl trisulfide, oxidized fat, omega-3 fatty acid, or oleylethanolamide. In certain embodiments, 10-50 mg / kg (e.g., 15±5, 20±5, 25±5, 30±5, 35±5, 40±5, or 45±5 mg / kg) of cystamine or a pharmaceutically acceptable salt thereof is administered to a subject within 30 minutes (e.g., 5 minutes, 10 minutes, 15 minutes, 20 minutes, 25 minutes, or 30 minutes) of administration of Compound 2 or a pharmaceutically acceptable salt thereof. In one embodiment, the cystamine or a pharmaceutically acceptable salt thereof is formulated for immediate release. In certain embodiments, the cystamine or a pharmaceutically acceptable salt thereof is administered simultaneously with administration of Compound 2 or a pharmaceutically acceptable salt thereof.

[0016] In certain embodiments, within 30 minutes (e.g., 5 minutes, 10 minutes, 15 minutes, 20 minutes, 20 minutes, or 25 minutes) of administration of Compound 2 or a pharmaceutically acceptable salt thereof, a dose of 10 to 50 mg / kg (e.g., 15±5, 20±5, 25±5, 30±5, 35±5, 40±5, or 45±5 mg / kg) of Compound 3, TIFF0007794914000004.tif22170 or a pharmaceutically acceptable salt thereof is administered to the subject. Optionally, Compound 3 or a pharmaceutically acceptable salt thereof is formulated for immediate release. In certain embodiments, Compound 3 or a pharmaceutically acceptable salt thereof is administered simultaneously with the administration of Compound 2 or a pharmaceutically acceptable salt thereof.

[0017] In any of the above methods, the cysteamine-sensitive disorder may be selected from cystinosis; neurodegenerative diseases; neurodevelopmental diseases; neuropsychiatric diseases; mitochondrial diseases; fibrotic diseases of the kidney, liver, or lungs; parasitic infections; sickle cell anemia; cancer; ischemic diseases, including ischemic heart disease or stroke; chronic obstructive pulmonary disease (COPD); cystic fibrosis (CF); bacterial infections; viral infections; non-alcoholic steatohepatitis (NASH); alcoholic steatohepatitis; and non-alcoholic fatty liver disease (NAFLD), or any other cysteamine-sensitive disorder described herein.

[0018] In a related aspect, the invention features a kit that includes: (i) a first pharmaceutical composition that includes Compound 2 or a pharmaceutically acceptable salt thereof; (ii) a second pharmaceutical composition that includes a reducing agent; and (iii) instructions for administering the second pharmaceutical composition at least two hours after the first pharmaceutical composition is administered to a subject for the treatment of a cysteamine-sensitive disorder.

[0019] In another aspect, the invention features a kit that includes: (i) a first pharmaceutical composition that includes Compound 2 or a pharmaceutically acceptable salt thereof; (ii) a second pharmaceutical composition that includes a pantetheinase inducer; and (iii) instructions for administering the first pharmaceutical composition and the second pharmaceutical composition to a subject for the treatment of a cysteamine-sensitive disorder.

[0020] In yet another aspect, the invention features a kit that includes: (i) a first pharmaceutical composition comprising Compound 2 or a pharmaceutically acceptable salt thereof; (ii) a second pharmaceutical composition comprising cystamine or a pharmaceutically acceptable salt thereof; and (iii) instructions for administering the second pharmaceutical composition within 30 minutes of administering the first pharmaceutical composition to a subject for the treatment of a cysteamine-sensitive disorder.

[0021] In yet another aspect, the invention features a kit that includes: (i) a first pharmaceutical composition comprising Compound 2, or a pharmaceutically acceptable salt thereof; (ii) a second pharmaceutical composition comprising Compound 3, or a pharmaceutically acceptable salt thereof; and (iii) instructions for administering the second pharmaceutical composition within 30 minutes of administering the first pharmaceutical composition to a subject for the treatment of a cysteamine-sensitive disorder.

[0022] In any of the above kits, the instructions can include instructions for practicing the method of administering Compound 2 described above.

[0023] The present invention relates to compound 3, The present invention features a pharmaceutical composition comprising TIFF0007794914000005.tif22170 or a pharmaceutically acceptable salt thereof. The pharmaceutical composition can comprise Compound 3 or a pharmaceutically acceptable salt thereof formulated for immediate release, delayed release, or sustained release. In certain embodiments, the pharmaceutical composition further comprises a second active agent comprising a cysteamine precursor or a pharmaceutically acceptable salt thereof. Optionally, the second active agent is formulated for immediate release. In certain embodiments, the second active agent is formulated for delayed release or sustained release. In one particular embodiment, Compound 3 or a pharmaceutically acceptable salt thereof is formulated for immediate release and the second active agent is formulated for delayed release.

[0024] In a related aspect, the invention features a method for treating a cysteamine-sensitive disorder in a subject, comprising administering to the subject a pharmaceutical composition containing a therapeutically effective amount of Compound 3, or a pharmaceutically acceptable salt thereof. The cysteamine-sensitive disorder may be selected from cystinosis; neurodegenerative diseases; neurodevelopmental diseases; neuropsychiatric disorders; mitochondrial diseases; fibrotic diseases of the kidney, liver, or lung; parasitic infections; sickle cell anemia; cancer; ischemic diseases, including ischemic heart disease or stroke; chronic obstructive pulmonary disease (COPD); cystic fibrosis (CF); bacterial infections; viral infections; nonalcoholic steatohepatitis (NASH); alcoholic steatohepatitis; and nonalcoholic fatty liver disease (NAFLD), or any other cysteamine-sensitive disorder described herein.

[0025] In another aspect, the present invention provides a method for administering Compound 3 at a dose of 50 to 150 milligrams per kilogram of body weight (mg / kg) (e.g., 60±10, 70±10, 80±10, 90±10, 100±25, 110±20, 120±10, 130±10, or 140±10 mg / kg). The present invention features a method for treating a cysteamine-sensitive disorder in a subject, comprising administering TIFF0007794914000006.tif22170, or a pharmaceutically acceptable salt thereof, to the subject one or more times daily (e.g., once, twice, or three times daily). In certain embodiments, a reducing agent is not administered to the subject within 2 hours (e.g., within 30 minutes, 1 hour, 90 minutes, or 2 hours) of administration of the dose. In certain embodiments, a reducing agent is administered to the subject between 2 and 8 hours after administration of the dose. In certain embodiments, a reducing agent is administered 3±1 hours, 4±1 hours, 5±1 hours, 6±1 hours, 7±1 hours, or 4±2 hours after administration of the dose of Compound 3 or a pharmaceutically acceptable salt thereof. In one embodiment, the reducing agent is selected from glutathione, glutathione diethyl ester, gamma glutamylcysteine, dihydrolipoic acid, N-acetylcysteine, homocysteine, pantetheine, 4-phosphopantetheine, dephospho-coenzyme A, coenzyme A, vitamin E, and ascorbic acid. In certain embodiments, Compound 3 or a pharmaceutically acceptable salt thereof is formulated for immediate release. In certain embodiments, Compound 3 or a pharmaceutically acceptable salt thereof is formulated as a powder and the dosage form is a sachet. In certain embodiments, a pantetheinase inducer selected from the group consisting of a PPAR alpha agonist, a PPAR gamma agonist, or an Nrf2 inducer is administered to the subject. In one embodiment, the pantetheinase inducer is an isothiocyanate present in cruciferous vegetables, sulforaphane, S-allyl cysteine, diallyl trisulfide, oxidized fat, omega-3 fatty acid, or oleylethanolamide. In certain embodiments, 10-50 mg / kg (e.g., 15±5, 20±5, 25±5, 30±5, 35±5, 40±5, or 45±5 mg / kg) of cystamine or a pharmaceutically acceptable salt thereof is administered to a subject within 30 minutes (e.g., 5 minutes, 10 minutes, 15 minutes, 20 minutes, 25 minutes, or 30 minutes) of administration of Compound 3 or a pharmaceutically acceptable salt thereof. In one embodiment, the cystamine or a pharmaceutically acceptable salt thereof is formulated for immediate release. In certain embodiments, the cystamine or a pharmaceutically acceptable salt thereof is administered simultaneously with administration of Compound 3 or a pharmaceutically acceptable salt thereof.

[0026] In any of the above methods, the cysteamine-sensitive disorder may be selected from cystinosis; neurodegenerative diseases; neurodevelopmental diseases; neuropsychiatric diseases; mitochondrial diseases; fibrotic diseases of the kidney, liver, or lungs; parasitic infections; sickle cell anemia; cancer; ischemic diseases, including ischemic heart disease or stroke; chronic obstructive pulmonary disease (COPD); cystic fibrosis (CF); bacterial infections; viral infections; non-alcoholic steatohepatitis (NASH); alcoholic steatohepatitis; and non-alcoholic fatty liver disease (NAFLD), or any other cysteamine-sensitive disorder described herein.

[0027] In a related aspect, the invention features a kit that includes: (i) a first pharmaceutical composition that includes Compound 3 or a pharmaceutically acceptable salt thereof; (ii) a second pharmaceutical composition that includes a reducing agent; and (iii) instructions for administering the second pharmaceutical composition at least two hours after the first pharmaceutical composition is administered to a subject for the treatment of a cysteamine-sensitive disorder.

[0028] In another aspect, the invention features a kit that includes: (i) a first pharmaceutical composition that includes Compound 3 or a pharmaceutically acceptable salt thereof; (ii) a second pharmaceutical composition that includes a pantetheinase inducer; and (iii) instructions for administering the first pharmaceutical composition and the second pharmaceutical composition to a subject for the treatment of a cysteamine-sensitive disorder.

[0029] In yet another aspect, the invention features a kit that includes: (i) a first pharmaceutical composition comprising Compound 3 or a pharmaceutically acceptable salt thereof; (ii) a second pharmaceutical composition comprising cystamine or a pharmaceutically acceptable salt thereof; and (iii) instructions for administering the second pharmaceutical composition within 30 minutes of administering the first pharmaceutical composition to a subject for the treatment of a cysteamine-sensitive disorder.

[0030] In any of the above kits, the instructions can include instructions for practicing the method of administering Compound 3 described above.

[0031] In another aspect, the invention features an acid addition salt of Compound 1, wherein the acid is hydrochloric acid, acetic acid, trifluoroacetic acid, or tartaric acid. In certain embodiments, the acid is acetic acid or tartaric acid. In certain embodiments, the acid is acetic acid.

[0032] In one embodiment, the acid addition salt has the formula: TIFF0007794914000007.tif23170 In some embodiments, the acid addition salt has the following formula: TIFF0007794914000008.tif23170

[0033] In a particular embodiment, the acid is tartaric acid.

[0034] In one embodiment, the acid addition salt has the formula: TIFF0007794914000009.tif21170 In one embodiment, the acid addition salt has the formula: TIFF0007794914000010.tif21170In one embodiment, the acid addition salt has the formula: TIFF0007794914000011.tif23170

[0035] In another aspect, the invention features a method for treating a cysteamine-sensitive disorder in a subject, comprising administering to the subject an effective amount of any of the foregoing acid addition salts.

[0036] In certain embodiments, the reducing agent is not administered to the subject within 2 hours (e.g., within 30 minutes, 1 hour, 90 minutes, or 2 hours) of administration of the dose. In certain embodiments, the reducing agent is administered to the subject between 2 hours and 8 hours after administration of the dose. In certain embodiments, the reducing agent is administered 3±1 hours, 4±1 hours, 5±1 hours, 6±1 hours, 7±1 hours, or 4±2 hours after administration of the dose of Compound 1 or a pharmaceutically acceptable salt thereof. In one embodiment, the reducing agent is selected from glutathione, glutathione diethyl ester, gamma glutamylcysteine, dihydrolipoic acid, N-acetylcysteine, homocysteine, pantetheine, 4-phosphopantetheine, dephospho-coenzyme A, coenzyme A, vitamin E, and ascorbic acid.

[0037] In certain embodiments, any of the above-mentioned acid addition salts are formulated for immediate release.In certain embodiments, any of the above-mentioned acid addition salts are formulated as powder, and the dosage form is a sachet, or is formulated in a bottle for resuspension in a drinkable aqueous solution.In certain embodiments, a pantetheinase inducer selected from the group comprising PPAR alpha agonist, PPAR gamma agonist, or Nrf2 inducer is administered to the subject.In one embodiment, the pantetheinase inducer is isothiocyanate, sulforaphane, S-allyl cysteine, diallyl trisulfide, oxidized fat, omega-3 fatty acid, or oleylethanolamide present in cruciferous vegetables. In certain embodiments, 10-50 mg / kg (e.g., 15±5, 20±5, 25±5, 30±5, 35±5, 40±5, or 45±5 mg / kg) of cystamine or a pharmaceutically acceptable salt thereof is administered to a subject within 30 minutes (e.g., 5 minutes, 10 minutes, 15 minutes, 20 minutes, 25 minutes, or 30 minutes) of administration of any of the aforementioned acid addition salts. In one embodiment, the cystamine or a pharmaceutically acceptable salt thereof is formulated for immediate release. In certain embodiments, the cystamine or a pharmaceutically acceptable salt thereof is administered simultaneously with administration of any of the aforementioned acid addition salts.

[0038] In certain embodiments, within 30 minutes (e.g., 5 minutes, 10 minutes, 15 minutes, 20 minutes, 20 minutes, or 25 minutes) of administration of Compound 1 or a pharmaceutically acceptable salt thereof, a dose of 10 to 50 mg / kg (e.g., 15±5, 20±5, 25±5, 30±5, 35±5, 40±5, or 45±5 mg / kg) of Compound 3, TIFF0007794914000012.tif22170 or a pharmaceutically acceptable salt thereof is administered to the subject. Optionally, Compound 3 or a pharmaceutically acceptable salt thereof is formulated for immediate release. In certain embodiments, Compound 3 or a pharmaceutically acceptable salt thereof is administered simultaneously with the administration of Compound 1 or a pharmaceutically acceptable salt thereof.

[0039] In any of the above methods, the cysteamine-sensitive disorder may be selected from cystinosis; neurodegenerative diseases; neurodevelopmental diseases; neuropsychiatric diseases; mitochondrial diseases; fibrotic diseases of the kidney, liver, or lungs; parasitic infections; sickle cell anemia; cancer; ischemic diseases, including ischemic heart disease or stroke; chronic obstructive pulmonary disease (COPD); cystic fibrosis (CF); bacterial infections; viral infections; non-alcoholic steatohepatitis (NASH); alcoholic steatohepatitis; and non-alcoholic fatty liver disease (NAFLD), or any other cysteamine-sensitive disorder described herein.

[0040] In yet another aspect, the disclosure features a method of synthesizing an asymmetric disulfide, the method including combining (a) 1 molar equivalent of an organic carboxylic acid or salt thereof; (b) 2 to 4 molar equivalents (e.g., 2, 2.5, 3, 3.5, or 4 molar equivalents) of cystamine or salt thereof; and (c) an amide coupling reagent to form a mixture in which greater than 90% (e.g., 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 99.5%) of the organic carboxylic acid or salt thereof is converted to the asymmetric disulfide of Formula (A) and less than 10% (e.g., 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or 0.5%) of the organic carboxylic acid remains unreacted or is converted to the symmetric disulfide of Formula (B); TIFF0007794914000013.tif21170, where R is an organic radical. In certain embodiments, the group RC(O)- is a moiety of less than 1,000 daltons, 700 daltons, 400 daltons, or 200 daltons. In some embodiments, RC(O)- is a natural or synthetic amino acid, or an ester or amide thereof.

[0041] In some embodiments, the organic carboxylic acid or its salt is selected from pantothenic acid, 4-phosphopantothenic acid, acetic acid, or a salt thereof. In certain embodiments, the organic carboxylic acid or its salt is pantothenic acid or a salt thereof. In some embodiments, the amide coupling reagent is hydroxybenzotriazole. In some embodiments, the amide coupling reagent is N,N'-dicyclohexylcarbodiimide. In some embodiments, the combining is in one or more solvents selected from dichloromethane, tetrahydrofuran, acetonitrile dimethylformamide, water, and combinations thereof.

[0042] In another aspect, the disclosure features a method of synthesizing an asymmetric disulfide, the method including: (a) 2 to 4 molar equivalents (e.g., 2 to 2.3, 2.2 to 3, 2.7 to 3.3, or 3.1 to 4 molar equivalents) of a thiol selected from cysteamine, N-acetylcysteamine, cysteine, N-acetylcysteine ​​amide, or a salt thereof, and (b) combining in one molar equivalent a disulfide selected from cystamine, pantethine, or a salt thereof.

[0043] In some embodiments, the molar ratio of thiol to disulfide is about 2:1 to about 4:1 (e.g., about 2:1 to about 2.5:1, about 2.5:1 to 3.5:1, about 2.7:1 to about 3.3:1, about 2.5:1 to about 3:1, about 3:1 to about 3.5, or about 3:1 to about 3.5, about 4:1, about 3.5:1 to about 4:1). In certain embodiments, the molar ratio of thiol to disulfide is about 2.5:1 to about 3.5:1 (e.g., about 2.5:1 to 2.7:1, about 2.7:1 to about 3.3:1, or about 3.3:1 to about 3.5:1). In other embodiments, the molar ratio of thiol to disulfide is about 2.7:1 to about 3.3:1 (eg, about 2.7:1 to about 3.1:1, or about 3.1 to about 3.3).

[0044] In some embodiments, the thiol is N-acetylcysteamine or a salt thereof.

[0045] In some embodiments, the disulfide is cystamine or a salt thereof.

[0046] In some embodiments, the organic solvent is methanol or ethanol. In certain embodiments, the organic solvent is methanol.

[0047] definition "Immediate release" refers to a manner in which an active agent (e.g., a cysteamine precursor, or a pharmaceutically acceptable salt thereof) formulated in a unit dosage form has a dissolution release profile in simulated gastric fluid, wherein at least 55%, 65%, 75%, 85%, or 95% of the agent is released within the first two hours of testing using USP compliant equipment.

[0048] "Controlled release" refers to a manner in which an active agent (e.g., a cysteamine precursor, or a pharmaceutically acceptable salt thereof) is released from the formulation in a manner that allows for control of either the anatomical release site or the rate of release, or both. Generally, the purpose of a controlled-release formulation is to extend the period during which therapeutic drug levels are present in the body (e.g., relative to an immediate-release formulation) and / or optimize drug delivery to the cysteamine absorption site, thereby reducing the number of doses that must be administered in a 24-hour period. Gastroretentive, delayed-release, sustained-release, and colon-targeted formulations are all examples of controlled-release formulations. Controlled-release formulations may also allow for a reduction in the peak concentration of the drug (Cmax) relative to that observed for an immediate-release formulation administered at the same dose level (i.e., a reduced cysteamine Cmax in the case of the cysteamine precursors of the present invention). Controlled release formulations of active agents can be achieved, for example, by embedding the active agent in a matrix material that dissolves or erodes slowly, so that the active ingredient slowly and regularly leaches out of the coating by either diffusion from the matrix or erosion of the surface of the matrix, or both, or by the formation of a gel with a semipermeable surface through which the drug slowly exits the semipermeable layer.

[0049] "Delayed release" refers to a pharmaceutical preparation, e.g., an oral dosage formulation, that passes substantially intact through the acidic environment of the stomach and dissolves in the more basic environment of the small intestine, thereby enabling the active agent (e.g., a cysteamine precursor or a pharmaceutically acceptable salt thereof) formulated in a unit dosage form to have a dissolution release profile in simulated gastric fluid in which less than 25%, 20%, 15%, 10%, or 5% of the drug is released within the first hour of testing, and additionally, a dissolution release profile in simulated intestinal fluid at pH 6.0, 6.3, or 6.5 in which at least 55%, 65%, 75%, 85%, or 95% of the drug is released within the first two hours of testing. In some embodiments, the delayed release of the active agent (e.g., a cysteamine precursor or a pharmaceutically acceptable salt thereof) results from the use of a pH-sensitive enteric coating on the oral dosage form. The enteric coating can be combined, for example, with a rapid or slow (sustained) release formulation, or a combination of the two, to extend the period over which the drug is released.

[0050] The term "sustained release" (also referred to in the literature as "extended release") refers to a drug formulation that provides sustained release of a drug over an extended period of time, e.g., 6-12 hours or more, compared to an immediate-release formulation of the same drug, whereby an active agent (e.g., a cysteamine precursor, or a pharmaceutically acceptable salt thereof) formulated in a unit dosage form has a dissolution release profile in simulated gastric or intestinal fluid in which at least 10-45% of the drug (i.e., 15-45%, 20-45%, 25-45%, 35-45%, 30-45%, or 40-45%) is released within the first 3 hours of the test, and when in simulated small intestinal fluid, 65%, 75%, 85%, 90%, 93%, 95%, or 97% or more of the drug is released within 8 hours. Preferably, but not necessarily, sustained release results in substantially constant blood levels of the drug within the therapeutic range for the disease being treated over an extended period of time. Preferably, the sustained release formulation of cysteamine precursor produces plasma cysteamine levels that fall within a concentration range of, for example, 5-50 μM, 5-40 μM, 5-35 μM, 5-30 μM, 5-25 μM, 5-20 μM, or between 10-50 μM, 10-45 μM, 10-40 μM, 10-35 μM, 10-30 μM, 10-25 μM, or 10-20 μM.

[0051] The term "colon-targeted" refers to a formulation or composition that provides drug release in the colon (where gut flora is much more dense than in the small intestine) and, optionally, in the distal ileum (which tends to be the most alkaline region of the gastrointestinal tract). One method for targeting drug release to the distal ileum and colon is to use a pH-sensitive coating that dissolves around pH 7 (e.g., pH 6.8, pH 6.9, pH 7.0), a typical pH in the ileum. Formulations designed for pH-dependent drug release in the ileum are highly likely to release the drug in the colon as well (especially if the drug is embedded in a sustained-release matrix), and / or some of the cysteamine precursor released in the ileum may still be transported to the colon in precursor form (i.e., not yet converted to cysteamine). Another type of colon-targeted formulation relies on enzymes produced by gut bacteria to degrade drug-encapsulating polymers that cannot be degraded by salivary, gastric, or pancreatic enzymes, thereby resulting in drug delivery in the colon. The density of intestinal flora is also high in the distal ileum, so the intestinal flora can begin to digest the polymer and thus release the drug in the distal ileum. Ileum and colon targeted formulations are collectively referred to herein as colon targeted formulations.

[0052] The term "unit dosage form" refers to physically separate units suitable as a monolithic dosage form, such as pills, tablets, caplets, hard capsules, or soft capsules, each containing a predetermined amount of a cysteamine precursor or a pharmaceutically acceptable salt thereof. A "hard capsule" refers to a capsule containing a membrane forming a two-part capsule-shaped container capable of holding a solid or liquid payload of drug and excipients. A "soft capsule" refers to a capsule formed into a single container holding a liquid, semi-solid, or solid payload of drug and excipients. Granules, powders, and liquids can also be provided in "unit dosage form" by using appropriate packaging. For example, granules or powders can be administered in sachets, and liquids in ampoules, vials, or plastic containers.

[0053] The term "microparticle," as used herein, refers to microbeads, microspheres, micropellets, nanoparticles, nanobeads, nanospheres, or other microparticles used in drug formulations, each having an average diameter of between 0.05 and 999 micrometers. Dozens, hundreds, or thousands of such microparticles can be used in a single unit dosage form; for example, they can be filled into a capsule, formulated as a powder, or suspended in a liquid.

[0054] The term "effective amount" of a drug, as used herein, is an amount sufficient to bring about a beneficial or desired result in a patient, such as disease remission, and such an "effective amount" will depend on the context in which it is applied, including the age and weight of the patient, the nature of the disease, including the organ(s) affected by the disease, the state or level of activity of the disease, the patient's sensitivity to cysteamine, and other factors.

[0055] As used herein, "pantetheine," "4-phosphopantetheine," "dephospho-coenzyme A," and "coenzyme A," as well as any analogs or derivatives convertible to one of these compounds in the gastrointestinal tract, all refer to the D enantiomer (sometimes also referred to as the R enantiomer, using more recent nomenclature). Each of these compounds contains a chiral carbon in the pantothenoyl moiety, which can exist in either the D (dextro) or L (levo) form, also referred to as the (R) or (S) form, respectively. The D-pantetheine enantiomer is the only pantetheine enantiomer that is a substrate for pantetheinase and, therefore, a cysteamine precursor. Similarly, only the D-enantiomers of compounds convertible to pantetheine, such as 4-phosphopantetheine, dephospho-coenzyme A, and coenzyme A, are useful in the compositions and methods of the present invention.

[0056] As used herein, a "disulfide compound" is a compound containing a sulfur atom chemically bonded to a second sulfur atom in the form R1-SS-R2, where R1 and R2 are organic compounds. R1 and R2 can be the same or different. Disulfide compounds are generally formed by the oxidation of two thiols (i.e., R1-S-H+R2-SH to R1-SS-R2+2H). + ) which can be reversibly converted to two thiols by reduction (i.e., R1-SS-R2+2H + gives R1-S-H+R2-SH). Disulfide compounds can also be formed by reacting one or two thiols with a dithiol (e.g., R1-S-H+R2-S-H+HS-R3-SH gives R1-SS-R3-SS-R2+4H). + where R1, R2, and R3 are organic compounds and H + is a hydrogen ion). The disulfide compounds of the present invention include 1) cysteamine mixed disulfide compounds of the formula C2H6NS-S-R1, where R1 is an organic moiety; 2) cysteamine mixed disulfide compounds of the formula C 11 H 21 3) pantetheine disulfide compounds of formula C 11 H 22 4) 4-phosphopantetheine disulfide compounds of formula N2O7PS-S-R1 (wherein R1 is an organic moiety); 21 H 34 N7O 13 5) dephosphocoenzyme A disulfide compounds of formula P2S-S-R1 (wherein R1 is an organic moiety); 21 H 35 N7O 16Bioactive sulfur-containing compounds include coenzyme A disulfide compounds of the formula PS-S-R (where R is an organic moiety), or 6) N-acetylcysteamine compounds of the formula CHNOS-S-R (where R is an organic moiety). Additional disulfides can be formed using dithiols, which are compounds capable of forming two disulfide bonds. At least one, and optionally both, disulfide bonds are with cysteamine or a compound that can degrade to cysteamine in the gastrointestinal tract. Alternatively, the dithiol is disulfide-linked to only one such compound, and the second thiol of the dithiol remains in thiol form, or the second thiol can be disulfide-linked to any thiol, including, for example, any of the thiols listed in Figure 17. In addition to pantetheine, compounds that can be degraded to cysteamine in the gastrointestinal tract include 4-phosphopantetheine, dephosphocoenzyme A, coenzyme A, or N-acetylcysteamine, or any analog or derivative that can be converted to one of these five compounds in the gastrointestinal tract (e.g., by chemical or enzymatic processes). Any such analog or derivative, referred to herein as a "suitable analog or derivative," is a thiol of the present invention and can be substituted for one of these five compounds. A "mixed disulfide" is a disulfide formed from two different thiols. A "cysteamine mixed disulfide" refers to a disulfide connecting cysteamine with another (non-cysteamine) thiol. A "pantetheine mixed disulfide" refers to a disulfide connecting pantetheine with another (non-pantetheine) thiol, and so forth. Generally, mixed disulfides are classified by the simpler of the two constituent thiols (e.g., cysteamine-pantetheine is called a cysteamine mixed disulfide). Thiols useful for forming disulfide cysteamine precursors include, for example, L-cysteine, N-acetylcysteine, glutathione, any of the thiols listed in Figure 17, and other thiols described herein. Some exemplary mixed disulfides are shown in Figures 2-10.The tables in Figures 18-21 show how the thiols in Figure 17 can be usefully combined to form disulfides. For brevity and clarity, the names of the two thiols connected via a disulfide bond are used herein to indicate the disulfide name rather than the formal chemical name (e.g., using IUPAC nomenclature). Thus, cysteamine-pantetheine refers to the disulfide formed from these two compounds. Three important exceptions to that rule are: a disulfide formed by reacting two pantetheines is commonly called pantethine, a disulfide formed by reacting two cysteines is commonly called cystine, and a disulfide formed by reacting two cysteamines is commonly called cysteamine.

[0057] As used herein, the terms "disulfide formed by reacting ..." or "compound formed by reacting ..." refer specifically to disulfides formed between two specified thiols. For example, the disulfide formed by reacting cysteamine with pantetheine (referred to as cysteamine-pantetheine) refers to a heterodimer formed between a cysteamine molecule and a pantetheine molecule. This definition does not reflect what actually occurs when two specified thiols react. That is, when cysteamine reacts with pantetheine under oxidizing conditions, three disulfides can be formed in varying proportions depending on the chemical conditions: cysteamine-cysteamine (i.e., cystamine), cysteamine-pantetheine (also pantetheine-cysteamine, which is the same for purposes of this invention), and pantetheine-pantetheine (i.e., pantethine). When the actual reaction product (i.e., a mixture of three disulfides) is meant, the text clearly states so.

[0058] "Cysteamine precursor" refers to a compound that can be converted to at least one cysteamine under physiological conditions. Means of conversion include reduction in the case of cysteamine-containing disulfides (i.e., cysteamine mixed disulfides), enzymatic hydrolysis in the case of pantetheinase substrates (pantetheine and compounds metabolically convertible to pantetheine in the gastrointestinal tract, such as 4-phosphopantetheine, dephosphocoenzyme A, and coenzyme A, and suitable analogs or derivatives thereof), or both reduction and enzymatic cleavage. Examples of precursors include, but are not limited to, cysteamine mixed disulfides, pantetheine disulfide, 4-phosphopantetheine disulfide, dephosphocoenzyme A disulfide, coenzyme A disulfide, and N-acetylcysteamine disulfide, as well as pantetheine, 4-phosphopantetheine, dephosphocoenzyme A, coenzyme A, and N-acetylcysteamine. The chemical relationships among cysteamine, pantetheine, 4-phosphopantetheine, dephospho-Coenzyme A, and Coenzyme A (the latter four compounds are cysteamine precursors) are shown below: Because the constituent thiols are all cysteamine precursors, homodimers of two pantetheine molecules (i.e., pantethine), or two 4-phosphopantetheine molecules, or two dephospho-Coenzyme A molecules, or two Coenzyme A molecules, or two N-acetylcysteamine molecules are each also disulfide cysteamine precursor compounds.

[0059] By "suitable analogue or derivative" is meant, with respect to the cysteamine precursors pantetheine, 4-phosphopantetheine, dephosphocoenzyme A, coenzyme A, or N-acetylcysteamine, or disulfides containing any of them, a compound that can be converted to pantetheine, 4-phosphopantetheine, dephosphocoenzyme A, coenzyme A, or N-acetylcysteamine in the gastrointestinal tract, whether by chemical or enzymatic processes.

[0060] "Compounds convertible to pantetheine" means compounds such as 4-phosphopantetheine, dephospho-coenzyme A, and coenzyme A that can be broken down to pantetheine in the gastrointestinal tract, as well as analogs or derivatives of compounds that can be converted to the parent compound in the gastrointestinal tract.

[0061] "Constituent thiol" when used in reference to a disulfide means a thiol (and optionally a dithiol) compound that reacts to form a disulfide.

[0062] "Cysteamine content" means the weight fraction of cysteamine precursors that can be converted to cysteamine in vivo upon chemical and / or enzymatic degradation.

[0063] The term "pharmaceutically acceptable salt," as used herein, refers to a salt that, within the scope of sound medical judgment, is suitable for use in contact with the tissues of humans and animals without undue toxicity, irritation, allergic response, and the like, and that is commensurate with a reasonable benefit / risk ratio. Pharmaceutically acceptable salts are well known in the art. For example, pharmaceutically acceptable salts are described in Berge et al., J. Pharmaceutical Sciences 66:1-19, 1977, and Pharmaceutical Salts: Properties, Selection, and Use (eds. P.H. Stahl and C.G. Wermuth), Wiley-VCH, 2008. The salts can be prepared in situ during the final isolation and purification of the compounds of the invention, or separately by reacting the free base group with a suitable organic or inorganic acid. Representative acid addition salts include acetate, adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, bitartrate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecyl sulfate, ethanesulfonate, fumarate, glucoheptonate, glycerophosphate, hemisulfate, heptanoate, hexanoate, hydrobromide, hydrochloride, hydroiodide, and 2-hydroxyethanesulfonate. Examples of the salts include phosphate, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, pamoate, pectinate, persulfate, 3-phenylpropionate, phosphate, picrate, pivalate, propionate, stearate, succinate, sulfate, tartrate, thiocyanate, toluenesulfonate, undecanoate, and valerate.Representative alkali or alkaline earth metal salts include sodium, lithium, potassium, calcium, magnesium, and the like, as well as non-toxic ammonium, quaternary ammonium, and amine cations, including, but not limited to, ammonium, tetramethylammonium, tetraethylammonium, methylamine, dimethylamine, trimethylamine, triethylamine, ethylamine, and the like.

[0064] "Gastroretentive," "gastric retention," and the like refer to a pharmaceutical composition that can remain in the stomach of a mammal, preferably a human, for an extended period of time, preferably the same length as, and more preferably longer than, that of food. Thus, "gastric retention" refers to the maintenance of a drug composition in the stomach for a period longer than the period of retention in the stomach when delivered in free form, e.g., in an oral delivery vehicle that is not considered gastric-retentive. A gastric-retentive formulation can be characterized by gastric retention for a period longer than the normal emptying time from the stomach, i.e., greater than about 2 hours, particularly greater than about 3 hours, and usually greater than about 4, 6, 8, or 10 hours. A gastric-retentive formulation typically remains in the stomach for about 3, 4, 6, 8, 10, or sometimes 18 hours or more after ingestion of a meal. However, it should be noted that, according to the present invention, retention of a controlled-release gastric-retentive drug delivery system is not observed after 48 hours or more, preferably 24 hours, of administration in the non-fasted state. Gastroretentive formulations include floating or buoyant formulations, swelling or expanding formulations, bioadhesive or mucoadhesive formulations, unfolding formulations, and magnetic formulations, or any combination thereof. Because it has proven difficult to maintain gastric retention with only one gastric retention mechanism, combinations of two or more gastroretentive formulations are common. Gastroretentive formulations are preferably administered with a meal.

[0065] The terms "floating," "floating," and "buoyant," used interchangeably, refer to a type of formulation capable of positioning the compositions of the present invention on or near the surface of gastric contents, which are chyme-like fluids in the fed state (gastric fluids in the fasted state or after gastric emptying). By floating on the gastric contents, the formulation has less opportunity to be propelled through the pylorus into the duodenum during contractions of the stomach muscles, which are seated or upright at the fundus of the stomach. Floating formulations may be composed of small (e.g., micron-scale), medium (e.g., millimeter-scale), or large (e.g., centimeter-scale) particles. Large compositions may simultaneously act via a swelling / distensible mechanism, as described herein. Formulations of any size may simultaneously act via a mucoadhesive mechanism.

[0066] The terms "swellable" and "expandable," used interchangeably, refer to the ability of a composition to increase in size upon contact with a fluid-containing medium, such as gastric juice or chyme. Preferably, "swellable" is characterized by an increase in the initial tablet size to a size that will not be easily removed from the stomach. Gastric emptying occurs through the pylorus. The average resting diameter of the pylorus in humans varies between fed and fasted states. In the fed state, it is about 1 centimeter or less, and in the fasted state, it is about 1.28 centimeters ± 7 millimeters. Preferably, "swellable" requires the composition to increase in size by 14 mm or more, 16 mm or more, 18 mm or more, 20 mm or more, or 22 mm or more in at least two dimensions, or alternatively, if only one dimension is so, to greater than 12 mm, 14 mm, or 16 mm in both the second and third dimensions.

[0067] "Mucooadhesion" refers to the ability of a composition to adhere to the mucus layer lining the gastrointestinal tract. In the case of gastroretentive formulations, "mucoadhesion" refers to adhesion to the mucus layer lining the stomach. Mucoadhesion is one of several techniques for extending gastric retention time, but the mucous layer of the stomach undergoes slow but continuous turnover, limiting the duration of mucoadhesion. Therefore, mucoadhesion is usually combined with other gastric retention methods to achieve prolonged gastric retention time. "Bioadhesion" refers to the ability of a composition to adhere to other molecules lining the gastrointestinal tract, including molecules on the surface of enterocytes.

[0068] "Unfolded" or "shape-changing," used interchangeably, refers to the ability of a composition to unfold, stretch, uncoil, relax, or otherwise unfold in the stomach, transforming into a composition of a size and / or geometry that does not readily pass through the pylorus and therefore remains in the stomach for an extended period of time. An "unfolded" or shape-changing formulation can be formulated in a capsule. Ideally, but not necessarily, the dimensions of the unfolded or unpackaged unfolded formulation are greater than 16 mm, 18 mm, 20 mm, or 22 mm in at least two dimensions, but alternatively, if only one dimension is so, the second and third dimensions are 12 mm, 14 mm, or 16 mm or greater.

[0069] "Magnetic formulation" refers to a composition containing a magnet or disseminated magnetized material that can interact with an externally applied magnetic field created by magnet(s) placed outside the body to cause retention of the composition in the stomach or small intestine for an extended period of time. Compositions targeted to the stomach preferably remain in the stomach for at least as long as food remains in the stomach, and more preferably longer than food does. Compositions targeted to the small intestine preferably remain in the stomach until substantially complete drug dissolution or until adequate magnetic strength is lost to hold the composition in place, whichever occurs first. The magnet or magnetic material used must be safe for human ingestion. Although external magnets can also be used to place pharmaceutical compositions containing magnets in other areas of the gastrointestinal tract, such as the colon, in most cases, magnetic formulations are a type of gastroretentive or small intestine-targeted formulation.

[0070] As used herein, a "therapeutically effective amount" refers to the amount that must be administered to a patient (human or non-human mammal) to ameliorate disease or modulate a biomarker surrogate for disease activity. Clinical endpoints for various diseases, including neurodegenerative, metabolic, fibrotic, ischemic, infectious, neoplastic, and genetic disorders, vary widely but are generally well known in the art. Specific biomarkers include, for example, (i) white blood cell (WBC) cystine levels, which are a surrogate for disease control in patients with cystinosis; (ii) Clinical Global Assessment (CGI) score, Clinician Interview-Based Assessment of Change and Caregiver Input (CIBIC-Plus), Alzheimer's Disease Cooperative Study-Clinician Global Assessment of Change (ADCS-CCGIC) score, Alzheimer's Disease Assessment Scale-Cognitive Subscale (ADAS-Cog) score, Alzheimer's Disease Cooperative Study-Modified Activities of Daily Living for Severe Dementia (ADCS-CCGIC) score, and the Alzheimer's Disease Cooperative Study-Modified Activities of Daily Living for Severe Dementia (ADCS-CCGIC). (iii) Indicators of cognitive, motor, or emotional status, including measures such as the ADCs-ADLsev score, Mini-Mental State Examination (MMSE), Neuropsychiatric Inventory (NPI) score, Unified Huntington's Disease Rating Scale (UDHRS), MATTIS test, Hopkins Trail Making Test, category fluency task, Unified Parkinson's Disease Rating Scale (UPDRS) score, or Parkinson's Disease Sleep Scale-II (PDSS-II) total score, can be used to measure treatment response in patients with neurodegenerative diseases. (iv) Biochemical measures of neurodegenerative disease activity include AD biomarkers (e.g., plasma beta-amyloid protein) or brain-derived neurotrophic factor (BDNF) levels. (iv) Indicators of metabolic and fibrotic liver disease include anatomical tests such as the NAFLD activity score (NAS) and liver biopsy-based liver fibrosis scores. (v) biochemical indicators of liver health including hepatic and adipose tissue insulin sensitivity, as measured by HOMA-IR and Adipo-IR indices, respectively, serum aminotransferase and gamma-glutamyl transpeptidase (GGT) levels, and CK-18-derived fragments in blood in relation to NAFLD, NASH, ASH, or inherited liver disease.(vi) Indicators of disease status related to mitochondrial disease include the Newcastle Pediatric Mitochondrial Disease Scale (NPMDS) score as a clinical endpoint. And (vii) biomarkers include levels of glutathione, total serum thiol, acetoacetate, beta-hydroxybutyrate, lactate, or malondialdehyde (a marker of oxidative stress). Other surrogate disease markers include modulation of immune response, modulation of gene or protein expression, or modulation of confirmed radiological disease measures (e.g., assessed by X-ray, CT scan, MRI scan, or PET scan). Methods for determining therapeutically effective amounts of cysteamine precursors are highly disease-specific and are well known to clinicians specializing in each of the above diseases.

[0071] As used herein, a "pharmaceutically acceptable excipient" is a natural or synthetic substance that may be included (together with an active ingredient) in the formulation of a composition suitable for use in humans and / or non-human mammals without undue adverse side effects (such as toxicity, irritation, or allergic reactions). Excipients may include, for example, antiadherents, antioxidants, binders, coatings, compression aids, disintegrants, dyes (colors), emollients, emulsifiers, fillers (diluents), film-forming or coating agents, flavors, fragrances, glidants (glidants), lubricants, preservatives (including antioxidants), printing inks, adsorbents, suspending or dispersing agents, solvents, colloidal stabilizers, sweeteners, and water. The US FDA maintains a database of "inactive ingredients" containing information on thousands of substances commonly used in drug formulations. This database can be searched for excipients commonly used in controlled-release, delayed-release, sustained-release, or extended-release formulations. Excipients include, but are not limited to, butylated hydroxytoluene (BHT), calcium carbonate, calcium phosphate (dibasic), calcium stearate, carbomer, croscarmellose, cross-linked polyvinylpyrrolidone, citric acid, crospovidone, cellulose derivatives including ethylcellulose, hydroxypropylcellulose, hydroxypropylmethylcellulose, or hypromellose, docusate sodium, gelatin, gelucire 43 / 01, lactose, magnesium stearate, maltitol, mannitol, methylcellulose, methylparaben, microcrystalline cellulose, polyethylene glycol, poly(ethylene oxide), polyvinylpyrrolidone, povidone, pregelatinized starch, propylparaben, shellac, silicon dioxide, sodium carboxymethylcellulose, sodium starch glycolate, sorbitol, starch (corn), stearic acid, sucrose, talc, titanium dioxide, vegetable oils, waxes including white, yellow, or beeswax, and xylitol.Excipients also include diluents (e.g., saline and aqueous buffer solutions), aqueous carriers, and non-aqueous carriers, such as water, ethanol, polyols (e.g., glycerol, propylene glycol, polyethylene glycol, etc.), and suitable mixtures thereof, vegetable oils such as olive oil, and injectable organic esters such as ethyl oleate. Excipients useful for formulating compositions with specific properties are described more specifically in the detailed description of the invention.

[0072] "Enteric coating" means an agent or compound added to the formulations described herein that protects the active ingredient(s) described herein (e.g., cysteamine precursors and enhancers of cysteamine precursor degradation and absorption) as they pass through the stomach. The enteric coating also protects the stomach from irritating pharmaceutical ingredients (e.g., cysteamine). Examples of commercially available enteric coating technologies include, but are not limited to, AcrylEZE, Opadry, Nutrateric, and Sureteric products (Colorcon, West Point PA), Advantia Performance Specialty Coatings (International Specialty Products, Wayne NJ), the Kollicoat product line (BASF Corporation, Ludwigshafen Germany), Aquacoat products (FMC BioPolymer), Eastman CAP (Eastman Chemical Co. Kingsman TN), the Eudragit product line (Evonik Industries), and the AQOAT, HP-50, and HP-55 product lines (Shin Etsu Pharma). Ashland Specialty Ingredients, Encap Drug Delivery, and Sanyo Chemical Industries, Ltd. also sell enteric coating systems.Examples of pH-sensitive film-forming polymers commonly used in enteric coated formulations include: (i) cellulosic polymers such as cellulose acetate phthalate (e.g., Aquacoat CPD, FMC; CAP, Eastman Chemical Co.), cellulose acetate succinate, cellulose acetate trimellitate, hydroxypropyl methylcellulose phthalate, hydroxypropyl methylcellulose acetate succinate (e.g., AquaSolve, Ashland Specialty Ingredients, Wilmington DE); (ii) polymethacrylates such as poly(methacrylic acid-ethyl acrylate) in 1:1 and 1:2 ratios (e.g., Eudragit L30D-55 and Eudragit L100-55 (Evonik Industries), AcrylEZE (Colorcon), Kollicoat MAE 30 DP and Kollicoat MAE 100P (BASF Pharma Ingredients and Services), Polyquid PA-30 (Sanyo Chemical (iii) polyvinyl derivatives such as poly(vinyl acetate) phthalate (e.g., Sureteric, Colorcon), and (iv) other copolymers such as half-esters of copolymers of styrene and maleic acid, half-esters of copolymers of vinyl ether and maleic acid, and copolymers of vinyl acetate and crotonic acid. Enteric coatings are also made using shellac (e.g., PROTECT, Sensient Pharmaceutical Coating Systems) or sodium alginate and zein (Encap Drug Delivery). Hydroxypropyl methylcellulose is also called hypromellose or HPMC. Examples of other excipients commonly used in enteric coating formulations include wet microcrystalline cellulose, wet powdered cellulose, gellan gum, and stearic acid. Enteric coatings can be applied to a variety of formulations, including tablets, capsules, and microparticles.

[0073] As used herein, "combination therapy" means that a patient (or non-human mammal) in need of treatment according to the present invention receives medications in addition to those disclosed herein that are not fully described or possibly contemplated herein. Combination therapy can be administered sequentially (before or after) or simultaneously with the cysteamine precursor therapy of the present invention.

[0074] "Treating" means treating a disease or disorder and subjecting a patient to a management regimen for the purpose of obtaining beneficial or desired results, such as amelioration of disease signs or symptoms, or improvement of biochemical, radiological, behavioral, or physical markers of disease activity or disease state. Examples of beneficial or desired results can include, but are not limited to, resolution of inflammation, resolution of biochemical imbalances, improvement in quality of life, improvement in cognitive and behavioral status, improvement in motor function, improvement in emotional and mood status, improvement in sleep, or more generally, alleviation or amelioration of one or more symptoms or conditions, reduction in the extent of disease, stabilization of the disease state, prevention of disease spread, delay or slowing of disease progression, improvement or alleviation of the disease, disorder, or condition, and partial or complete remission of significant disease manifestations.

[0075] The term "mammal" is intended to mean both human and non-human mammals.

[0076] By "delivering" is meant providing and / or administering an active ingredient(s) described herein by oral administration of a tablet, capsule, liquid, powder, granules, particulates, sachet, suppository, etc. (collectively referred to as "pharmaceutical composition" or simply "composition") containing the active ingredient(s) and (optionally) one or more carriers and / or diluents and / or adjuvants or other excipients. The composition may be provided with instructions for delivery, including any color-coding or alphanumeric text description on the surface or packaging of the composition, as well as instructions regarding whether the composition should be taken at a specific time or with food (e.g., (specific types or amounts of food), liquid, meal (including details regarding type of meal), or other medication), and whether the patient should remain upright or seated for a period of time following drug administration.

[0077] Some disease acronyms, gene names, and other medical terms are represented by abbreviations. Disease acronyms include MELAS (mitochondrial encephalocardiopathy, lactic acidosis, and stroke-like episodes) and MERFF (myoclonic epilepsy with ragged-red fibers). Gene names include POLG (DNA polymerase gamma, encoding the catalytic subunit of mitochondrial DNA polymerase); OCT1, OCT2, and OCT3 (encoding organic cation transporters 1, 2, and 3) (also known as SLC22A1, SLC22A2, and SLC22A3, respectively); PANK2 (encoding pantothenate kinase 2); VNN1 (encoding vanin 1, also known as pantetheinase); VNN2 (encoding vanin 2, also known as GPI-80 and pantetheinase).

[0078] As used herein, "cysteamine-sensitive disease" refers to a disease for which there is evidence that cysteamine may be an effective treatment. Evidence can be derived from either clinical or preclinical studies of the disease in mammals (e.g., humans, dogs, mice, rats, monkeys, rabbits), or from in vitro studies of disease mechanisms. Cysteamine-sensitive diseases comprise a broad and heterogeneous group of diseases with widely varying symptoms and etiologies. Diseases and disorders for which there is evidence of cysteamine effectiveness can be classified by etiology, with the important caveat that the mechanism of cysteamine effectiveness is not always clear and may have an unknown mechanism of action. Important categories of cysteamine-sensitive diseases include: (i) disorders of cystine transport, of which cystinosis is the most well-known; (ii) disorders associated with oxidative damage, including neurodegenerative and liver diseases; (iii) disorders associated with pathological enzyme activity, including neurodegenerative diseases, inherited mitochondrial diseases, and diseases associated with mutant MECP2 and POLG; (iv) fibrotic disorders, including fibrosis of the kidney, liver, or lung; (v) metabolic disorders, including metabolic syndrome X, diabetes, and the spectrum of non-alcoholic fatty liver disease leading to non-alcoholic steatohepatitis (NASH); (vi) infectious diseases, including certain viral infections (e.g., influenza), bacterial infections (e.g., Pseudomonas aeruginosa), and parasitic infections (e.g., malaria); (vii) ischemic diseases, including ischemia-reperfusion injury of the heart and other organs; (viii) diseases associated with abnormal adiponectin metabolism; and (ix) amelioration of the adverse effects of cancer and cancer treatments.

[0079] As used herein, the term "about" means ±20% of the recited value. [Brief explanation of the drawings]

[0080] [Figure 1]

[0049] Figure 11 shows the chemical structure of Coenzyme A, from which dephospho-Coenzyme A, 4-phosphopantotheine, pantetheine, pantothenic acid, or cysteamine molecules can be derived by enzyme-catalyzed reactions (shown in Figure 11). [Figure 2]Two chemical structures of disulfides of the present invention are shown. The top chemical structure shows a mixed cysteamine disulfide molecule, with cysteamine on the left and a second thiol (designated RS-) on the right. The bottom chemical structure shows pantetheine disulfide, with pantetheine on the left and a second thiol (designated RS-) on the right. Figures 3, 4, and 5 show exemplary mixed cysteamine disulfides. As shown schematically in Figures 18 and 21, other mixed cysteamine disulfides can be formed with the thiols listed in Figure 17. [Figure 3] Four chemical structures of cysteamine mixed disulfides are shown. Specifically, mixed cysteamine disulfide with the partner thiol allyl mercaptan, L-cysteine, L-cysteine ​​ethyl ester, and N-acetylcysteine ​​are shown, as indicated on the labels. [Figure 4] Two exemplary chemical structures of cysteamine mixed disulfides and one exemplary N-acetylcysteamine mixed disulfide are shown. Two cysteamine mixed disulfides are formed between cysteamine and N-acetylcysteamine, and between cysteamine and N-acetylcysteinamide. Also shown (labeled) is the mixed disulfide formed between N-acetylcysteamine and N-acetylcysteinamide. [Figure 5] As shown on the label, two chemical structures of exemplary cysteamine mixed disulfides are shown: those formed between cysteamine and pantetheine, and those formed between cysteamine and glutathione. [Figure 6] 1 shows the chemical structure of an exemplary cysteamine mixed disulfide formed between cysteamine and coenzyme A. [Figure 7] Two chemical structures are shown: (a) an exemplary pantetheine mixed disulfide formed between pantetheine and cysteine; (b) an exemplary N-acetylcysteamine mixed disulfide formed with pantetheine. [Figure 8]The chemical structures of two exemplary mixed disulfides are shown, one formed between pantetheine and N-acetylcysteine, and the other formed between dithiol dihydrolipoic acid and two cysteamines (one disulfide bonded to each of the two thiols of dihydrolipoic acid), as indicated on the label. [Figure 9] 1 shows the chemical structure of an exemplary pantetheine mixed disulfide formed between pantetheine and glutathione. [Figure 10] 1 shows the chemical structure of an exemplary 4-phosphopantetheine mixed disulfide formed between 4-phosphopantetheine and coenzyme A. [Figure 11]

[0023] Figure 1 is a schematic diagram of a portion of the coenzyme A, pantetheine, and cysteamine metabolic pathway, including both intracellular metabolism (solid lines) and catabolic reactions (dotted lines) occurring in the gastrointestinal tract. Some reactions occur in both locations (e.g., phosphatases are present in the cytoplasm and the gastrointestinal tract). Compounds are shown in regular type, and enzymes are shown in italics. Both compounds and enzymes have various alternative names for those shown in the diagram. This diagram is not a complete depiction of coenzyme A, pantetheine, and cysteamine metabolism, but is simply intended to convey that coenzyme A, dephosphocoenzyme A, 4-phosphopantetheine, and pantetheine can be catabolized to cysteamine (and pantothenate) in the intestine. [Figure 12]The anatomy of the gastrointestinal (GI) tract is shown in schematic form (above). Below is a table summarizing certain anatomical and physiological parameters for each segment of the GI tract related to the in vivo production and uptake of cysteamine from the cysteamine precursors of the present invention. In particular, the table indicates the anatomical sites where cysteamine formation and uptake occur, as well as the levels of physiological variables that affect the in vivo rate of cysteamine production from cysteamine precursors (e.g., via disulfide bond reduction and pantetheinase cleavage) and the rate of cysteamine absorption along the GI tract (e.g., via organic cation transporters 1, 2, and 3). For example, pH affects disulfide exchange reactions. Glutathione (GSH) levels are a proxy for the redox environment, affecting the equilibrium between oxidized and reduced disulfides and thiols, including the reduction of disulfide cysteamine precursors. The absorption surface area and transit time, along with the levels of pantetheine digestive enzymes and cysteamine transporters, affect the rate of cysteamine production from pantetheine and subsequent cysteamine absorption. Other physiological variables in this diagram affect the performance of certain types of formulations. For example, some types of gastroretentive formulations swell to a size that prevents them from passing through the pylorus. Some pH-sensitive pharmaceutical coatings dissolve in the duodenum at pH 5.5, pH 6, or near pH 6.5, while other coatings dissolve near pH 7, more typical of the ileum. Some types of colon-targeted formulations consist, in part, of polymers that are refractory to digestion by human (or mammalian) enzymes but can be degraded by enzymes produced by intestinal bacteria, thereby resulting in the release of cysteamine precursors co-formulated with the polymer. The values ​​or ranges provided in the table are derived from literature sources and may not encompass the full range of normal human variation. Nevertheless, the degree of variation shown may, in part, explain the wide interindividual variation in cysteamine uptake and metabolism observed clinically. [Figure 13]This table shows the classification of cysteamine precursors and some of their notable pharmacological properties. Cysteamine precursors are classified on the left (bottom) side of the table according to (i) whether they are thiols or disulfides; (ii) if they are disulfides, whether they are cysteamine-containing mixed disulfides (including cysteamine-pantetheine), pantetheine-containing mixed disulfides (excluding cysteamine-pantetheine), or contain other thiols that can be degraded to pantetheine in the gastrointestinal tract; and (iii) how many cysteamines are generated (under the # symbol) upon chemical reduction and / or enzymatic degradation. "Other thiols or dithiols" refers to any dithiol, including any thiol that is not cysteamine, pantetheine, 4-phosphopantetheine, dephosphocoenzyme A, coenzyme A, or N-acetylcysteamine. (See Figure 17 for exemplary thiols and dithiols.) While the decomposition of disulfide cysteamine precursors containing "other thiols" produces only one cysteamine, disulfide cysteamine precursors containing dithiols can decompose to produce one or two cysteamines, since one dithiol can, for example, bond to two cysteamines (see Table 21 for a summary of how thiols and dithiols can be combined). The table further indicates, under "Steps to Generate Cysteamine," which chemical and / or enzymatic steps are required to generate cysteamine from each class of cysteamine precursor. For example, cysteamine mixed disulfides containing cysteamine plus another thiol (e.g., cysteine) require only one step, disulfide bond reduction. Similarly, thiol pantetheine requires only one step, pantetheinase cleavage. Other cysteamine precursors require two steps. For example, the pantetheine homodimer pantethine requires disulfide bond reduction followed by pantetheinase cleavage. Still other cysteamine precursors require more than two steps, for example, 4-phosphopantetheine homodimer requires disulfide bond reduction, phosphatase cleavage, and pantetheinase cleavage.Dephosphocoenzyme A and disulfides containing coenzyme A require additional steps. For some disulfide cysteamine precursors, the number of degradation steps to cysteamine differs between the two thiols produced by disulfide bond reduction, as shown in the table. Furthermore, the table indicates classes of compounds that can be coformulated or coadministered with cysteamine precursors to enhance in vivo cysteamine production, and indicates which class(es) of accelerators are useful for each class of cysteamine precursor. For example, to enhance disulfide bond reduction, any disulfide cysteamine precursor can be productively coformulated or coadministered with a reducing agent (abbreviated as RA in the table). Cysteamine precursors that are or contain pantetheine, or any thiol that can be degraded to pantetheine, can be productively coformulated or coadministered with an inducer of the enzyme pantetheinase (abbreviated as PI in the table). Pantetheine disulfide can be productively coformulated or coadministered with both a reducing agent and a pantetheinase inducer. Enhancers of cysteamine absorption (e.g., inducers of cysteamine transporters, such as organic cation transporters) or inhibitors of cysteamine catabolism are not shown in the table, because such compounds can be productively coformulated or coadministered with all classes of cysteamine precursors. At the far right (top), the table summarizes in a few words the salient pharmacological properties of different classes of cysteamine precursors, which may be affected by the number of degradative steps required to produce cysteamine, the yield of cysteamine, or the presence of enhancers of in vivo cysteamine production. The very brief descriptions provided are not exhaustive and should not be construed as limiting. [Figure 14]1 is a diagram of an exemplary pharmaceutical composition. Salient characteristics of the exemplary composition are shown, including: (i) type of dosage form (e.g., tablet, capsule, powder, liquid), (ii) characteristics of the formulation with respect to anatomical localization of drug release (e.g., gastroretentive formulations remain in the stomach; enteric-coated formulations may be designed to release drug in the small intestine; colon-targeted formulations are designed to release drug in the ileum or colon), and (iii) duration of drug release (immediate release: IR, or sustained release: SR), (iv) type of cysteamine precursor(s), (v) dose (provided as a range), (vi) dosage (provided as a range), (vii) dosage (provided as a range), (viii) dosage (provided as a range), (viiii) duration of drug release (immediate release: IR, or sustained release: SR), (viii) dosage (provided as a range), (viiii) duration of drug release (immediate release: IR, or sustained release: SR), (viiii) duration of drug release (immediate release: IR, or sustained release: SR), (viiii) type of cysteamine precursor(s), (viiii) dosage (provided as a range), (viii) dosage (provided as a range), (viiii) duration of drug release (immediate release: IR, or sustained release: SR ... (vi) type of co-formulated enhancer(s) of in vivo cysteamine production, if any; (vii) dose of enhancer compound (provided as a range); (viii) recommendation for administering the composition with food (e.g., applesauce or yogurt) or a meal (e.g., dinner) or whether food is optional ("food OK"); (ix) site(s) of cysteamine precursor release within the gastrointestinal tract; and (vii) site(s) where cysteamine is generated in vivo (e.g., by disulfide bond reduction or pantetheinase cleavage). The compositions in Figure 13 are each limited to a single type of formulation with respect to site and time of drug release. Such compositions (including many variants not shown) can be administered in various combinations, providing flexibility for individualizing administration. Other exemplary compositions with more active ingredients and / or more complex formulations are shown in Figures 14 and 15. [Figure 15] Figure 1 shows exemplary pharmaceutical compositions with (i) one or two drug release profiles (e.g., composition G includes an immediate-release component and an extended-release component), (ii) at least two cysteamine precursor(s) and up to two enhancers. Recommendations for administration with or without food are provided, as are site(s) of drug release and in vivo conversion of cysteamine precursors to cysteamine. The exemplary compositions, and many other compositions not shown, can be combined in various ratios. [Figure 16]

[0033] Figures 14 and 15 show exemplary multiple-dose regimens in which two or more compositions are administered together or sequentially at short intervals. Salient features of exemplary compositions are shown in Figures 14 and 15. Examples include compositions that provide an enhancer (e.g., a reducing agent) for cysteamine precursor degradation, but not the cysteamine precursor. Separate formulation of enhancers allows them to be co-administered with the cysteamine precursor-containing composition in various ratios to optimize in vivo cysteamine production or uptake. Separate formulation of enhancers further allows for control of the site and timing of enhancer release to optimize in vivo cysteamine production or uptake. [Figure 17] Figure 17 shows a list of exemplary thiols and dithiols that can be combined to produce thiol- or disulfide-type cysteamine precursors (compounds 2-6). The chemical formula, Chemical Abstracts Service (CAS) registry number, and formula molecular weight of each thiol or dithiol are provided. In some cases, the CAS number is specific to a particular enantiomer. To facilitate concise reference of these thiols in Figures 18-21, each thiol is numbered (leftmost column of Figure 17). [Figure 18]Two tables are included that show how the thiols and dithiols in Figure 17 can be combined to create two classes of disulfide cysteamine precursors: cysteamine mixed disulfides and pantetheine disulfides. The five columns in each of the two tables list, from left to right: (i) The two thiols that react to form a disulfide are indicated by a number in the leftmost column of Figure 17 (thiols are numbered 1-29, dithiols are numbered 30-35). Thus, for example, the notation "1 + 28" represents the disulfide formed by reacting thiol 1 (cysteamine) with thiol 28 (tiopronin). All disulfides in the left table contain cysteamine (compound 1) plus a second thiol (any of compounds 2-35). All disulfides in the right table contain pantetheine (compound 2) plus a second thiol (any of compounds 2-35). (ii) Formula molecular weight (MW) of the disulfide shown in the first column. For example, the MW of disulfide 1+28 is 238.35 Daltons (the sum of the masses of the two constituent thiols minus 2 to account for the two missing protons). Note that for thiols 13 and 14 (L-cysteine ​​ethyl ester HCl and L-cysteine ​​methyl ester HCl), the mass of the salt form is used. The actual mass of the free disulfide is 36.46 Daltons less than the indicated mass. (iii) The number of cysteamines that can be produced upon degradation of the cysteamine precursor in vivo. Disulfides are sorted into those that yield two cysteamines, listed above the horizontal bold line, and those that yield one cysteamine, listed below. (iv) The fraction of the cysteamine precursor that can be converted to free cysteamine in vivo. For example, the fraction of disulfide 1+28 that can be converted to the 238.35 Dalton cysteamine is 32.4%. The disulfides that yield one cysteamine are ranked from highest to lowest by the fraction of their molecular weight that can be converted to cysteamine. (v) The number of decomposition steps (chemical or enzymatic) required to obtain cysteamine from the disulfide cysteamine precursor.For disulfides above the horizontal bold line where both thiols are degradable to cysteamine (or one of the two thiols is cysteamine itself), two numbers are shown, indicating the number of steps for each thiol component of the disulfide. The order of the two numbers corresponds to the order in which the two thiols are listed in the first column of the table. For disulfides where only one of the thiols is degradable to cysteamine (below the horizontal bold line), only one number is shown, indicating the number of decomposition steps for that thiol. For example, in Disulfide Table 1B, the disulfide represented by "2 + 5" refers to pantetheine (thiol 2) disulfide linked to coenzyme A (thiol 5). The MW of this disulfide is 1,352.36. Upon degradation in the intestine, this disulfide yields two cysteamines. The combined weight of the two cysteamines is 154.3 daltons, which is 11.4% of the mass of the disulfide, as shown in column 4. The degradation pathway from the disulfide to the two cysteamines involves two steps in the case of the pantetheine moiety (step 1: reduction of the disulfide bond, step 2: pantetheinase cleavage) and four or more steps (indicated by 4+) in the case of the coenzyme A moiety (step 1: reduction of the disulfide bond, step 2: ectonucleotide diphosphatase-catalyzed elimination of the nucleotide (other catabolic pathways are possible), step 3: dephosphorylation to pantetheine, step 4: pantetheinase cleavage). Thus, the numbers in column 5: 2 / 4+ indicate the number of degradation steps from the disulfide to the cysteamine for the pantetheine and coenzyme A moieties, respectively. [Figure 19] Two tables are included showing how the thiols and dithiols of Figure 17 can be combined to create two classes of disulfide cysteamine precursors: 4-phosphopantetheine disulfides and dephospho-Coenzyme A disulfides. The five columns in each of the two tables provide the same information as Figure 18. Again, note that for thiols 13 and 14 (L-cysteine ​​ethyl ester HCl and L-cysteine ​​methyl ester HCl), the mass of the salt form is used. The actual mass of the free disulfide is 36.46 daltons less than the mass shown. [Figure 20] Two tables are included showing how the thiols and dithiols in Figure 17 can be combined to create two classes of disulfide cysteamine precursors, coenzyme A disulfide and N-acetylcysteamine disulfide. The five columns in each of the two tables provide the same information as in Figure 18. Again, note that for thiols 13 and 14 (L-cysteine ​​ethyl ester HCl and L-cysteine ​​methyl ester HCl), the mass of the salt form is used. The actual mass of the free disulfide is 36.46 daltons less than the mass shown. [Figure 21] Two tables are included showing how to conjugate a dithiol to two thiols to create a disulfide that can yield two cysteamines (top table) or one cysteamine (bottom table) upon in vivo degradation. Thiol and dithiol numbering is as in Figure 17. Within each table, the various possible dithiol-thiol-thiol combinations are grouped by the dithiol moiety (compounds 30-35) for simplicity, and the molecular weight and cysteamine yield for each group are provided as ranges. Three exemplary dithiol-thiol-thiol combinations are shown at the bottom of each table, including the specific MW, the percent of MW convertible to cysteamine, and the number of degradation steps to cysteamine (see the description of Figure 18 above). Additional details are provided in the explanatory text below the two tables. [Figure 22] 1 shows the initial thiol activation step used in the chemical synthesis of mixed (unsymmetric) disulfides. [Figure 23]Figure 17 shows one synthetic scheme used to make cysteamine-pantetheine disulfide (referred to as TTI-0102, where 01 refers to cysteamine, which is thiol 1 in Figure 17, and 02 refers to pantetheine, which is thiol 2 in Figure 17). The primary amine of cysteamine is first protected with tert-butyloxycarbonyl (Boc), and then the -SH of cysteamine-Boc is activated with bis(5,5-dimethyl-2-thioxo-1,3,2-dioxaphosphorinan-2-yl)disulfane (abbreviated as PDTA) in the presence of 2,3-dichloro-5,6-dicyanobenzoquinone (DDQ) in dichloromethane (DCM). The Boc group is then removed with acid, and the activated cysteamine is reacted with (R)-pantetheine. [Figure 24] The second synthetic scheme used to prepare cysteamine-pantetheine disulfide (TTI-0102) is shown. (R)-pantetheine is activated with bis(5,5-dimethyl-2-thioxo-1,3,2-dioxaphosphorinan-2-yl)disulfane (abbreviated as PDTA) in the presence of 2,3-dichloro-5,6-dicyanobenzoquinone (DDQ) in dichloromethane (DCM). The activated (R)-pantetheine is then reacted with cysteamine in sodium hydride (NaH) and tetrahydrofuran (THF). [Figure 25] Figure 17 shows the synthetic scheme used to make N-acetylcysteamine-pantetheine disulfide (referred to as TTI-0602, where the numbers 6 and 2 refer to the two combined thiols, as numbered in Figure 17). N-acetylcysteamine is activated with bis(5,5-dimethyl-2-thioxo-1,3,2-dioxaphosphorinan-2-yl)disulfane (PDTA) in the presence of 2,3-dichloro-5,6-dicyanobenzoquinone (DDQ) in dichloromethane (DCM). The activated N-acetylcysteamine is then reacted with (R)-pantetheine in triethanolamine (TEA) in DCM. [Figure 26]Figure 17 shows the synthetic scheme used to make N-acetylcysteine-pantetheine disulfide (referred to as TTI-1502, where the numbers 15 and 2 refer to the two combined thiols, as numbered in Figure 17). N-acetylcysteine ​​is activated with bis(5,5-dimethyl-2-thioxo-1,3,2-dioxaphosphorinan-2-yl)disulfane (PDTA) in the presence of 2,3-dichloro-5,6-dicyanobenzoquinone (DDQ) in dichloromethane (DCM). The activated N-acetylcysteine ​​is then reacted with (R)-pantetheine in sodium hydride (NaH) and tetrahydrofuran (THF). [Figure 27] Contains the nuclear magnetic resonance (NMR) spectrum of TTI-0102 obtained on a Varian INOVA 500. The inset structure of TTI-0102 is labeled with letters a–i to indicate specific bonds, which are also highlighted on the NMR spectrum. [Figure 28] Contains a nuclear magnetic resonance (NMR) spectrum of TTI-0602 obtained on a Varian INOVA 500. The inset structure of TTI-0602 is labeled with letters a–g to indicate specific bonds, which are also highlighted on the NMR spectrum. [Figure 29] Contains a nuclear magnetic resonance (NMR) spectrum of TTI-1502 obtained on a Varian INOVA 500. The inset structure of TTI-1502 is labeled with letters a through i to indicate specific bonds, which are also highlighted on the NMR spectrum. [Figure 30A] Figure 30 includes concentration-time curves of cysteamine in plasma after oral gavage administration of cysteamine hydrochloride (30 mg / kg, Figure 30A) or Compound 2 (also known as TTI-0602) (120 mg / kg, Figure 30B) to Sprague-Dawley rats, as described in Example 10. Values ​​for both curves are the average of three rats. Standard deviations are indicated by error bars. [Figure 30B]Figure 30 includes concentration-time curves of cysteamine in plasma after oral gavage administration of cysteamine hydrochloride (30 mg / kg, Figure 30A) or Compound 2 (also known as TTI-0602) (120 mg / kg, Figure 30B) to Sprague-Dawley rats, as described in Example 10. Values ​​for both curves are the average of three rats. Standard deviations are indicated by error bars. [Figure 31A] 31A includes plasma cysteamine concentration-time curves after administration of Compound 2 (also known as TTI-0602) at doses of 30 mg / kg, 60 mg / kg, or 120 mg / kg by oral gavage to Sprague-Dawley rats (3 rats per dose) as described in Example 10 (FIG. 31A), and plasma cysteamine, N-acetylcysteamine, and pantothenic acid concentration-time curves after administration of 120 mg / kg of Compound 2 (also known as TTI-0602) by oral gavage to Sprague-Dawley rats as described in Example 10 (FIG. 31B). [Figure 31B] 31A includes plasma cysteamine concentration-time curves after administration of Compound 2 (also known as TTI-0602) at doses of 30 mg / kg, 60 mg / kg, or 120 mg / kg by oral gavage to Sprague-Dawley rats (3 rats per dose) as described in Example 10 (FIG. 31A), and plasma cysteamine, N-acetylcysteamine, and pantothenic acid concentration-time curves after administration of 120 mg / kg of Compound 2 (also known as TTI-0602) by oral gavage to Sprague-Dawley rats as described in Example 10 (FIG. 31B). [Figure 32] 1 includes a chart showing the concentrations of cysteamine (micromolar) in the liver and kidney 10.5 hours after administration of 120 mg / kg of Compound 2 (also known as TTI-0602) by oral gavage to Sprague-Dawley rats, as described in Example 10. [Figure 33]Figure 1 shows the concentration-time curves of cysteamine in plasma after administration of either (i) cysteamine hydrochloride (30 mg / kg), or cystamine dihydrochloride (30 mg / kg), or pantethine (30 mg / kg), or Compound 1 (also known as TTI-0102) (100 mg / kg) by oral gavage to male Sprague-Dawley rats, as described in Example 1. Concentrations are the average of three rats, and error bars indicate the standard deviation. [Figure 34A] Figure 34A shows the plasma cysteamine concentration-time curve (Figure 34A) and the pharmacokinetic parameters (Figure 34B) derived from each dose of Compound 1 (also known as TTI-0102) administered by oral gavage to male Sprague-Dawley rats at doses of 30 mg / kg, 60 mg / kg, or 100 mg / kg, as described in Examples 11-13. Abbreviations: Cmax = maximum cysteamine concentration; Tmax = time at which maximum cysteamine concentration occurs; AUClast = area under the drug concentration-time curve from 0 to 6 hours; T1 / 2 = half-life of cysteamine in plasma; MRTlast = mean residence time based on 6 hours of data. [Figure 34B] Figure 34A shows the plasma cysteamine concentration-time curve (Figure 34A) and the pharmacokinetic parameters (Figure 34B) derived from each dose of Compound 1 (also known as TTI-0102) administered by oral gavage to male Sprague-Dawley rats at doses of 30 mg / kg, 60 mg / kg, or 100 mg / kg, as described in Examples 11-13. Abbreviations: Cmax = maximum cysteamine concentration; Tmax = time at which maximum cysteamine concentration occurs; AUClast = area under the drug concentration-time curve from 0 to 6 hours; T1 / 2 = half-life of cysteamine in plasma; MRTlast = mean residence time based on 6 hours of data. [Figure 35A]Figure 35A shows bar graphs depicting the concentrations of cysteamine (Figure 35A) and pantetheine (Figure 35B) in the gastrointestinal tract of male Sprague-Dawley rats 6 hours after oral gavage administration of cysteamine hydrochloride (30 mg / kg) or Compound 1 (also known as TTI-0102) at doses of 30 mg / kg, 60 mg / kg, or 100 mg / kg. Gastrointestinal contents obtained from the stomach, proximal small intestine, distal small intestine, and cecum / colon were analyzed as described in Examples 11-13. All samples from four intestinal segments were reduced with TCEP. Proximal and distal small intestinal contents were also analyzed without TCEP. The numbers above each bar indicate the value for that sample. To facilitate comparison of cysteamine and pantetheine levels in the same sample, samples are arranged in upper and lower panels. [Figure 35B] Figure 35A shows bar graphs depicting the concentrations of cysteamine (Figure 35A) and pantetheine (Figure 35B) in the gastrointestinal tract of male Sprague-Dawley rats 6 hours after oral gavage administration of cysteamine hydrochloride (30 mg / kg) or Compound 1 (also known as TTI-0102) at doses of 30 mg / kg, 60 mg / kg, or 100 mg / kg. Gastrointestinal contents obtained from the stomach, proximal small intestine, distal small intestine, and cecum / colon were analyzed as described in Examples 11-13. All samples from four intestinal segments were reduced with TCEP. Proximal and distal small intestinal contents were also analyzed without TCEP. The numbers above each bar indicate the value for that sample. To facilitate comparison of cysteamine and pantetheine levels in the same sample, samples are arranged in upper and lower panels. [Figure 36A] Figure 1 shows a synthetic scheme for the hemiacylation of cystamine to produce compound 1 (cysteamine-pantetheine asymmetric disulfide). DCC is the abbreviation for the coupling reagent N,N'-dicyclohexylcarbodiimide. HOBt is the abbreviation for 1-hydroxybenzotriazole hydrate. DMF is the abbreviation for the solvent dimethylformamide. [Figure 36B]

[0033] Figure 1 shows a scheme for the synthesis of Compound 1 via thiol-disulfide exchange. The illustrated reaction, involving cysteamine (a thiol) and pantetheine (a disulfide), is one of two thiol-disulfide exchange reactions discussed in Example 14. [Figure 37] Figure 36A shows the proton nuclear magnetic resonance (H NMR) spectrum of TTI-0102 produced by hemiacylation of cystamine (shown in Figure 36A). The structure of TTI-0102 is shown, and the peaks in the H NMR spectrum are annotated with lowercase letters to match. The table shows the numerical values ​​of all peaks, their delta values ​​(expressed in parts per million, or ppm), and peak heights. [Figure 38] The predicted and observed proton peak heights for the H NMR spectrum shown in Figure 37 are tabulated, with the protons labeled (a, b, c, d, e, f, g, h, i) as in Figure 37. DETAILED DESCRIPTION OF THE INVENTION

[0081] The present invention features compositions and methods that enable the in vivo production of cysteamine from precursor compounds (cysteamine precursors) in controlled amounts and at controlled locations in the gastrointestinal tract, as well as methods for treating cysteamine-sensitive conditions, syndromes, and diseases. The methods and compositions of the present invention can include any one of Compounds 1-3 shown below, or a pharmaceutically acceptable salt thereof. TIFF0007794914000014.tif73170 Compounds 1-3 can be administered to a subject following administration of a compound such as a reducing agent or pantetheinase inducer, alone or in combination with a second active agent that is a cysteamine precursor, or in combination with an agent that modifies cysteamine release or uptake.

[0082] Cysteamine is a small, highly reactive thiol molecule (NH2-CH2-CH2-SH) present in all living organisms, from bacteria to humans. Its IUPAC name is 2-aminoethanethiol. Other common names include mercaptamine, beta-mercaptoethylamine, 2-mercaptoethylamine, decarboxycysteine, and thioethanolamine. In humans, cysteamine is produced by the enzyme pantetheinase (also known as pantothenate or vitamin B5), which cleaves pantetheine into cysteamine and pantothenic acid. Human pantetheninase is encoded by the Vanin 1 and Vanin 2 genes (abbreviated VNN1 and VNN2) and is widely expressed, including in the gastrointestinal tract. Thus, dietary pantetheine, present in many foods (e.g., nuts and dairy products), is cleaved in the gastrointestinal lumen to produce cysteamine and pantothenic acid, which are then absorbed. In particular, cysteamine can be transported across the gastrointestinal epithelium by organic cation transporters (OCTs), a family of transporters including organic cation transporter 1 (OCT1), OCT2, and OCT3, which have been shown to transport cysteamine within enterocytes. Based on its ability to be converted to cysteamine in the gastrointestinal tract, pantetheine is a cysteamine precursor. Cysteamine precursors represent a class of compounds that may have advantages over cysteamine salts in terms of (i) tolerability and side effects, (ii) pharmacokinetics and administration interval, (iii) manufacturing, and (iv) product stability. More generally, administering cysteamine precursors capable of generating cysteamine in vivo at various rates and using formulation methods to deliver these precursors to selected sites in the gastrointestinal tract at selected times could be useful in therapeutic regimens by providing much better control of cysteamine pharmacokinetics, which, until now, has been a major obstacle to the widespread use of cysteamine and other thiols.

[0083] Cysteamine precursor Pantetheine and its catabolic products, cysteamine and pantothenate, are intermediate compounds in the biosynthesis of coenzyme A in plants and animals (see Figure 11 for a diagram of the relevant metabolic and catabolic pathways). Some compounds in the coenzyme A biosynthetic pathway, such as 4-phosphopantetheine, dephosphocoenzyme A, and coenzyme A, can be catabolized to pantetheine and then cysteamine and pantothenate in the human gastrointestinal tract. Thus, 4-phosphopantetheine, dephosphocoenzyme A, and coenzyme A are cysteamine precursors because they can be converted to cysteamine in the intestine. N-acetylcysteamine is also a cysteamine precursor through deacetylation by intestinal or cellular deacetylases (e.g., deacetylases that convert N-acetylcysteine ​​to cysteine ​​in vivo).

[0084] Pantethine is a dimer of two pantetheine molecules joined by a disulfide bond. In other words, pantethine is the oxidized form of pantetheine. The interconversion of pantethine into two pantetheines is not enzymatically mediated and does not require ATP. Instead, this reaction is primarily controlled by the redox environment in the intestine. In vivo, pantetheine predominates in reducing environments, which tend to prevail, especially within cells, whereas in more oxidative environments, such as the stomach, the equilibrium shifts toward pantethine. A small clinical study by Wittwer (Wittwer et al., J. Exp. Med. 76:4 (1985)) showed that upon oral administration, a significant fraction of pantethine is chemically reduced to pantetheine in the human gastrointestinal tract and subsequently cleaved to cysteamine and pantothenate. Therefore, pantethine is a cysteamine precursor. Pantetheine herein refers to the D-enantiomer.

[0085] The pantothenoyl moiety of pantetheine contains a chiral carbon. Therefore, two enantiomeric forms of pantetheine exist, traditionally referred to as D-pantetheine and L-pantetheine (also referred to as R-pantetheine and S-pantetheine). Only the D-enantiomer of pantetheine can be cleaved by pantetheinase, and therefore only the D-enantiomer is eligible as a cysteamine precursor. The two enantiomers of pantetheine can combine in four ways (D-,D-; D-,L-; L-,D-; and L-,L-pantetheine) to form the disulfide pantethine. Only D-,D-pantethine can be chemically reduced to two D-pantetheines and then cleaved to produce two cysteamines. Therefore, the D-,D-form of pantethine is highly preferred, and the term pantethine, as used herein, refers to the D-,D-enantiomer. The pantetheine-related compounds 4-phosphopantetheine, dephospho-coenzyme A, and coenzyme A must also be in the D-stereoisomeric configuration to yield D-pantetheine (and thus cysteamine) upon intestinal degradation. Thus, "4-phosphopantetheine," "dephospho-coenzyme A," and "coenzyme A," as well as any analogs or derivatives thereof, refer to the D-enantiomers herein. Neither pantetheine, 4-phosphopantetheine, dephospho-coenzyme A, nor coenzyme A are absorbed by enterocytes; rather, each compound must be catabolized to pantothenate and cysteamine, which are absorbed (see Shibata et al., J. Nutr. 113:2107 (1983)).

[0086] Analogs or derivatives of pantetheine, 4-phosphopantetheine, dephosphocoenzyme A, or the D-stereoisomer of coenzyme A that can be converted to the parent compound in the gastrointestinal tract (e.g., by natural enzymatic or chemical processes) can also be used to form thiol or disulfide-type cysteamine precursors, and are referred to herein as "suitable analogs or derivatives." For example, there are many physiological forms of coenzymes (e.g., acetyl-CoA, succinyl-CoA, malonyl-CoA, etc.) that are readily degraded to coenzyme A in the intestine. Any acetylated, alkylated, phosphorylated, lipidated, or other analogs can be used as cysteamine precursors. Analogs of pantetheine, 4-phosphopantetheine, dephosphocoenzyme A, or coenzyme A, as well as methods for producing them, have been described in the literature (van Wyk et al., Chem Commun 4:398 (2007)).

[0087] Pantetheine can form disulfides with thiols other than itself, called pantetheine mixed disulfides, which constitute another class of cysteamine precursors. The thiols reacted with pantetheine are preferably naturally occurring thiols or non-naturally occurring thiols known to be safe in humans based on a history of human or animal use. For example, mixed disulfides can be formed by reacting pantetheine with 4-phosphopantetheine, dephosphocoenzyme A, or coenzyme A, compounds present in the human body and many foods. Such mixed disulfides yield two cysteamines upon reduction and degradation in the intestine. Pantetheine linked to N-acetylcysteamine also yields two cysteamines upon reduction and degradation in the intestine. In certain embodiments, disulfide cysteamine precursors capable of yielding two cysteamines are preferred. Figures 18-21 show the cysteamine yields of different classes of disulfide cysteamine precursors. 4-phosphopantetheine, dephospho-coenzyme A, or analogs or derivatives of coenzyme A that can be converted to the parent compound in the gastrointestinal tract via chemical or enzymatic processes (i.e., suitable analogs or derivatives) can also be linked to pantetheine to form the pantetheine mixed disulfide cysteamine precursor or can be linked to other thiols.

[0088] Pantetheine mixed disulfides can also be formed by reacting pantetheine with a thiol that is not itself decomposable to cysteamine, such as L-cysteine, homocysteine, N-acetylcysteine, N-acetylcysteineamide, N-acetylcysteine ​​ethyl ester, N-acetylcysteamine, L-cysteine ​​ethyl ester hydrochloride, L-cysteine ​​methyl ester hydrochloride, thiocysteine, allyl mercaptan, furfuryl mercaptan, benzyl mercaptan, thioterpineol, 3-mercaptopyruvate, cysteinylglycine, gamma glutamylcysteine, gamma-glutamylcysteine ​​ethyl ester, glutathione, glutathione monoethyl ester, glutathione diethyl ester, mercaptoethylgluconamide, thiosalicylic acid, thiocysteine, tiopronin, or diethyldithiocarbamic acid. See Figure 17 for Chemical Abstracts Service (CAS) registry numbers, molecular formulas, and molecular weights of exemplary thiol compounds that can react with pantetheine to form pantetheine mixed disulfides. Disulfides formed by pantetheine and any of thiols 6-35 (see Figure 17 for thiol numbering) yield one cysteamine upon reduction of the disulfide bond and pantetheinase cleavage. These second thiols are not convertible to cysteamine in the intestine but can nevertheless promote cysteamine production, e.g., by stimulating pantetheinase activity or participating in disulfide exchange with cysteamine-containing disulfides, or can provide therapeutic benefits complementary to those provided by cysteamine, e.g., by acting as reducing agents or by other mechanisms.

[0089] Dithiol compounds such as dihydrolipoic acid (DHLA), meso-2,3-dimercaptosuccinic acid (DMSA), 2,3-dimercaptopropanesulfonic acid (DMPS), 2,3-dimercapto-1-propanol, bucillamine, or N,N'-bis(2-mercaptoethyl)isophthalamide can also react with pantetheine to form either pantetheine mixed disulfides with one free thiol group or tripartite compounds with two disulfide bonds connecting two pantetheine molecules to the dithiol. The former category of mixed pantetheine disulfides yields one cysteamine upon reduction and pantetheinase cleavage of the disulfide bond, while the latter category yields two cysteamines. See Figure 21 for a table showing how cysteamine, pantetheine, 4-phosphopantetheine, dephosphocoenzyme A, coenzyme A, or N-acetylcysteamine can be combined with various dithiols to produce useful cysteamine precursors. Alternatively, two different thiols can be attached to a dithiol to yield a cysteamine precursor, i.e., a compound that can ultimately be degraded to cysteamine in the gastrointestinal tract, so long as one of the thiols is cysteamine, pantetheine, 4-phosphopantetheine, dephosphocoenzyme A, coenzyme A, or N-acetylcysteamine, or a suitable analog or derivative thereof. Tables 2A and 2B in Figure 21 show some of the salient properties of such cysteamine precursors, including the molecular weight and cysteamine yield range (i.e., the percent of cysteamine precursor that can be converted to cysteamine in vivo), and for selected examples, the number of in vivo degradation steps from cysteamine precursor to cysteamine.

[0090] Like pantetheine, 4-phosphopantetheine, dephosphocoenzyme A, coenzyme A, or N-acetylcysteamine, or suitable analogs or derivatives, can (i) react with themselves to form homodimeric disulfides, or (ii) react with each other in various pairs to form mixed disulfides, or (iii) react with other thiols (which cannot be converted to cysteamine in vivo) to form mixed disulfides.All such disulfides are cysteamine precursors.The first two categories can produce two cysteamines upon reduction and decomposition in the intestine, while the third category can produce only one cysteamine.

[0091] For example, any of the thiols listed in Figure 17 can be reacted with 4-phosphopantetheine (as shown in Figure 19), dephosphocoenzyme A (Figure 19), coenzyme A (Figure 20), or N-acetylcysteamine (Figure 20) to form a mixed disulfide cysteamine precursor. Other naturally occurring thiols can also be used, as can unnatural thiols known to be safe in humans. Figures 18-21 schematically illustrate some of the combinations of thiols and dithiols that can react to form disulfide cysteamine precursors. Conversion of such compounds to cysteamine in the human gastrointestinal tract requires (i) reduction of the disulfide bond to generate the free thiol; (ii) degradation by intestinal enzymes (e.g., phosphatases, diphosphatases, phosphodiesterases) to generate pantetheine in the case of 4-phosphopantetheine, dephosphocoenzyme A, coenzyme A-containing disulfides, or suitable analogs or derivatives thereof; and (iii) cleavage of the pantetheine by pantetheinase. The disulfide-containing N-acetylcysteamine must be reduced and deacetylated in the intestine, blood, or tissues.

[0092] Cysteamine itself can also be reacted with other thiols to form mixed disulfide cysteamine precursors. For example, cysteamine can be reacted with pantetheine, 4-phosphopantetheine, dephosphocoenzyme A, coenzyme A, or N-acetylcysteamine, analogs or derivatives of the five thiols that can be degraded to the parent compounds in the gastrointestinal tract, or with any of the other thiols listed in Figure 17 to form any of the disulfides shown in Figures 18-20. Two cysteamines can be conjugated to a dithiol via two disulfide bonds to produce another type of disulfide cysteamine precursor (Figure 21). Figure 8 shows the chemical structure of such a cysteamine precursor, dihydrolipoate disulfide linked to two cysteamines. Upon reduction of the disulfide bond, two cysteamines are released, along with dihydrolipoic acid, a potent reducing agent, which may complement the therapeutic properties of cysteamine in certain disease settings.

[0093] In summary, cysteamine precursors can be divided into three main categories: (i) thiols that can be degraded to cysteamine, (ii) mixed disulfides containing cysteamine, including disulfides formed with dithiols, (ii) disulfides containing pantetheine, and (iii) disulfides containing 4-phosphopantetheine, dephosphocoenzyme A, or coenzyme A, or a suitable analog or derivative. Each of the latter three categories can be further degraded depending on the second thiol: (a) pantetheine, or a suitable analog or derivative, (b) 4-phosphopantetheine, dephosphocoenzyme A, or coenzyme A, or a suitable analog or derivative, or (c) a thiol that is not itself a cysteamine precursor (e.g., L-cysteine, homocysteine, N-acetyl-cysteine, N-acetylcysteine ​​amide, N-acetylcysteine ​​ethyl ester, N-acetylcysteamine, L-cysteine ​​ethyl ester). ester hydrochloride, L-cysteine ​​methyl ester hydrochloride, thiocysteine, allyl mercaptan, furfuryl mercaptan, benzyl mercaptan, 3-mercaptopyruvate, thioterpineol, glutathione, cysteinylglycine, gamma glutamylcysteine, gamma-glutamylcysteine ​​ethyl ester, glutathione monoethyl ester, glutathione diethyl ester, mercaptoethylgluconamide, thiosalicylic acid, thiocysteine, tiopronin, or diethyldithiocarbamate). Dithiol compounds such as dihydrolipoic acid, meso-2,3-dimercaptosuccinic acid (DMSA), 2,3-dimercaptopropanesulfonic acid (DMPS), 2,3-dimercapto-1-propanol, bucillamine, or N,N′-bis(2-mercaptoethyl)isophthalamide can also be combined with cysteamine, pantetheine, 4-phosphopantetheine, dephospho-coenzyme A, or coenzyme A, or suitable analogs or derivatives, to form disulfides.

[0094] Pharmacological properties of cysteamine precursors The temporal and spatial patterns of in vivo cysteamine production from cysteamine precursors can vary widely depending on the type of cysteamine precursor. Cysteamine precursors that require multiple chemical and enzymatic reactions to produce cysteamine will, on average, produce cysteamine later than those that require only one step. This property of cysteamine precursors can be used to design multiple pharmaceutical compositions that vary in the rate and duration of in vivo cysteamine production. Furthermore, pharmaceutical compositions can be administered in combinations and ratios that result in the desired pharmacological outcome. For example, cysteamine mixed disulfides can be administered to provide elevated plasma cysteamine levels immediately after drug administration. The only step required to produce cysteamine from cysteamine mixed disulfides is the reduction of the disulfide bond. Depending on the identity of the second thiol, a second cysteamine can be produced following one or more decomposition steps. Because the second cysteamine can only be produced after disulfide bond reduction and another step, it is necessarily produced slower than the first cysteamine, thereby extending the period during which cysteamine is produced in the intestine and absorbed into the blood. Because cysteamine free base and cysteamine salts (e.g., Cystagon® and Procysbi®) have very short half-lives, this extension of in vivo cysteamine production from cysteamine precursors represents a significant advance over current therapies.

[0095] In one approach, when the second thiol is pantetheine (i.e., cysteamine-pantetheine disulfide), a pantetheinase cleavage step is required to generate a second cysteamine. Because pantetheinase is generally located on the surface of enterocytes, it is only in contact with a portion of the intestinal contents at any given time, thereby extending the period during which cysteamine is generated. This combination of early and late cysteamine generation from a single disulfide molecule has several advantages: (i) cysteamine becomes available upon disulfide bond reduction, providing an early therapeutic benefit; (ii) pantetheine cleavage occurs over time (pantetheinase is expressed at various levels throughout the gastrointestinal tract), extending the duration of the therapeutic effect; (iii) the temporally and spatially extended cysteamine production via both bond reduction and pantetheine cleavage reduces the high peak cysteamine concentrations strongly associated with side effects, while (iv) avoids saturation of pantetheinase or cysteamine uptake mechanisms, such as transport by OCT. Thus, long-term elevation of blood cysteamine levels provides patients with both more effective drug therapy and a less toxic and more convenient dosage form.

[0096] Alternatively, if the second thiol is L-cysteine ​​(i.e., cysteamine-L-cysteine ​​disulfide), only one cysteamine is produced upon reduction of the disulfide, and there is no long-term cysteamine production. However, as described below, cysteamine-L-cysteine ​​disulfide can be formulated for release in virtually any part of the gastrointestinal tract, including the ileum or colon, where a cysteamine precursor capable of rapid cysteamine release may be useful. Furthermore, cysteine ​​has also been shown to promote pantetheinase activity and have beneficial effects in several disease models. Therefore, cysteamine-L-cysteine ​​disulfide may be a useful complement to another cysteamine precursor or may be useful in treating diseases that respond to both cysteamine and cysteine.

[0097] Disulfides containing thiols, such as 4-phosphopantetheine, dephosphocoenzyme A, or coenzyme A, which require two or more catabolic reactions to produce cysteamine, or suitable analogs or derivatives thereof, are degraded more efficiently in the small intestine, where they are more exposed to the digestive enzymes present in pancreatic juice than in the stomach or large intestine. Disulfides created by reacting two such thiols with each other or with thiols other than cysteamine start to produce cysteamine at a later time point and over a longer period than, for example, cysteamine-L-cysteine ​​disulfide. On average, 4-phosphopantetheine, dephosphocoenzyme A, or coenzyme A, or suitable analogs, produce cysteamine more slowly than pantetheine, and the same is true for disulfides containing these compounds.

[0098] Cysteamine precursors such as pantetheine and compounds degradable to pantetheine in the intestine, as well as disulfides containing either of these compounds, all yield pantothenate along with cysteamine upon cleavage by pantetheinase. Pantothenate, or vitamin B5, is a water-soluble compound present in food and synthesized by intestinal bacteria. When pantothenate is administered in large doses, excess amounts are excreted in the urine. A review of pantothenate by the Panel on Folate, Other B Vitamins, and Choline of the US Institute of Medicine Standing Committee on the Scientific Evaluation of Dietary Reference Intakes (National Academies Press (US), 1998) stated, "There are no reported adverse effects of oral pantothenic acid in humans or animals."

[0099] Cysteamine Precursor Mixture The methods and compositions of the present invention can include a mixture of cysteamine precursors to take advantage of their different pharmacological properties. In particular, personalized improvement of cysteamine plasma levels (or personalization for a given patient's needs) can be achieved by using a mixture of cysteamine precursors. For example, the cysteamine-pantetheine mixed disulfide described above has a fixed 1:1 ratio of cysteamine to pantetheine. However, cysteamine is rapidly absorbed and eliminated from the body (elimination half-life: approximately 25 minutes), resulting in a sudden peak in blood levels, while pantetheine provides cysteamine over several hours (via pantetheinase cleavage). Therefore, because cysteamine production from pantetheine is spread over a long period of time, a dose of cysteamine-pantetheine mixed disulfide that results in therapeutic cysteamine levels early on (from cysteamine released upon reduction of the disulfide bond) may later result in subtherapeutic cysteamine levels. Therefore, a 1:1 ratio of cysteamine to pantetheine may not be ideal for certain patients or purposes. Adding more pantetheine to the dosage form maintains blood cysteamine within the therapeutic concentration range for a longer period of time.In order to increase the ratio of pantetheine to cysteamine, either thiol pantetheine or disulfide pantethine or another pantetheine-containing disulfide can be co-formulated or co-administered with, for example, cysteamine-pantetheine mixed disulfide, to achieve blood cysteamine levels within the therapeutic range for a longer period of time.The ratio of the two cysteamine precursors can be adjusted to achieve desired pharmacokinetic parameters, such as maximizing the area under the cysteamine concentration-time curve (AUC), or minimizing the peak concentration (Cmax) of cysteamine, or maximizing the trough concentration (Cmin), or maintaining cysteamine blood levels above the threshold, or any combination of such parameters.

[0100] Cysteamine precursors such as 4-phosphopantetheine, dephosphocoenzyme A, or coenzyme A, and the disulfides formed from these three compounds, require more catabolic steps to produce cysteamine than pantetheine. Therefore, the rate of cysteamine production from these cysteamine precursors is, on average, slower and more prolonged than that of pantetheine or certain pantetheine disulfides. Therefore, the coadministration or co-formulation of 4-phosphopantetheine, dephosphocoenzyme A, or coenzyme A, or their disulfides with the cysteamine-pantetheine combination, and optionally pantetheine or pantethine, provides another method for controlling cysteamine pharmacokinetics by selecting an appropriate cysteamine precursor. In particular, the use of such cysteamine precursors can be used to further extend the time during which cysteamine is produced in the gastrointestinal tract.

[0101] 4-Phosphopantethine, dephosphocoenzyme A, and coenzyme A-containing disulfides The formal biosynthetic pathway for coenzyme A, shown schematically in Figure 11, requires five steps catalyzed by four enzymes (CoA synthase catalyzes the final two steps). The initial step of phosphorylation of pantothenate by pantothenate kinase controls flux through the pathway. Until recently, it was thought that none of the intermediate compounds in the coenzyme A synthesis (or catabolic) pathway were efficiently absorbed in the gastrointestinal tract. Rather, only the catabolic product of pantetheine (pantothenate and cysteamine) is absorbed in the intestine. Two important consequences of our understanding of the coenzyme A pathway for cysteamine precursor therapy are that (i) cysteamine precursors must be broken down to cysteamine in the intestine, then absorbed and transported to the site of therapeutic effect (e.g., liver, central nervous system), and (ii) cellular coenzyme A synthesis necessarily begins with pantothenate (because other metabolic intermediates do not cross the cell membrane).

[0102] However, 4-phosphopantetheine efficiently crosses cell membranes (Srinivasan et al., Nature Chemical Biology 11:784 (2015)). This observation has implications for the design and use of the cysteamine precursors described herein to treat various diseases and disorders. First, it enables a therapeutic approach involving in situ cysteamine production in multiple tissues and organs (as opposed to only the intestine), including diseased tissue. Second, it enables cellular delivery of the coenzyme A precursor (4-phosphopantetheine) downstream of the initial synthesis step catalyzed by pantothenate kinase, which can be used to treat pantothenate kinase-deficient subjects. Methods for using cysteamine precursors to treat these two categories of diseases are described below and illustrated with several examples.

[0103] In one approach, because these organs (and others) all contain pantetheinases expressed from either the VNN1 or VNN2 gene, diseases of the kidney, liver, lungs, and connective tissue, as well as infectious diseases, can be effectively treated. This method involves (i) administering to the patient a cysteamine precursor that can be degraded in the intestine to yield one or two molecules of 4-phosphopantetheine, some of which (ii) are absorbed by enterocytes and enter the blood (where 4-phosphopantetheine is relatively stable), then (iii) pass through the diseased organ via the circulation, where (iv) they can be degraded by phosphatases and pantetheinases to yield cysteamine at the site of disease.

[0104] Advantages of this treatment method include: (i) higher cysteamine concentrations at the site of disease that can be achieved with cysteamine absorbed from the gut per equivalent dose; (ii) lower plasma cysteamine concentrations, resulting in lower toxicity (4-phosphopantetheine is a circulatory delivery vehicle); (iii) a longer blood half-life than cysteamine (approximately 25 minutes for cysteamine versus more than 3 hours for 4-phosphopantetheine), which allows for longer dosing intervals and thereby increased patient convenience; and (iv) the ability to selectively target cysteamine to diseased tissues where pantetheinase overexpression is pathogenic, including, for example, metabolic diseases such as NASH (Sato W. et al., Hepatol Res. 34:256 (2006)) and certain inflammatory diseases (Naquet P. et al., Biochem Soc Trans. 42:1094 (2014)). Because inflammation is often present at sites of infection, selective cysteamine production at sites of infection is also possible, where cysteamine has antibacterial, antiviral, or antiparasitic properties. Thus, 4'-phosphopantetheine can be absorbed in the intestine, circulate in the blood, and then degraded to cysteamine in organs or diseased tissues that express pantetheinase, either constitutively, as in the kidney, or as a manifestation of active disease, as in inflammation.

[0105] 4-Phosphopantethine - disulfide formation in renal disease As mentioned above, pantetheinase (encoded by both the VNN1 and VNN2 genes) is expressed at high levels in the kidney. Therefore, some circulating 4-phosphopantetheine is degraded in the kidney to produce cysteamine. The advantages of kidney-specific cysteamine production include higher tissue levels than achieved through cysteamine absorption by the gastrointestinal tract and fewer side effects associated with elevated blood levels of cysteamine (e.g., foul-smelling breath and sweating, nausea, vomiting, loss of appetite, and stomach pain). Renal diseases responsive to cysteamine therapy include fibrotic diseases (e.g., glomerulonephritis) and metabolic diseases, including nephropathic cystinosis (renal failure is a major complication that can be delayed by up to 10 years with cysteamine therapy).

[0106] Cystinuria is another inherited kidney disease associated with recurrent kidney stones (nephrolithiasis). On average, adult patients require surgery every three years for pain, infection, or other complications associated with kidney stones, and the average patient has undergone seven surgical procedures for nephrolithiasis by middle age. Patients with cystinuria are at high risk for kidney loss, requiring nephrectomy. A small but significant proportion of cases (1–3%) develop end-stage renal disease and must be treated with dialysis or kidney transplantation.

[0107] Cystinuria is caused by mutations in one of two genes (SLC3A1 and SLC7A9) that encode the low-affinity cystine transporter rBAT (heterodimer). Disease transmission is autosomal recessive; individuals who inherit two defective copies of either gene develop cystinuria.

[0108] In healthy human subjects, only 0.4% of cystine filtered through the glomerulus ultimately is excreted in the urine. The other 99.6% is reabsorbed in the proximal tubule by rBAT (and to a lesser extent by other transporters). When rBAT is defective, cystine collects in the renal pelvis, leaving high concentrations of cystine in the urine. Cystine can precipitate as stones, causing ureteral obstruction and severe pain. Kidney stones also increase the risk of infection. (Not all patients with cystinuria develop stones; the spectrum of the disease is quite broad.)

[0109] Initial treatment for patients with cystinuria who develop stones involves dietary modification, drinking up to 5 liters of fluid per day, and alkalinizing the urine to approximately pH 7.5, which increases cystine solubility. Second-line therapy involves the administration of thiol compounds capable of forming mixed disulfides with cysteine. Mixed disulfides are more soluble than cystine and therefore remain dissolved in the urine. Thiols such as penicillamine and tiopronin have been used in this way, but are poorly tolerated by most patients. Alpha-mercaptopropionylglycine is also approved by the US FDA for cystinuria, but is poorly tolerated by approximately one-third of patients.

[0110] Orally administered cysteamine precursors are a useful class of therapeutic compounds for cystinuria. They can be degraded to 4-phosphopantetheine in the intestine, then absorbed and enter the circulation, and finally degraded to pantetheine, which is then degraded to cysteamine by pantetheinase in the kidney.Cysteamine easily forms a mixed disulfide with cysteine ​​through disulfide exchange with cystine, and cysteamine-cysteine ​​disulfide is more soluble in aqueous solutions (e.g., urine) than cystine.This therapeutic approach involves the formation of cysteamine in the kidney, so it requires a lower dose of cysteamine precursor than that required for cysteamine formed in the intestine and absorbed therefrom (only a small portion of it reaches the kidney).

[0111] Other renal diseases amenable to cysteamine therapy, including fibrotic diseases associated with oxidative damage and genetic disorders, including diseases caused by mutations that convert arginine codons to cysteine ​​codons, can be treated using a similar approach. The renal blood supply, which is a major component of cardiac output, ensures delivery of a significant portion of absorbed 4-phosphopantetheine to the kidney.

[0112] More generally, cysteamine precursors that can be degraded to 4-phosphopantetheine (including 4-phosphopantetheine disulfide) are useful for providing therapeutic doses of cysteamine to all organs that express significant levels of phosphatases and pantetheinases, for example, to treat lung diseases associated with oxidative damage.

[0113] Useful cysteamine precursors for these therapeutic methods include coenzyme A, dephosphocoenzyme A, and disulfides containing 4'-phosphopantetheine, each of which can be degraded to 4'-phosphopantetheine in the gastrointestinal tract by disulfide bond reduction (in the case of disulfides containing 4'-phosphopantetheine) or by disulfide bond reduction followed by enzymatic degradation (in the case of disulfides containing coenzyme A and dephosphocoenzyme A). In some embodiments, cysteamine precursors that provide two molecules of 4'-phosphopantetheine are preferred over those that provide one. For example, 4'-phosphopantetheine-dephosphocoenzyme A mixed disulfide or homodimeric 4'-phosphopantetheine disulfide can deliver more in situ cysteamine generating capacity than cysteine-4-phosphopantetheine mixed disulfide. Another useful class of cysteamine precursors contains a dithiol linked to one or two thiols that can degrade to 4'-phosphopantetheine. For example, dihydrolipoic acid is linked to one or two molecules of 4'-phosphopantetheine via disulfide bonds.

[0114] More generally, any disulfide consisting of 4′-phosphopantetheine, dephosphocoenzyme A, or coenzyme A and another thiol can serve as a source of 4′-phosphopantetheine after reduction of the disulfide bond and (in the case of dephosphocoenzyme A or coenzyme A) partial degradation in the gastrointestinal tract. After transport across the gastrointestinal epithelium and reaching the circulation, 4′-phosphopantetheine can be degraded to pantetheine by serum phosphatases (however, this is a slow reaction), and then to cysteamine and pantothenate in the blood by pantetheinase (a fast reaction), or 4′-phosphopantetheine can be degraded upon contact with tissues expressing phosphatases and pantetheinases. Phosphatases (collectively) are widely expressed, including acid phosphatases encoded by the ACP1, ACP2, ACP5, and ACPT genes, and alkaline phosphatases encoded by the ALPI, ALPL, ALPP, and ALPPL2 genes. Tissues that express the pantetheinase encoded by VNN1 include the liver, kidney, heart, and gastrointestinal tract, while the pantetheinase encoded by VNN2 is expressed in the kidney, bladder, pancreas, spleen, lung, hematopoietic system (e.g., bone marrow, lymph nodes, tonsils), connective tissue (smooth muscle, adipose tissue), and to a lesser extent in the thyroid gland, adrenal gland, heart, and reproductive organs (testes, ovaries, fallopian tubes, endometrium). The VNN3 gene has been described as a pseudogene, but several reports have described differential VNN3 expression, suggesting a functional role. VNN3 is widely expressed. Data on tissue and cell line expression of vanillin family genes can be found in public databases such as the Protein Atlas (www.proteinatlas.org) and in several publications (e.g., Jansen, P.A.M. et al. Expression of the Vanin Gene Family in Normal and Inflamed Human Skin: Induction by Proinflammatory Cytokines. J. Investigative Dermatology 129:2167-2174, 2009).

[0115] Pantothenate kinase-associated neurodegeneration (PKAN) A second therapeutic approach in which disulfide cysteamine precursors delivering 4-phosphopantetheine can be used therapeutically is exemplified by the disease known as pantothenate kinase-associated neurodegeneration (PKAN). Preclinical and clinical evidence suggests that cysteamine is therapeutically effective in several neurodegenerative disorders, including Parkinson's disease, Huntington's disease, and neurodegeneration due to brain iron accumulation (NBIA). NBIA refers to a rare, clinically heterogeneous group of disorders variably associated with progressive extrapyramidal signs, delayed motor development, and cognitive decline, among other symptoms. Age of onset ranges from infancy to late adulthood. The onset of symptoms varies widely, as does the rate of progression. Consequently, diagnosis is usually suggested by the observation of abnormal iron accumulation in the basal ganglia on brain MRI scans. Cerebellar atrophy may also be present. NBIA is associated with mutations in one of 10 genes: PANK2, PLA2G6, C19orf12, FA2H, ATP13A2, WDR45, COASY, FTL, CP, and DCAF17. With the exception of mutations in the WDR45 gene, which is located on the X chromosome, NBIA is transmitted as an autosomal recessive disorder.

[0116] The most common type of NBIA (30–50% of all cases) is pantothenate kinase-associated neurodegeneration (PKAN), caused by mutations in the gene encoding pantothenate kinase 2 (PANK2). Mitochondrially localized pantothenate kinase 2 phosphorylates pantothenate to generate 4-phosphopantothenate, which is converted to 4-phosphopantothenoyl-cysteine ​​and subsequently decarboxylated to 4-phosphopantetheine (see Figure 11). Providing a source of 4′-phosphopantetheine, a downstream metabolite of the PANK2-catalyzed step, overcomes the requirement for a functional PANK2 enzyme. Both coenzyme A and dephosphocoenzyme A can be degraded to 4′-phosphopantetheine in the gastrointestinal tract. Therefore, any disulfide consisting of 4′-phosphopantetheine, dephosphocoenzyme A, or coenzyme A and another thiol can complement the PANK2 defect.

[0117] In certain embodiments, disulfides containing 4'-phosphopantetheine, dephospho-Coenzyme A, or Coenzyme A can be administered to patients suffering from PANK2 deficiency to ameliorate disease symptoms. Specifically, the disulfides are shown in Figure 19 (Tables 1C and 1D), Figure 20 (Table 1E), and Figure 21 (subsets of compounds containing at least one 4'-phosphopantetheine, one dephospho-Coenzyme A, or one Coenzyme A; thiols 3, 4, and 5, respectively, in the figure nomenclature).

[0118] The disulfide cysteamine precursors of the present application are particularly suitable for carrying out the therapeutic methods outlined above. Disulfides provide an effective method for delivering 4'-phosphopantetheine (and ultimately cysteamine) because (i) disulfides are stable in air (i.e., oxygen-stable), and therefore easier to formulate and store than thiols and are stable for longer periods of time; (ii) the thiol group is protected until the disulfide is reduced in the small intestine near the site of absorption; and (iii) a second thiol with additional or complementary therapeutic properties can be delivered simultaneously. For example, in some embodiments, cysteamine-4-phosphopantetheine mixed disulfide, cysteamine-dephosphocoenzyme A mixed disulfide, and cysteamine-coenzyme A mixed disulfide are useful therapeutic compounds.

[0119] N-acetylcysteamine disulfide (compound 3) In certain embodiments, the cysteamine precursor is Compound 3 or a pharmaceutically acceptable salt thereof. The homodimer of two N-acetylcysteamines is an effective delivery vehicle for cysteamine that can be used in two ways: it can be administered as a single agent or in combination with one or more other cysteamine precursors. In either case, the goal is to provide sustained blood N-acetylcysteamine and cysteamine levels within the therapeutic range (e.g., greater than 5 micromolar and less than 75 micromolar, or greater than 10 micromolar and less than 65 micromolar in plasma) for as long as possible.

[0120] In those embodiments in which Compound 3 is administered as a single agent, it is preferably formulated in a manner that provides at least two release profiles: an early-release profile and a late-release profile. Early-release formulations (also known as immediate-release) begin releasing Compound 3 within 10 minutes of oral administration. Late-release formulations begin releasing a significant amount of Compound 3 after approximately 2-4 hours. The two formulations can be mixed so that they can be taken together in a single dosage form. The ratio of the dose of Compound 3 formulated for early release to the dose formulated for late release is at least 1:2 and can range up to 1:8 (e.g., 1:2, 1:3, 1:4, 1:5, 1:6, 1:7, 1:8). In one embodiment, both the early and late-release dosage components are formulated as microbeads. Microbeads with two release profiles can be manufactured separately and then mixed together in the desired ratio to produce a dosage form (e.g., a sachet). This approach facilitates the manufacture of doses with different ratios of early- to late-release microbeads. Different ratios of the two types of microbeads can be used to individualize treatment for patients.

[0121] In some embodiments, Compound 3 is formulated with three release profiles: early, mid-, and late. The early-release component begins releasing Compound 3 within 10 minutes of oral administration, the mid-release component begins releasing a significant amount of Compound 3 approximately 2-4 hours after ingestion, and the late formulation begins releasing approximately 3-6 hours after ingestion. The three release components can be mixed so that they can be taken together in a single dosage form. The ratio of Compound 3 in the three dosage components (early:mid:late) is at least 1:2:2. The amount of Compound 3 in the mid- and late components can independently vary between 2-8 times the amount of the early component, while the late release component is at least equal to the amount of the mid-release component (e.g., 1:2:8, 1:4:6, 1:4:4, 1:5:5, 1:6:8, etc.). In one embodiment, the early, intermediate, and late release dosage components are all formulated as microbeads, which can be manufactured separately and then mixed together in a desired ratio (e.g., a ratio customized for gastrointestinal and hepatic physiology) to produce a dosage form (e.g., a sachet).

[0122] In certain embodiments, the later dose component, or both the middle and later dose components, are formulated for extended retention in the stomach (gastroretentive formulation). In other embodiments, the later dose component, or both the middle and later dose components, are formulated for sustained release. In certain embodiments, the two-component or three-component dosage form is taken with a meal, preferably a meal containing at least 500 calories, more preferably at least 700 calories. Preferably, the meal is nutritionally complex (e.g., containing several types of whole foods) with at least 25% of the caloric content coming from fat.

[0123] In these embodiments, in which Compound 3 is co-administered with at least one additional cysteamine precursor, the compound is formulated to provide a release profile that complements the release profile of the at least one other cysteamine precursor, so that the cysteamine precursors together provide plasma cysteamine concentrations within the therapeutic range for as long as possible. In preferred embodiments, Compound 3 provides cysteamine in the first 1 to 3 hours after administration, and the at least one additional cysteamine precursor provides cysteamine for 3 to 6, 3 to 8, 4 to 10, or 3 to 12 hours, for example, over a 12-hour dosing interval. In such embodiments, Compound 3 may be formulated for immediate release. In certain embodiments, the at least one additional cysteamine precursor co-administered with Compound 3 is Compound 1 or a pharmaceutically acceptable salt thereof.

[0124] Stimulators of cysteamine production from cysteamine precursors The methods and compositions of the present invention can utilize promoters of cysteamine production.Additional flexibility in controlling blood levels of cysteamine can be achieved by combining cysteamine precursors with promoters of the steps required to chemically and enzymatically decompose cysteamine precursors to cysteamine in the intestine, to absorb cysteamine into the blood, and to prevent cysteamine from being rapidly catabolized in the intestine, blood, or tissues.Specific promoters exist for each of these several steps.Therefore, any of the cysteamine precursors described herein can be optionally co-formulated or co-administered with agents that promote cysteamine production or intestinal uptake, or slow cysteamine degradation, or administered sequentially.

[0125] The first step in converting disulfide cysteamine precursors to cysteamine is the reduction of the disulfide to produce two thiols. The redox environment in the gastrointestinal tract may not contain enough reducing equivalents to quantitatively reduce cysteamine precursors to their respective thiols, thereby limiting cysteamine production. For example, concentrations of the reducing agents glutathione and cysteine ​​in gastric fluid are very low or undetectable (Nalini et al., Biol Int. 32:449 (1994)). Furthermore, in a small clinical study of high-dose pantethine, much of the pantetheine was excreted unchanged in the feces, apparently reflecting incomplete disulfide bond reduction (Wittwer et al., J. Exp. Med. 76:4 (1985)). To address this potential limitation, reducing agents can be coadministered or coformulated with disulfide cysteamine precursors, or administered before or after the cysteamine precursors, making them available when and where needed. The reducing agent can promote the reduction of the disulfide bond, liberating two thiols, or it can promote a thiol-disulfide exchange reaction in which a thiol (A) and a disulfide (BC) react to produce a new disulfide (AB or AC) and a thiol (B or C), thereby releasing one of the thiols in the original disulfide (e.g., cysteamine, pantetheine, or a compound that can be decomposed to cysteamine).

[0126] Various reducing agents can be used to promote disulfide reduction or thiol-disulfide exchange in the gastrointestinal tract. The reducing agent can directly reduce the disulfide cysteamine precursor, or in turn reduce the disulfide cysteamine precursor or other disulfides, such as glutathione disulfide, involved in thiol-disulfide exchange. In some embodiments, physiological compounds (i.e., substances normally found in the body) or food-derived compounds with reducing ability can be used to promote the reduction of the disulfide cysteamine precursor or promote the thiol-disulfide exchange reaction. Physiological reducing agents such as the thiol glutathione or cysteine ​​(all of which are present in the small intestine as a result of bile and enterocyte secretion) can be used, as can other compounds that may normally be present in the body and in food, such as ascorbic acid (vitamin C), tocopherol (vitamin E), or dithiol dihydrolipoic acid, strong reducing agents. Other widely available reducing agents can also be used, including thiols such as N-acetylcysteine ​​and non-thiols such as nicotinamide adenine dinucleotide (NADH), as well as any of the thiols listed in Figure 17. Preferred reducing agents include those known to be safe at the doses required to produce a change in the local gastrointestinal redox environment. Up to several grams of reducing agent, e.g., 0.5 to 5 grams, may be required per administration period. Disulfide cysteamine precursors that can benefit from simultaneous administration of reducing agents are shown in Figure 13. In particular, compounds 1-3 can benefit from simultaneous administration or appropriate subsequent administration of one or more reducing agents, as described herein. Two or more reducing agents may also be combined. Preferably, the reducing agent has a molecular weight of less than 300 daltons.

[0127] Adults produce 400 to over 1,000 milliliters (mL) of bile daily, with 750 mL being estimated as the average volume (Boyer, Compr. Physiol. 3:32 (2013)). Bile is produced throughout the day by the liver. Some is stored in the gallbladder, while the remainder provides a steady, slow flow of bile, even in the fasting state (bile serves excretory functions and aids in digestion and fat absorption). Meals stimulate duodenal secretion of the peptide hormones secretin and cholecystokinin, which stimulate bile production and gallbladder contraction, respectively. The concentration of thiols in bile is approximately 4 mM and consists primarily of glutathione, but also gamma-glutamylcysteine, cysteinylglycine, and cysteine ​​(Eberle et al., J. Biol. Chem. 256:2115 (1981); Abbott and Meister, J. Biol. Chem 258:6193 (1984)).

[0128] Cysteine ​​and, to a lesser extent, glutathione, are also secreted into the lumen of the gastrointestinal tract by enterocytes to regulate luminal redox potential. Thiol concentrations in intestinal fluid from rat jejunum have been measured directly, independently of contributions from bile. They range from 60 to 200 μM in fasted rats and 120 to 300 μM in fed animals (Hagen et al., Am. J. Physiol. 259:G524 (1990); Dahm and Jones, Am. J. Physiol. 267:G292 (1994)). Furthermore, unlike bile secretion, maintenance of luminal thiol levels is a dynamic process, such that increases in intestinal levels of oxidized molecules (e.g., disulfide cysteamine precursors) can be countered, at least to some extent, by increased cysteine ​​production by enterocytes (Dahm and Jones, J. Nutr. 130:2739 (2000)). The human small intestine secretes approximately 1.8 liters of fluid per day, and the colon approximately 0.2 liters, for a total of approximately 2 liters. The concentration of thiols (mainly cysteine) in the secretions varies with the region of the gastrointestinal tract, the redox capacity of the lumen, and the diet.

[0129] The total concentration of gastrointestinal thiols (both bile and enterocyte-derived) influences the rate and extent of disulfide bond reduction and / or thiol-disulfide exchange required to convert cysteamine precursors to thiols, which are the essential first step in their degradation to cysteamine. The amount of reducing equivalents available in the upper gastrointestinal tract after a meal can be estimated by making several assumptions. For example, assuming (i) 200 mL of bile is secreted 1 hour after a large meal and an additional 100 mL 2–3 hours later, and (ii) the thiol concentration in bile is 4 mM, the milliequivalent of thiol reducing power in bile is 0.3 L × 0.004 mol / L = 0.0012 moles of thiol (1.2 mmol). Furthermore, assume that small intestinal enterocytes secrete an additional 400 milliliters during the 4 hours after a meal, resulting in an additional 0.4 liters × 0.0002 moles / liter = 80 micromoles of luminal thiol at a 200 μM thiol concentration. Combined with bile thiol, a total of approximately 1.28 millimoles is available to reduce dietary disulfides and maintain the intestinal redox potential. This is not an estimate of the upper limit of thiol secretion (which may be considerably larger), but rather a value for the normal level of thiol in the small intestine several hours after a meal.

[0130] A 0.5 gram dose of cysteamine-(R)-pantetheine disulfide (MW: 353.52 g / L) contains approximately 1.41 mmol of disulfide bonds and therefore can, in principle, be converted to thiols (either via disulfide bond reduction or thiol-disulfide exchange) by endogenous levels of thiols (ignoring the need for luminal thiols for other physiological purposes).

[0131] More commonly, a cysteamine precursor dose in excess of 1.25 millimolar can benefit from the coadministration of an exogenous reducing agent. Many natural products commonly present in food can provide reducing power to promote the reduction of cysteamine precursors or thiol-disulfide exchange, including the major endogenous intestinal thiols cysteine ​​or glutathione. Cysteine ​​or glutathione analogs such as N-acetylcysteine, N-acetylcysteine ​​ethyl ester, or N-acetylcysteine ​​amide can also be used. Ascorbic acid is another agent capable of reducing disulfide bonds (Giustarini et al., Nitric Oxide 19:252 (2008)). For example, the dose of ascorbic acid required to provide reducing power equivalent to 1 gram of the disulfide cysteamine precursor cysteamine-(R)-pantetheine disulfide can be calculated as follows:

[0132] The molecular weight of ascorbic acid (176.12 g / mol) is approximately half that of cysteamine-(R)-pantetheine disulfide (353.52 g / mol), also known as compound 1. Thus, one gram of ascorbic acid has an equimolar reducing equivalent to the number of disulfide bonds in a 2-gram dose of compound 1. Although the U.S. Food and Nutrition Board's recommended daily intake of vitamin C is only 75 milligrams for women and 90 milligrams for men, many people take much higher doses, including doses of 1 gram or more per day, with apparently fewer or no adverse effects.

[0133] Similar reasoning derives the amount of other reducing agents needed to match the dose of Compound 1 in molar terms. For example, cysteine ​​(molecular weight: 121.15 daltons) is approximately 34% of the mass of Compound 1, N-acetylcysteine ​​(molecular weight: 163.195 daltons) is approximately 46% of the mass of Compound 1, alpha-lipoic acid (molecular weight: 208.34 daltons) is approximately 59% of the mass of Compound 1, and so on. Alpha-lipoic acid and N-acetylcysteine ​​are widely available in 600 and 1,000 mg capsules and tablets, respectively, including extended-release formulations, which exhibit unregulated status, both in vitamin stores and online. Similar calculations can be performed for other disulfide cysteamine precursors based on their molecular weights.

[0134] Because bile is the primary source of thiols and bile is continuously diluted along the length of the small and large intestines, excess reducing power for the reduction of cysteamine precursors may be more available in the jejunum, ileum, or colon than in the duodenum. Therefore, formulations designed to release reducing agents in the distal small intestine and / or large intestine may be particularly useful supplements for disulfide cysteamine precursors. Sustained-release formulations of ascorbic acid and other reducing agents are commercially available. Alternatively, ascorbic acid can be co-formulated with cysteamine precursors to ensure simultaneous delivery of both agents.

[0135] Although the electrochemical capacities (reducing powers) associated with different biological reducing agents are known and provide guidance for their use, the ability of such agents to reduce different disulfide cysteamine precursors is best determined empirically.

[0136] The reaction rate of thiol-disulfide exchange reactions is strongly affected by pH (i.e., slowed by low pH). Such exchange reactions are an alternative mechanism to disulfide bond reduction to liberate cysteamine from cysteamine mixed disulfide or pantetheine from pantethesine disulfide. To increase the reaction rate of thiol-disulfide exchange reactions, basic compounds may be co-administered or co-formulated with disulfide cysteamine precursors, so that they are available when and where needed. Physiological compounds such as bicarbonate are present in high concentrations in pancreatic juice and can be used to regulate local gastrointestinal pH.

[0137] An essential step in the conversion of many cysteamine precursors to cysteamine is the enzyme pantetheinase, encoded by the human VNN1 and VNN2 genes. Pantetheine and pantetheine disulfide (including pantethine) require this enzyme to produce cysteamine. Pantetheinase is also ultimately required for the production of cysteamine from compounds convertible to pantetheine in the gastrointestinal tract, such as 4-phosphopantetheine, dephosphocoenzyme A, coenzyme A, and appropriate analogs and derivatives. Normal levels of pantetheinase in the gastrointestinal tract may not be sufficient to quantitatively cleave all pantetheine molecules provided by a pharmacological dose. To address this limitation, compounds that induce pantetheinase expression can be coadministered or coformulated with cysteamine precursors containing pantetheine or compounds convertible to pantetheine to increase the amount of pantetheinase in the gastrointestinal tract at the time and place needed (i.e., when and where pantetheine is present). Agents that induce pantetheinase expression include both physiological substances, including certain dietary components, and pharmaceutical agents, including FDA-approved drugs.Physiological inducers of VNN1 include various substances that act through the transcription factors NF-E2-related factor-2 (more commonly referred to by the acronym Nrf2), peroxisome proliferator-activated receptor alpha (PPAR alpha), and peroxisome proliferator-activated receptor gamma (PPAR gamma).

[0138] Factors that induce Nrf2 activation (through nuclear translocation) include both natural products and certain drugs. For example, sulforaphane, an isothiocyanate found in cruciferous vegetables such as broccoli, Brussels sprouts, cabbage, and cauliflower, induces VNN1 expression via Nrf2. Sulforaphane-rich foods (e.g., Brussels sprouts) can be used to induce pantetheinase expression, or sulforaphane can be administered as a pure substance in a pharmaceutical composition. Certain food-derived thiols, including S-allyl cysteine ​​and diallyl trisulfide (both present in onions, garlic, and garlic extracts), also induce Nrf2 and can be included in the diet along with cysteamine precursors. Alternatively, either compound can be obtained in pure form and administered in a pharmaceutical composition. Lipids present in certain foods, including some polyunsaturated fatty acids, oxidized fats, omega-3 fatty acids, and the naturally occurring lipid oleylethanolamide (OEA), also induce Nrf2 and / or PPARα. Foods rich in oxidized fats, such as French fries and other fried foods, can be co-administered with cysteamine precursors, which require pantetheinase cleavage to produce cysteamine. Omega-3 fatty acids are present in fish and are available in fish oil extracts and in pure form for use in pharmaceutical compositions.

[0139] Naturally occurring PPAR alpha ligands include endogenous compounds such as leukotriene B4, 8-hydroxyeicosatetraenoic acid, and arachidonic acid and arachidonic acid metabolites, including certain family members.Pharmacological PPAR alpha ligands include fibrates (e.g., benzafibrate, ciprofibrate, clinofibrate, clofibrate, fenofibrate, gemfibrozil), pirinixic acid (Wy14643), and di(2-ethylhexyl) phthalate (DEHP).Any natural or synthetic PPAR alpha ligand can be co-formulated or co-administered with cysteamine precursors that require pantetheinase cleavage to produce cysteamine.For a general review of PPAR ligands, see Grygiel-Gorniak, B. Nutrition Journal 13:17 (2014).

[0140] Natural and synthetic PPARG agonists can also be used to stimulate Nrf2-mediated transcription of the pantetheinase genes VNN1 and / or VNN2. Natural product PPARG agonists include arachidonic acid and metabolites, including 15-hydroxyeicosatetraenoic acid (15(S)-HETE, 15(R)-HETE, and 15(S)-HpETE), 9-hydroxyoctadecadienoic acid, 13-hydroxyoctadecadienoic acid, 15-deoxy-(delta)12,14-prostaglandin J2, and prostaglandin PGJ2, as well as honokiol, amorfurthin 1, amorfurthin B, and amorphastilbole. Other natural products that activate both PPARG and PPARA include genistein, biochanin A, sargaquinoic acid, sargahydroquinoic acid, resveratrol, and amorphastilbole. Natural product PPARG agonists are described and reviewed in Wang et al., Biochemical Pharmacology 92:73 (2014). Pharmacological PPAR gamma agonists include thiazolidinediones (also known as glitazones, e.g., pioglitazone, rosiglitazone, and lobeglitazone). Heme from red meat also induces VNN1 expression. PPARA or PPARG agonists that stimulate pantetheinase expression can be coadministered or coformulated with cysteamine precursors containing pantetheine or compounds that can be degraded to pantetheine in the intestine. Two or more inducers of pantetheinase expression can be combined to enhance expression or reduce the dose of any single agent.

[0141] Another important step in making cysteamine bioavailable throughout the body is absorption across the intestinal epithelium. Cysteamine uptake from the intestinal lumen is mediated by transporters, whose natural levels may not be high enough to transport all cysteamine in the intestinal lumen. Therefore, compounds that induce cysteamine transporter expression can be coadministered or coformulated with cysteamine precursors to enhance cysteamine absorption. Cysteamine is transported across the intestinal epithelium by organic cation transporters 1, 2, and 3 (encoded by the OCT1, OCT2, and OCT3 genes, also known as SLC22A1, SLC22A2, and SLC22A3 genes), and possibly other transporter proteins. Inducers of organic cation transporter expression include the transcription factors PPAR alpha and PPAR gamma, pregnane X receptor (PXR), retinoic acid receptor (RAR), and (in the case of OCT1) RXR receptors, as well as glucocorticoid receptors. Therefore, either natural or synthetic ligands for these receptors can be used to increase OCT expression and, consequently, promote cysteamine uptake by intestinal epithelial cells. Agents that stimulate the expression of cysteamine transporter(s) may be co-administered or co-formulated with any type of cysteamine precursor.

[0142] The elimination half-life of cysteamine in the human body (the time from Cmax to half Cmax after an intravenous bolus) is approximately 25 minutes. A portion of a cysteamine dose is converted to various disulfides, including mixed disulfides with free cysteine, cysteinyl residues in proteins, and glutathione. Pharmacological intervention cannot prevent this excretion process, and in either case, a pool of cysteamine remains available for further disulfide exchange. However, there is a cysteamine catabolic pathway that irreversibly converts cysteamine and effectively removes it from the body. The enzyme cysteamine dioxygenase, which oxidizes cysteamine to hypotaurine, is an important factor in cysteamine excretion. Hypotaurine is then further oxidized to taurine. Coadministration of a cysteamine precursor with one or both of these catabolic products can slow cysteamine catabolism by inhibiting the end product. Thus, in certain embodiments, the cysteamine precursor is co-formulated, co-administered, or administered in an optimal temporal sequence with hypotaurine and / or taurine.

[0143] Figure 13 shows a classification of cysteamine precursors based on the thiol constituent, the number of cysteamine molecules that can be produced, the metabolic steps required to produce cysteamine, potentially useful enhancers of in vivo cysteamine production, and cysteamine release profiles. Compounds (not shown in Figure 13) that induce higher expression of cysteamine transporter(s) are useful for all types of cysteamine precursors. Compounds (not shown in Figure 13) that alkalinize intestinal contents, thereby promoting thiol-disulfide exchange and / or reduction of disulfide bonds, are useful for disulfide cysteamine precursors.

[0144] In summary, flexibility in controlling cysteamine blood levels can be achieved by co-combining or co-administering (i) one or more cysteamine precursors with selected properties, (ii) one or more enhancers of in vivo cysteamine precursor degradation and / or cysteamine absorption, (iii) one or more inhibitors of cysteamine catabolism, (iv) one or more formulations (e.g., immediate, delayed, sustained, gastroretentive, or colon-targeted, or a combination), and (v) a dosing schedule that allows optimal simultaneous delivery of the cysteamine precursor(s) and enhancer(s) to targeted segments of the gastrointestinal tract in amounts that can be effectively degraded and absorbed. The result of personalized application of these tools is maintenance of cysteamine blood levels within the therapeutic range for extended periods, resulting in superior pharmacological effects against diseases compared to existing compounds and formulations.

[0145] Pharmaceutical Composition The present invention provides compositions formulated to achieve therapeutically effective plasma concentrations of cysteamine over extended periods of time to (i) reduce side effects associated with high peak cysteamine concentrations, (ii) reduce therapeutic underperformance caused by subtherapeutic trough concentrations of cysteamine, and (iii) improve patient convenience and therefore compliance with therapy by reducing the number of daily doses. The compounds and formulations of the present invention are also designed to (i) provide improved organoleptic properties compared to existing cysteamine formulations, (ii) reduce contact of free cysteamine with the gastric epithelium, a known cause of gastrointestinal side effects, and (ii) minimize the dose of cysteamine precursor required to achieve therapeutic cysteamine blood levels by matching the dose and delivery site(s) with the relevant digestion and absorption processes in the gastrointestinal tract, which may be achieved by (iii) optimizing the degradation and absorption of cysteamine precursors by co-formulation or co-administration with enhancers of those processes.

[0146] In the case of the compositions of the present invention, pharmaceutical excipients are included in all formulations to prevent exposure of the cysteamine precursor or its salt in the mouth. Formulation methods for masking bitter or other unpleasant tastes include coatings that can be applied in several layers. Flavoring agents and dyes can also be used. Methods for producing pharmaceutical compositions with acceptable mouthfeel and / or taste are known in the art (see, for example, pharmaceutical formulation textbooks cited elsewhere). The patent literature also provides methods for producing organoleptically acceptable pharmaceutical compositions (see, for example, U.S. Patent Publication No. 2010 / 0062988).

[0147] Gastric retentive composition The first composition provides cysteamine precursor or its salt in gastroretentive formulation.Various gastroretentive technologies are known in the art, and some of them have been successfully used in commercial products.For review, see, for example, Pahwa et al., Recent Patents in Drug Delivery and Formulation, 6:278 (2012) and Hou et al., Gastric retentive dosage forms: a review.Critical Reviews in Therapeutic Drug Carrier Systems 20:459 (2003).

[0148] Gastric retention formulations provide sustained release of cysteamine precursors in the stomach. Depending on the type of cysteamine precursor, subsequent in vivo cysteamine production may begin in the stomach or the small intestine, the tissue where cysteamine is most efficiently absorbed. Some cysteamine precursors may continue to be converted to cysteamine in the large intestine, even if released from a pharmaceutical composition in the stomach or small intestine. For example, disulfide cysteamine precursors released in the stomach may remain primarily in an oxidized state in the acidic, oxidizing environment of the stomach and begin to release cysteamine after encountering a reducing agent (e.g., bile glutathione) in the small intestine. Gastric retention compositions produce elevated blood cysteamine levels for 1 to 4 hours, preferably 1 to 6 hours, more preferably 1 to 8 hours, 1 to 10 hours, or longer after ingestion.

[0149] Contrary to recommendations for cysteamine bitartrate (see, e.g., Procysbi® FDA full prescribing information), gastroretentive formulations of cysteamine precursors should be administered with food, preferably with a meal containing sufficient caloric content and nutrient density to delay gastric emptying. Nutrient-dense meals induce osmoreceptors and chemoreceptors in the small intestine (and to a lesser extent in the stomach), which have the effect of stimulating neural and hormonal signals that reduce gastric motility and thereby delay emptying. Delaying gastric emptying is a mechanism for prolonging the effects of gastroretentive compositions. However, filling the stomach with large amounts of food or liquid tends to promote gastric motility and increase the emptying rate, and therefore nutrient density is a more important dietary characteristic than volume. Solid foods that must be broken down into small particles in the antrum and pylorus before emptying into the duodenum prolong gastric retention compared to liquid or semi-liquid foods. Among liquid foods, high-viscosity liquids can delay gastric emptying compared to low-viscosity liquids. Foods with high osmolality contents trigger duodenal osmoreceptors, signaling gastric emptying to slow. Release of cysteamine precursors in the stomach (e.g., from gastroretentive formulations) can increase the osmolality of the gastric and, therefore, duodenal contents.

[0150] In certain embodiments, disulfide cysteamine precursors are preferred for gastroretentive formulations because the acidic, oxidizing environment of the stomach tends to maintain the disulfide in an oxidized form, thereby limiting the exposure of the gastric epithelium to cysteamine, which is thought to be one of the causes of cysteamine toxicity. Upon entering the duodenum and mixing with bile, which contains high (millimolar) concentrations of glutathione, cysteine, and other reducing agents, the disulfide is reduced, thereby generating free thiols where it is exposed to pantetheinase and where cysteamine transporters are expressed on enterocytes.

[0151] The presence of fat in the small intestine is the most potent known inhibitor of gastric emptying, leading to a decrease in the relaxation and contraction of the proximal stomach in the pylorus region.Once fat is absorbed in the small intestine and no longer induces an inhibitory signal to the stomach, gastric motility resumes its normal pattern.Therefore, gastroretentive preparations can ideally be administered with a meal containing fatty foods.Protein-rich meals also slow gastric emptying, to a lesser extent, and carbohydrate-rich meals slow gastric emptying, to an even lesser extent.

[0152] The gastroretentive composition can also be administered with compounds that slow gastric emptying, including certain lipids; for example, fatty acids with at least 12 carbon atoms stimulate cholecystokinin release from enteroendocrine cells and reduce gastric motility, while fatty acids with shorter carbon atoms are less effective. In some embodiments, food or meals can be supplemented with fatty acids or triglycerides containing fatty acids with carbon chains of 12 or more (e.g., oleic acid, myristic acid, triethanolamine myristate, fatty acid salts).

[0153] When fat and protein reach the duodenum, they stimulate the secretion of several gut hormones, including ghrelin, cholecystokinin (CCK), and glucagon-like peptide 1 (GLP1). CCK delays gastric emptying by binding to the CCK1 receptor (CCK1R, formerly known as the CCK-A receptor). In some embodiments, an orally active CCK agonist or mimetic, a positive allosteric modulator of CCK1R, or an agent that promotes the release of endogenous CCK or inhibits CCK degradation, or an agent that otherwise prolongs CCK action through some combination of these or other mechanisms, is administered with the gastroretentive composition to delay gastric emptying and prolong the gastric retention of the gastroretentive composition. CCK is a peptide that exists in several forms ranging from 8 to 53 amino acids (e.g., CCK-8, CCK-53). Oral administration of peptides is ineffective because they are digested in the gastrointestinal tract. Small molecule CCK agonists have been developed and tested by several research groups, for example, SR-146, 131 and related compounds were developed by scientists at Sanofi (U.S. Patent Nos. 5,731,340 and 6,380,230, incorporated herein by reference).

[0154] Certain protease inhibitors induce CCK production or release, extend its half-life, or otherwise enhance its effects, including both food-derived mixtures and pure compounds. For example, the ingestion of potato-derived protease inhibitor concentrates is associated with elevated CCK levels, as is the ingestion of soybean peptone and soybean beta-conglycinin peptone. Camostat is a synthetic protease inhibitor with pleiotropic effects, including stimulating endogenous CCK release and consequently slowing gastric emptying. Camostat mesylate is a pharmaceutical salt widely used in humans. FOY-251 is the active metabolite of camostat. In some embodiments, an agent that stimulates CCK production or release, extends CCK half-life, or otherwise enhances CCK effects is co-formulated or co-administered with a gastric-retentive composition in an amount that delays gastric emptying. In some embodiments, camostat, FOY-251, or a prodrug, derivative, or active metabolite of camostat, or a pharmaceutically acceptable salt thereof, is co-formulated or co-administered with the gastroretentive composition in an amount ranging from 50 to 300 mg / kg or 100 to 250 mg / kg.

[0155] Gastric emptying is also slowed by acidifying the chyme. For example, citric acid and acetic acid have been shown to delay gastric emptying. In some embodiments, the food or meal includes a natural source of citric acid (e.g., the pulp or juice of oranges, lemons, limes, grapefruits, or other citrus-rich fruits), or acetic acid (e.g., vinegar, pickles, or other pickled vegetables), or lactic acid (e.g., sauerkraut or kimchi). In some embodiments, a sufficient amount of acidic food or liquid is administered with the gastric retention composition to lower the pH of the gastric chyme below pH 4 or below pH 3.5.

[0156] Glucagon-like peptide-1 (GLP1) is another gut hormone released by cells in the duodenum in response to food, particularly ingested fat, and influences gastric emptying. Orally administered GLP1 receptor agonists have been discovered by several research groups (e.g., Sloop et al., Diabetes 59:3099 (2010)). Positive allosteric modulators of the GLP1 receptor (which are not agonists per se but enhance endogenous GLP1) are another category of GLP1R stimulants (see, e.g., Wootten et al., J. Pharmacol. Exp. Ther. 336:540 (2011); Eng et al., Drug Metabolism and Disposition 41:1470 (2013); also see U.S. Patent Nos. 2006 / 0287242, 2007 / 0021346, 2007 / 0099835, 2013 / 0225488, and 2013 / 0178420, each of which is incorporated herein by reference). Among compounds that positively regulate GLP-1 receptor signaling in the presence of endogenous GLP1 is quercetin, which acts by binding to an allosteric site on the GLP-1 receptor and positively influencing receptor signaling upon binding of an endogenous ligand (GLP-1, a peptide, exists in several forms). Some quercetin analogs are also positive regulators of endogenous GLP1. Quercetin is a flavonol found in many fruits, vegetables, leaves, and grains. It is used as an ingredient in dietary supplements, beverages, and foods. In some embodiments, a GLP-1 receptor agonist or a positive allosteric modulator of GLP-1 is coformulated or coadministered with the gastroretentive composition in an amount sufficient to delay gastric emptying. In some embodiments, the GLP-1 receptor agonist or positive allosteric modulator is quercetin or an analog, derivative, or active metabolite of quercetin. Certain small molecule drugs can also slow gastric emptying time and can be co-administered or co-formulated with the gastroretentive composition.

[0157] Gastric emptying is also slowed by acidifying the chyme. For example, citric acid and acetic acid have been shown to delay gastric emptying. In some embodiments, the food or meal contains a natural source of citric acid (e.g., oranges, grapefruits, or other citrus-rich fruits), or acetic acid (e.g., vinegar, pickles, or other pickled vegetables), or lactic acid (e.g., sauerkraut or kimchi). In some embodiments, the pH of the chyme is reduced to below 4 or below 3.5 by administering an acidic food or liquid with the gastroretentive composition.

[0158] U.S. Patent No. 8,741,885 describes a method for prolonging the gastric retention of a gastroretentive pharmaceutical composition (e.g., a floating, swellable, or mucoadhesive composition) by combining an active pharmaceutical ingredient with an opioid. The purpose of the co-formulated opioid is to delay gastric emptying. Gastroparesis, or severe congestive gastrointestinal motility, is a well-known and potentially serious complication of opioid therapy.

[0159] Sustained Release Compositions The second composition provides a cysteamine precursor or its salt in a non-gastroretentive sustained-release formulation. Sustained-release formulations are well known in the art: Wen, H. and Park, K. (eds.) Oral Controlled Release Formulation Design and Drug Delivery: Theory to Practice. Wiley, 2010; Augsburger, LL and Hoag, SW (eds.) Pharmaceutical Dosage Forms-Tablets, Vol. 3: Manufacture and Process Control. CRC Press, 2008. The sustained-release component can be a tablet, a powder, or a capsule filled with microparticles. Optionally, the particles can differ in size, composition (e.g., type or concentration of sustained-release polymer), or type or thickness of coating agent, or the number and composition of layers when coated with multiple layers of coating agent, so that the drug is released from individual particles at different rates or at different onset times, thereby providing a longer drug release period in the aggregate compared to a formulation in which all particles are substantially identical. Sustained-release formulations may optionally be coated with a pH-sensitive material (called an enteric coating) that prevents dissolution in the stomach. Microparticles in a single composition may have one or more different coating types or thicknesses. For example, the pH at which the coating dissolves may be different. Two or more types of microparticles used in such a mixed composition may be manufactured separately to precise specifications and then blended in a ratio that achieves extended drug release in vivo.

[0160] Sustained-release compositions can provide extended release of cysteamine precursors in the stomach and / or small intestine (but not the former, if enteric-coated), resulting in sustained in vivo cysteamine production. Sustained-release formulations can be designed to release the drug over a period approximately equal to the sum of the mean gastric transit time and small intestinal transit time, e.g., 3-5 hours when administered in a fasted state, or 5-8 hours when administered with food or a meal. Alternatively, sustained-release formulations can be designed to release the drug for longer than the sum of the mean gastric transit time and small intestinal transit time, such that cysteamine precursors continue to be released in the large intestine. In some embodiments, such sustained-release compositions can release cysteamine precursors over a period of 4-8 hours when administered in a fasted state, or 6-10 hours or more when administered with a meal.

[0161] Sustained-release formulations can provide elevated blood cysteamine levels for 1 to 4 hours, preferably 1 to 6 hours, more preferably 1 to 8 hours, and even more preferably 1 to 10 hours or more after ingestion. Sustained-release formulations of cysteamine precursors can be administered with food or between meals, and optionally with enhancers of cysteamine precursor breakdown or cysteamine absorption. Because food tends to inhibit the absorption of free cysteamine, especially fatty foods, it is generally recommended that cysteamine salts be taken on an empty stomach, although small amounts of applesauce or similar foods are acceptable.

[0162] Mixed preparations Some compositions necessarily have two types of formulations: those that primarily control the rate of drug release and those that primarily control the anatomical site of drug release. For example, gastroretentive formulations always contain a drug in a sustained-release formulation. Otherwise, there is no point in long-term gastric retention. However, there are ways to combine immediate-release and sustained-release components into a single gastroretentive formulation. For example, the immediate-release component can form an outer layer that rapidly dissolves or disintegrates in the stomach, leaving a core sustained-release component that remains in the stomach via one or more of the gastric retention mechanisms described herein. However, not all types of formulations can be productively combined. For example, enteric-coated gastroretentive formulations are counterproductive because the gastroretentive formulation is designed to release the drug in the stomach, and gastric release is blocked by a coating that is resistant to dissolution in an acidic medium.

[0163] Compositions with different temporal or anatomical drug release profiles, when combined with appropriate cysteamine precursors and, optionally, enhancers of cysteamine production or absorption, provide blood cysteamine levels within the therapeutic range for 0.5 to 6 hours, more preferably 0.5 to 8 hours, and most preferably 0.5 to 12, 0.5 to 15 hours, or longer. Examples of productive combination formulations include mixed formulations containing up to two drug-releasing components, as well as separately formulated compositions that can be combined in various amounts and ratios to tailor the amount and timing of in vivo cysteamine production and absorption to individual patient needs.

[0164] A third composition provides a co-formulation of a first enteric-coated component formulated for delayed release of a cysteamine precursor or salt thereof in the small intestine and a second component of enteric-coated microparticles formulated for sustained release of a cysteamine precursor or salt thereof throughout the small intestine and proximal portions of the large intestine, the co-formulation providing the first component to initially achieve elevated blood cysteamine levels and the second component to maintain blood cysteamine levels over time.

[0165] The fourth composition provides a mixed formulation comprising (i) a sustained-release gastroretentive formulation of a cysteamine precursor or its salt, and (ii) an immediate-release formulation of a cysteamine precursor or its salt designed to release the drug in the stomach. The second component of the mixed formulation is located on the outer surface of the composition and begins to dissolve immediately upon contact with stomach contents. It is the first to produce cysteamine, although not necessarily in the stomach. The first (gastroretentive) component provides extended release of the cysteamine precursor in the stomach, followed by in vivo cysteamine production throughout the small intestine and, depending on the properties of the cysteamine precursor, in the large intestine. The combined in vivo production and absorption of cysteamine from the two components begins within one hour after administration of the mixed composition and continues for at least five hours, preferably for 8, 10, 12 hours or more within the therapeutic concentration range.

[0166] In the fifth composition, the first component is formulated for immediate release in the stomach and contains a cysteamine precursor, preferably cysteamine mixed disulfide or pantetheine disulfide, or a salt thereof, and the second component is formulated for sustained release of the cysteamine precursor or its salt. The first component is on the outer surface of the composition so that the second component remains intact after dissolution or disintegration of the first component. This mixed formulation of the fifth composition can produce an initial increase in plasma cysteamine concentrations from the immediate-release component, and maintain elevated levels of cysteamine from the second (sustained-release) component with continued in vivo cysteamine production for 6, 8, 10, or more hours. Release of the cysteamine precursor (or several different cysteamine precursors) along the gastrointestinal tract from the stomach to the large intestine allows the amount of cysteamine precursor to be matched to the levels of pantethetinase and cysteamine transporter in all segments of the intestine, thereby maximizing cysteamine production and absorption. Continuous intestinal production and absorption of cysteamine avoids reliance on a high Cmax to prolong exposure, thereby reducing the cysteamine side effects associated with high peak levels. Thus, mixed formulations of cysteamine precursors allow for the administration of cysteamine to a number of disorders sensitive to the effects of cysteamine.

[0167] In the sixth composition, the first component is formulated for immediate release in the stomach and contains a cysteamine precursor, preferably cysteamine mixed disulfides or pantetheine disulfide, or a salt thereof, and the second component is formulated for sustained release of the cysteamine precursor or a salt thereof in the ileum and / or colon. This mixed formulation of the sixth composition can produce an initial rise in plasma cysteamine levels from the immediate-release component, followed by a second rise in plasma cysteamine levels from the ileum and colon-targeted component, around the time that the first peak rapidly declines. The second component can begin releasing the cysteamine precursor 4 to 8 hours after administration, depending on whether it is administered with or without food. The controlled release of the cysteamine precursor (or a different cysteamine precursor) along the gastrointestinal tract, from the stomach to the large intestine, allows the amount of cysteamine precursor to be matched with the levels of pantethetinase and cysteamine transporter in all segments of the intestine, maximizing cysteamine production and absorption.

[0168] compound The pharmaceutically acceptable composition of the present invention comprises one or more cysteamine precursors or its pharmaceutically acceptable salt(s).The salt of the present invention includes, but is not limited to, salts of alkali metals, such as sodium and potassium; salts of alkaline earth metals, such as calcium, magnesium, and barium; and salts of organic bases, such as amine bases and inorganic bases.Exemplary salts can be found in Remington's Pharmaceutical Sciences, 17th ed., Mack Publishing Company, Easton, Pa., 1985, p.1418; Berge et al., J. Pharmaceutical Sciences 66:1 (1977); and Pharmaceutical Salts: Properties, Selection, and Use (P.H. Stahl and C.G. Wermuth, eds.), Wiley-VCH, 2008, each of which is incorporated herein by reference in its entirety.

[0169] The compositions of the present invention can contain a cysteamine precursor or its salt in a gastroretentive formulation or mixed formulation to achieve a plasma cysteamine concentration within the therapeutic range within the first four hours after administration, preferably within the first two hours after administration, and most preferably within the first hour. The plasma cysteamine concentration preferably remains within the therapeutic range for at least five hours, preferably six hours, more preferably eight, ten hours, or more. The formulation may contain a thiol cysteamine precursor that can be enzymatically degraded to produce cysteamine, such as pantetheine, or a compound that can be degraded to pantetheine (and then cysteamine) in the gastrointestinal tract, such as 4-phosphopantetheine, dephosphocoenzyme A, or coenzyme A, or a derivative or prodrug thereof that can be degraded to pantetheine (and then cysteamine) in the gastrointestinal tract. Alternatively, the cysteamine precursor may be formed by reacting cysteamine or a compound that can be degraded to produce cysteamine with another thiol-containing organosulfur compound to form a disulfide compound. Disulfide cysteamine precursors or salts thereof can be prepared by reacting cysteamine with a thiol cysteamine precursor, such as pantetheine, 4-phosphopantetheine, dephosphocoenzyme A, coenzyme A, or N-acetylcysteamine, or by reacting cysteamine with N-acetylcysteine ​​(NAC), N-acetylcysteine ​​amide, N-acetylcysteine ​​ethyl ester, homocysteine, glutathione (GSH), allyl mercaptan, furfuryl mercaptan, benzyl mercaptan, or thioterpineol. It can be formed by reacting with other thiols, including (grapefruit mercaptan), 3-mercaptopyruvate, L-cysteine, L-cysteine ​​ethyl ester, L-cysteine ​​methyl ester, thiocysteine, cysteinylglycine, gamma-glutamylcysteine, gamma-glutamylcysteine ​​ethyl ester, glutadione monoethyl ester, glutathione diethyl ester, mercaptoethylgluconamide, thiosalicylic acid, tiopronin, or diethyldithiocarbamate.Thiol cysteamine precursors or cysteamine can also be reacted with dithiols, such as dihydrolipoic acid, meso-2,3-dimercaptosuccinic acid (DMSA), 2,3-dimercaptopropanesulfonic acid (DMPS), 2,3-dimercapto-1-propanol (dimercaprol), bucillamine, or N,N'-bis(2-mercaptoethyl)isophthalamide (BDTH2), to form disulfide cysteamine precursors. See Figure 17 for a list of thiols that can be used to form disulfide cysteamine precursors, and Figures 18-21 for tables summarizing thiol pairs that can be combined to form disulfide cysteamine precursors. Other thiols suitable for forming cysteamine precursors are known in the art. For example, PCT Patent Publication No. WO 1993 / 006832, which is incorporated herein by reference in its entirety, discloses additional useful thiols not included in FIG. 17, including N,N-dimethylcysteine, thiocholine, aminopropanethiol, aminobutanethiol, and aminopentanethiol, among others.

[0170] Depending on the properties of the cysteamine precursor used (e.g., the number of decomposition steps required to form cysteamine), the formed disulfides may delay cysteamine release in the stomach and / or facilitate its in vivo production and absorption in the small intestine. Figure 13 shows a classification of cysteamine precursors and summarizes selected pharmacologically relevant properties. Figures 18-21 provide information on the cysteamine yield of many disulfide cysteamine precursors. The stomach is generally a more oxidative and acidic environment than the small intestine. As gastric contents enter the duodenum, they mix with pancreatic juice, which contains bicarbonate, which neutralizes gastric acid, and bile, which contains millimolar concentrations of the physiological reducing agent glutathione and related thiols, including cysteine. As a result, disulfides tend to remain oxidized in the stomach and are more likely to be reduced or participate in disulfide exchange reactions with thiols in the small intestine. Disulfide exchange reactions are generally catalyzed by thiolate ions, which are much more nucleophilic than thiol forms, and thiolate ion formation is unfavorable in the acidic environment of the stomach.

[0171] For example, pantetheine, a thiol cysteamine precursor, can form a homodimer disulfide, in which two pantetheines are covalently linked to form pantethine (a disulfide cysteamine precursor). In some preferred embodiments, the cysteamine precursor provides multiple cysteamines, such as by a mixed cysteamine disulfide formed by conjugating cysteamine with pantetheine, 4-phosphopantetheine, dephosphocoenzyme A, or coenzyme A, or by a corresponding mixed pantetheine disulfide formed by oxidizing pantetheine with 4-phosphopantetheine, dephosphocoenzyme A, or coenzyme A, or by a suitable prodrug or analogue that can be converted to the parent compound in the gastrointestinal tract. Also, 4-phosphopantetheine can be disulfide-linked to dephosphocoenzyme A or coenzyme A, or dephosphocoenzyme A can be disulfide-linked to enable the cysteamine precursor to generate two cysteamines in vivo. Figure 13 shows the number of cysteamines that can be generated in vivo from different classes of cysteamine precursors. Figures 18-21 show specific disulfide cysteamine precursors. Those that yield two cysteamines in vivo are listed at the top of the table, and the fractional yield (percent) of cysteamine for each disulfide is also shown, as well as the number of decomposition steps required to generate cysteamine. In some embodiments, the reactive thiol group of cysteamine or organosulfur can be modified to include substituents such as acetyl groups, ester groups, glutamyl, succinyl, phenylalanyl, polyethylene glycol (PEG), and / or folate.

[0172] In a preferred embodiment, the compositions of the present invention contain pantetheine, pantetheine-containing pantetheine, or a salt thereof in a gastroretentive formulation and / or a mixed formulation, and can maintain elevated blood levels of cysteamine for 5 to 10 hours or more after administration. The compositions may be cysteamine precursors that require chemical reduction or enzymatic conversion of at least one parent compound to cysteamine, thereby delaying the release of cysteamine. The formulations may also contain pantetheine or a compound that can be degraded to pantetheine in the gastrointestinal tract (e.g., 4-phosphopantetheine, dephosphocoenzyme A, or coenzyme A; collectively, pantetheine precursors), where the thiol group of pantetheine or the pantetheine precursor reacts with the thiol group of another organosulfur compound to form a disulfide compound. Because pantetheinase is expressed at higher levels in the intestine than in the stomach, and the lumen of the small intestine is a more reducing environment than the stomach, the pantetheine component of the disulfide cysteamine precursor is converted to cysteamine in the small intestine and subsequently absorbed. For example, pantetheine can form a homodimeric disulfide, in which two pantetheines are covalently linked to form pantethine. Pantetheine-containing cysteamine precursors can also include pantetheine mixed disulfides, in which pantethethiol reacts with a thiol group to form a disulfide. In preferred embodiments, the pantetheine precursor provides two or more cysteamines, such as a mixed disulfide formed from cysteamine and pantetheine (which, when reduced and subsequently cleaved by pantetheinase, yields two cysteamines and one pantothenic acid), or a mixed disulfide pantetheine-coenzyme A (which, when reduced, subsequently degraded, and then cleaved by pantetheinase, yields two cysteamines, two pantothenic acids, and ADP). Other disulfide cysteamine precursors that yield two cysteamines upon degradation in the intestine are shown in Figures 18-21.In some embodiments, the reactive thiol group of pantetheine or the organosulfur compound may be modified to include substituents such as acetyl groups, methyl esters, ethyl esters, glutamyl, succinyl, phenylalanyl, polyethylene glycol (PEG), and / or folate.

[0173] The distinction between cysteamine precursors that require pantetheinase cleavage to produce cysteamine and those that require only chemical reduction to produce cysteamine (cysteamine mixed disulfides) is important because the kinetics of precursor compound conversion to cysteamine are generally more rapid in the second category, provided that a suitable reducing environment exists in the intestine (or can be created pharmacologically). A further distinction can be made between cysteamine precursors that require reduction followed by pantetheinase cleavage (e.g., pantethine) and those that require initial reduction, then degradation to pantetheine, followed by pantetheinase cleavage (e.g., 4-phosphopantethine, dephosphocoenzyme A, or coenzyme A-containing disulfides). The additional degradation step(s) required by the latter class of disulfide cysteamine precursors slows and extends the duration of cysteamine production over a longer period.

[0174] The compounds of the present invention can be prepared by various methods known to those skilled in the art of chemical synthesis. Methods for preparing thiols, including cysteamine, pantetheine, 4-phosphopantetheine, dephospho-coenzyme A, or coenzyme A, and other thiols (see Figure 17), are well known in the art. Coenzyme A, pantethine, N-acetylcysteamine, and glutathione are commercially available as dietary supplements. Most of the other thiols in Figure 17 are readily available from chemical companies.

[0175] Synthesis of cysteamine precursors The compounds of the present invention, containing both thiol and disulfide cysteamine precursors, can be prepared from readily available starting materials using methods and procedures known in the art, such as those described in Mandel et al., Organic Letters, 6:4801 (2004). Methods for producing pantethine are described in U.S. Patent Nos. 3,300,508 and 4,060,551, each of which is incorporated herein by reference. Methods for converting liquid pantetheine to a solid are disclosed in Japanese Patent Publications JP-A-S50-88215 and JP-A-S55-38344. Where typical or preferred process conditions (i.e., reaction temperature, time, molar ratio of reactants, solvent, pressure, etc.) are given, it will be understood that other process conditions can also be used unless otherwise specified. Optimum reaction conditions may vary depending on the particular reactants or solvents used, but such conditions can be determined by routine optimization procedures by one of ordinary skill in the art.

[0176] In a preferred embodiment, the composition of the present invention comprises one or more disulfide cysteamine precursors. Disulfides, which are oxidized forms of thiols, can be easily formed from constituent thiols without expensive reagents or equipment. Furthermore, disulfides are not susceptible to oxidation, which can limit the long-term stability of thiol compounds exposed to air. Therefore, disulfide forms of cysteamine precursors are more preferable than thiol forms in terms of production, cost, storage cost, shipping, and patient convenience (i.e., long shelf life).

[0177] In some embodiments, mixed disulfide cysteamine precursors are synthesized by conjugating two different thiols to form three reaction products: thiols A and B can conjugate to form disulfides AA, AB, and BB. For example, disulfides formed by reacting cysteamine with pantetheine include cysteamine-cysteamine (referred to as cystamine), cysteamine-pantetheine, and pantetheine-pantetheine (referred to as pantethine). All three compounds are useful in providing cysteamine, and the different steps involved in converting each compound to cysteamine may be pharmacologically beneficial by extending the period during which cysteamine is produced in vivo by reduction of the disulfide bond or by a combination of reduction and enzymatic degradation steps. Therefore, co-combining all three oxidation products without purification (except for removing unwanted impurities such as unreacted thiols and / or solvents) may be pharmacologically useful. This is particularly true when two reacted thiols are each convertible to cysteamine (e.g., pantetheine, 4-phosphopantetheine, dephosphocoenzyme A, coenzyme A, N-acetylcysteine, or a suitable analog and prodrug), or when cysteamine itself is reacted with a thiol convertible to cysteamine. As a result, in certain embodiments, all three disulfides formed by reacting two different thiols, each convertible to cysteamine (or one of which is cysteamine), are co-formulated into a single composition. This synthesis and formulation method does not require more complex synthesis steps or post-synthesis purification steps required to separate the mixed disulfide from the two homodimeric disulfides simultaneously produced in the oxidation reaction. (Unreacted thiols and other impurities must, of course, be removed before formulating the pharmaceutical composition.)

[0178] The advantages of producing and simultaneously combining three disulfide mixtures are not fully realized in the case of disulfide cysteamine precursors, which are produced by reacting a thiol that can be converted to cysteamine with a second thiol that cannot be converted to cysteamine.For example, the three disulfides formed by reacting pantetheine with N-acetylcysteine ​​(NAC) are pantetheine-pantetheine (pantethine), pantetheine-NAC, and NAC-NAC.The first two compounds are cysteamine precursors, while the third (NAC-NAC) is not.However, NAC-NAC may nevertheless have beneficial pharmacological properties in terms of regulating the intestinal redox environment or beneficial medical properties, as a result of producing two NAC molecules upon chemical reduction. Thus, in certain embodiments, all three disulfide products formed by reacting cysteamine or a thiol convertible to cysteamine in vivo with a second thiol that is not convertible to cysteamine in vivo are co-formulated in a single composition.

[0179] The expected ratio of reaction products when two different thiols are oxidized depends on the molar ratio of the two thiols, the absolute concentrations of the two thiols, pH, and / or the chemical environment surrounding the sulfhydryl group of each thiol. When the ratio of thiol A to thiol B is 1:1, the expected molar ratio of reaction products AA, BB, and AB is approximately 1:1:2. (Deviation from the expected ratio may result from differences in the chemical bonds adjacent to the thiols, which may affect the reaction rate of disulfide bond formation, which may be influenced, for example, by the electronegativity of the atom bonded to the sulfhydryl. Any deviation can be predicted or measured using methods known in the art.) The ratio of reaction products can be altered by changing the molar ratio of the two thiols. For example, to increase the ratio of AA and AB relative to BB, the molar concentration of thiol A may be increased relative to the molar concentration of thiol B. When reacting two thiols, one of which is cysteamine or a compound that can be decomposed into cysteamine (thiol A) and the other is a thiol that cannot be decomposed into cysteamine (thiol B), the molar concentration of the first thiol can be increased relative to the molar concentration of the second thiol to increase the proportion of cysteamine precursor produced. For example, reacting thiols A and B in a 2:1 molar ratio increases the proportion of AA and AB (both cysteamine precursors) to BB (not a cysteamine precursor).

[0180] In certain embodiments, the inclusion of a catalyst can promote the oxidation of two different thiols and / or alter the mixture of reaction products (reviewed in Musiejuk and Witt (2015)). For example, oxidizing agents such as hydrogen peroxide or dimethyl sulfoxide (DMSO), or metals such as copper, manganese, or telluride, or iodine, diethyl azodicarboxylate (or related compounds), or dichlorodicyanoquinone (DDQ) can be added. Optimal solvent systems, catalyst concentrations, and reaction conditions can be empirically determined to achieve optimal catalyst performance.

[0181] In other embodiments, asymmetric disulfides can be produced via a thiol-disulfide exchange reaction between a thiol and a symmetric disulfide. This type of reaction, like the oxidation of two different thiols, provides a mixture of all possible products (symmetric and asymmetric disulfides). However, by providing a molar excess of the symmetric disulfide over the thiol, the formation of the asymmetric disulfide is favored and may even become the major reaction product under optimized conditions. Examples 15 and 16 describe the synthesis of pantetheine-cysteamine disulfide via thio-disulfide exchange. This method utilizes cysteamine as the thiol and pantethine as the disulfide, as well as pantetheine as the thiol and cystamine as the disulfide. In preferred embodiments of the thiol:disulfide exchange reaction, the molar ratio of thiol to disulfide (e.g., cysteamine:pantethine) is 2:1 to 4:1, 2.5:1 to 3.5:1, or 2.7:1 to 3.3:1. In certain embodiments, the solvent is methanol and the reaction time is 1 to 20 hours, or 1 to 12 hours, or 1 to 6 hours. In certain embodiments, the product of the thiol:disulfide exchange reaction (e.g., TTI-0102) is then precipitated (e.g., as described in Example 17).

[0182] Alternatively, in another embodiment, the ratio of cysteamine precursors used in the pharmaceutical composition can be adjusted by combining the three reaction products of the mixed disulfide oxidation reaction with pure disulfides. For example, when the thiols cysteamine (C) and pantetheine (P) are oxidized in a 1:1 molar ratio, they are combined to form three products: CC, PP, and CP in an approximately 1:1:2 ratio. Pure pantethine (PP) can be added to the mixture in any desired amount to extend the mixture's in vivo cysteamine-generating properties. Doubling the starting amount of pantethine results in a 1:2:2 ratio. Adding four times the starting amount of pantethine results in a 1:2:5 ratio.

[0183] Two independently produced mixed disulfide reaction products can also be combined to achieve a new ratio of cysteamine precursors. For example, when cysteamine-pantetheine reaction products (CC, PP, and CP) are combined with equimolar amounts of reaction products from the N-acetylcysteine ​​(NAC)-cysteamine (C) oxidation reaction (CC, NAC-NAC, and C-NAC in a ratio of 1:1:2), the mixture will contain five compounds, one of which, NAC-NAC, cannot be converted to cysteamine. The other four disulfides, PP, CC, CP, and C-NAC, are present in a molar ratio of approximately 1:2:2:2. Optionally, pantetheine can be added to make the ratio, for example, 2:2:2:2 (more simply expressed as 1:1:1:1), or a larger amount can be added to make the ratio 1:1:1:5. Thus, the molar ratio of disulfides in pharmaceutical compositions can be controlled by various methods. In another example, cysteamine-pantetheine reaction products (CC, PP, and CP) may be combined with a 4-phosphopantetheine (4P)-cysteamine (C) oxidation reaction (i.e., CC, 4P-4P, and C-4P in a 1:1:2 ratio) to produce a mixture of five disulfides in a 1:1:1:2:2 ratio.

[0184] In summary, when one thiol is oxidized to produce a cysteamine precursor disulfide, only one product will be present (e.g., pantetheine + pantetheine = pantethine). When two thiols are oxidized, three products will be present, two or three of which will be cysteamine precursors, depending on whether one or both of the thiols are decomposable to cysteamine or cysteamine itself. Mixtures of cysteamine precursors are most easily produced by combining the products of these two types of reactions. Mixtures can contain various molar ratios of pure disulfides or ternary disulfide mixtures. However, heterodimeric cysteamine precursors can also be used in pure form, after purification, or in combination with other homo- or heterodimeric cysteamine precursors.

[0185] Alternatively, more sophisticated chemical methods can be used to selectively synthesize specific mixed disulfides (also called asymmetric disulfides) (e.g., cysteamine and pantetheine can be combined to form essentially only the disulfide cysteamine-pantetheine). These methods employ a wide range of sulfur protecting groups and strategies for their removal. The most widely used method involves replacing a sulfenyl derivative with a thiol or its derivatives. Commonly utilized sulfenyl derivatives include sulfenyl chloride, S-alkyl and S-aryl thiosulfates (Bunte salts), S-(alkylsulfanyl)isothiourea, benzothiazol-2-yl disulfide, benzotriazolyl sulfide, dithioperoxyesters, (alkylsulfanyl)dialkylsulfonium salts, 2-pyridyl disulfides and derivatives, N-alkyltetrazolyl disulfides, sulfenamides, sulfenyl dimesylamine, sulfenyl thiocyanate, 4-nitroarenesulfenanilide, thiolsulfinates and thiolsulfonates, sulfanylsulfinamidine, thionitrite, sulfenyl thiocarbonate, thioimides, thiophosphonium salts, and 5,5-dimethyl-2-thioxo-1,3,2-dioxaphosphorinan-2-yl disulfide. Further methods include the reaction of thiols with sulfinylbenzimidazoles, rhodium-catalyzed disulfide exchange, electrochemical methods, and the use of diethyl azodicarboxylate. These and other methods are reviewed by Musiejuk, M. and D. Witt. Organic Preparations and Procedures International 47:95 (2015). Thus, with modest effort, specific mixed (asymmetric) disulfides of interest can be prepared. Examples 1 and 2 provide synthetic procedures for the mixed disulfides of the present invention.

[0186] In yet other embodiments, mixed disulfides can be synthesized from symmetric disulfides by preferentially attaching a substituent (e.g., an acyl group) to one end of the symmetric disulfide (i.e., hemiacylation). For example, because cysteamine and pantetheine differ in the pantothenate moiety, the disulfide cystamine can be hemiacylated with pantothenate to produce cysteamine-pantetheine disulfide. This procedure can achieve yields of over 95% when the reactant concentrations are optimized and a coupling agent is added to promote acylation. Because cystamine contains reactive amino groups at both ends, it is an attractive starting point for creating asymmetric disulfides. Example 14 describes the efficient synthesis of pantetheine-cysteamine disulfide via hemiacylation of cystamine with the substituent pantothenic acid via a reactive intermediate. In certain embodiments, the molar ratio of acyl group to disulfide is 1:2 to 1:4. In certain embodiments, the acylation reaction is facilitated by the addition of N,N'-dicyclohexylcarbodiimide (DCC) in a molar ratio of DCC:acyl group of 3:1 to 5:1. In certain embodiments, the acylation reaction is facilitated by the addition of 1-hydroxybenzotriazole (HOBt) in a molar ratio of HOBt:acyl group of 1:1 to 1:3.

[0187] stereochemistry Some of the compounds of the present invention exist in multiple enantiomeric forms. In particular, pantetheine, 4-phosphopantetheine, dephosphocoenzyme A, and coenzyme A contain chiral carbon atoms in the pantethenoyl moiety. Therefore, each of these compounds can exist as either a D- or L-enantiomer, or as a racemic mixture of the two with respect to the pantethenoyl group. However, human pantetheinase (encoded by the VNN1 and VNN2 genes) is specific for D-pantetheine. (Bellussi et al., Physiological Chemistry and Physics 6:505 (1974)) Therefore, only D-pantetheine (and not L-pantetheine) is a cysteamine precursor, and therefore the present invention relates only to D-pantetheine and the D-enantiomers of 4-phosphopantetheine, dephosphocoenzyme A, and coenzyme A, as well as analogs or prodrugs that can be converted to these compounds in the gastrointestinal tract. Similarly, pantetheine, 4-phosphopantetheine, dephosphocoenzyme A, and all disulfide containing coenzyme A, or any suitable analog or prodrug, use only the D-enantiomer.

[0188] Preferred is the L-enantiomer of amino acid and amino acid derivative.Therefore, in this specification, "cysteine" refers to L-cysteine, homocysteine ​​to L-homocysteine, and cysteine ​​derivatives such as N-acetylcysteine, N-acetylcysteineamide, N-acetylcysteine ​​ethyl ester, cysteine ​​methyl ester, cysteine ​​ethyl ester, cysteinylglycine and gamma glutamylcysteine ​​are all formed using the L-enantiomer of cysteine.

[0189] In the case of dihydrolipoic acid, the R enantiomer is preferred because it is the enantiomer made in the human body. Generally, for compounds normally present in the human body or present in food, the naturally occurring enantiomer is preferred.

[0190] Salt Form and Crystallization The pharmaceutical properties of any compound, including the cysteamine precursors of the present invention, may be improved by association with a counterion or salt. Specific properties that may be improved include stability (e.g., low hygroscopicity, low susceptibility to oxidation, high resistance to extremes of heat, humidity, and pH), increased tendency to form crystals, and ease of formulation (with respect to the properties of solid forms of the compound, such as powders).

[0191] The potential for salt forms to improve the crystallization properties of compounds is particularly important because crystallization is a cheaper, faster, and more scalable purification method than, for example, column chromatography. Methods for inducing precipitation of small molecules are known in the art (see, for example, the review by Chen et al., Crystal Growth and Design, 11(4), 2011). Crystallization can be induced by cooling a saturated solution of the target compound, by adding an antisolvent (a liquid in which the target compound is poorly soluble) to a solution of the compound (Mostafa et al., Chemical Engineering Science 63:5457-5467, 2008), by introducing a surface that promotes crystal formation (e.g., scratched glass), by adding crystalline seeds, or by adding compounds that will co-crystallize with the target compound, including polymers (Edueng et al., Journal of Controlled Release 256:193-202, 2017) (Korotkova and Karatchvil, Procedia Chemistry 10:473-476, 2014). Industrial crystallization processes can be carried out in batch mode or, increasingly, via continuous processes (Zhang et al., Engineering 3:354-364, 2017).

[0192] Crystallization can be used in one of two ways: the desired disulfide compound can be selectively crystallized in the presence of impurities (the impurities remain in solution and can therefore be easily removed after crystallization), or one or more impurities can be crystallized while the desired compound remains in solution. The two approaches can be combined sequentially (one after the other).

[0193] Because cysteamine precursors are often administered in relatively large doses (more than 1 gram per day, or up to 10 grams per day in adults), any salt must be safe. Therefore, salts that are natural products, present at significant levels in the diet, and have pleasant organoleptic properties are preferred. Examples of such salts include, but are not limited to, acetate, citrate, and tartrate. The salt form of a cysteamine precursor that induces crystal formation may not overlap with a pharmaceutically preferred salt form. In such cases, a salt form may be specifically generated to aid crystallization, and then redissolved in a solvent to convert the salt into the preferred pharmaceutical salt. Larger salts that tend to promote crystal formation include benzoates and naphthoates (naphthoic acid).

[0194] Separation method Most of the synthetic routes outlined above are unable to produce mixed disulfides with purity greater than 99% (i.e., within the range required by drug regulatory agencies). Crystallization may also not be selective enough to reduce drug impurities to acceptable levels. Therefore, robust separation methods may be required. Therefore, in certain embodiments, any synthetic method can be combined with an efficient scheme for separating the desired product (i.e., the mixed disulfide cystamine precursor) from other compounds (including other disulfides) produced in the synthesis. Useful separation methods other than crystallization include various chromatographic procedures, including resins that separate small molecules based on size, charge, hydrophobicity, affinity, or other properties.

[0195] formulation When used as a pharmaceutical, cysteamine precursors, or their pharmaceutically acceptable salts, solvates, or prodrugs, can be administered in the form of pharmaceutical compositions. These compositions can be prepared by various methods well known in the pharmaceutical industry, and can be made to release the drug in a specific part of the gastrointestinal tract at a controlled time using various excipients and formulation techniques. For example, formulations can be tailored to address specific diseases, achieve the blood level of cysteamine required to achieve therapeutic efficacy, allow the desired duration of drug effect, and provide a set of compositions with various drug release characteristics that can be administered in different combinations to account for patient-to-patient variations in cysteamine metabolism. Administration is primarily via the oral route, and may be supplemented by suppositories. Cysteamine precursors can also be co-formulated with agents that promote the production or absorption of cysteamine in vivo, including, for example, reducing agents, buffers, pantetheinase inducers, or inducers of cysteamine uptake by intestinal epithelial cells.

[0196] Pharmaceutical compositions can contain one or more pharmaceutically acceptable carriers. In preparing pharmaceutical compositions for use in the methods of the present invention, cysteamine precursors, their pharmaceutically acceptable salts, solvates, or prodrugs are typically mixed with excipients, diluted with excipients, or enclosed in carriers in the form of, for example, capsules, tablets, sachets, paper, vials, or other containers. The active ingredients of the present invention can be administered alone or as a mixture in the presence of pharmaceutically acceptable excipients or carriers. The excipients or carriers are selected based on the mode and route of administration, the region of the gastrointestinal tract targeted for drug release, and the intended time profile of drug release. When the excipient serves as a diluent, it can be a solid, semi-solid, or liquid substance (e.g., saline) that acts as a vehicle, carrier, matrix, or other medium for the active ingredient. Therefore, the composition can be in the form of tablets, powders, granules, lozenges, sachets, cachets, elixirs, suspensions, emulsions, solutions, syrups, and soft and hard gelatin capsules.As known in the art, the type and amount of excipients vary depending on the intended drug release characteristics.The resulting composition can contain additional agents such as preservatives or coatings.

[0197] Suitable pharmaceutical carriers and pharmaceutical ingredients for use in pharmaceutical formulations are described in well-known references in the field, such as Remington: The Science and Practice of Pharmacy, 21st Ed., Gennaro, Ed., Lippencott Williams & Wilkins (2005), and USP / NF (United States Pharmacopoeia and National Pharmaceutical Standards), or corresponding European or Japanese references. Examples of suitable excipients are lactose, dextrose, sucrose, sorbitol, mannitol, starch, acacia gum, calcium carbonate, calcium phosphate, alginate, tragacanth, gelatin, calcium silicate, microcrystalline cellulose, cellulose derivatives, polyvinylpyrrolidone, poly(lactic-co-glycolic acid) (PLGA), cellulose, water, syrup, methylcellulose, vegetable oil, polyethylene glycol, hydrophobic inert matrix, carbomer, hypromellose, gelucire 43 / 01, docusate sodium, and white wax. The formulation may additionally include lubricants such as talc, magnesium stearate, and mineral oil; wetting agents; emulsifying and suspending agents; preservatives such as methyl and propylhydroxybenzoates; sweeteners; and flavoring agents. Other exemplary excipients and their use details are described in Handbook of Pharmaceutical Excipients, 6th Edition, Rowe et al., Eds., Pharmaceutical Press (2009).

[0198] The pharmaceutical composition can include a cysteamine precursor salt, optionally co-formulated or co-administered with other agents that promote the in vivo breakdown of cysteamine precursors to cysteamine or promote the intestinal absorption of cysteamine. The pharmaceutical composition can also include other therapeutic agents that complement the pharmacological effects of cysteamine in the target disease. Exemplary enhancers of in vivo cysteamine production or absorption, and exemplary therapeutic agents that can be included in the compositions described herein, are provided herein.

[0199] The compositions of the present invention may contain a single active ingredient (i.e., a single cysteamine precursor), or a combination of a first and a second active ingredient in a single unit dosage form, or a combination of a first, a second, a third, and optionally a fourth active ingredient, and optionally a fifth ingredient, in a single unit dosage form. In compositions with two active ingredients, both ingredients may be cysteamine precursors, or one ingredient may be a promoter of in vivo cysteamine production (e.g., a reducing agent that promotes the reduction of disulfide cysteamine precursors or an agent that induces increased intestinal expression of pantetheinase), or a promoter of intestinal absorption of cysteamine (e.g., an agent that induces increased expression of one or more organic cation transporters, such as OCT1, OCT2, or OCT3). In compositions with three or four active ingredients, all ingredients may be cysteamine precursors, or one or two ingredients may be a promoter of in vivo cysteamine production and / or intestinal absorption. In the composition having two or more cysteamine precursors, the type of cysteamine precursor is selected to achieve the production of cysteamine in vivo over a sustained period.For example, the mixed disulfide cysteamine precursor, which only requires the reduction of disulfide bond to produce one cysteamine, will therefore begin to produce cysteamine immediately after reaching the area of ​​the gastrointestinal tract that has an oxidation-reduction environment that is conducive to the reduction of disulfide bond, can be mixed with pantetheine or pantetheine disulfide, which requires both the reduction of disulfide bond and pantetheinase cleavage to produce cysteamine, and can optionally be combined with a compound that can be degraded to pantetheine in the intestine, or a disulfide containing such a compound that requires an additional step to produce pantetheine and thus cysteamine.Compounds that can be degraded to pantetheine in the intestine include 4-phosphopantetheine, dephosphocoenzyme A, coenzyme A, and suitable analogs and derivatives.The time course of cysteamine production in vivo varies depending on the number of decomposition steps between cysteamine precursor and cysteamine.In some embodiments, the composition comprising multiple cysteamine precursors is formulated as a powder, granules, or liquid, i.e., a formulation type that can accommodate large amounts of drug.

[0200] Pharmaceutical compositions can also include one or more agents that enhance the performance of the formulation. For example, a gastroretentive composition can include a compound that slows gastric emptying to extend the residence of the composition in the stomach.

[0201] In a composition containing two cysteamine precursor components, the first and second components may be present in a ratio of, for example, about 1:1.5 to about 1:4. In a composition containing three cysteamine precursor components, the first, second, and third components may be present in a ratio of, for example, about 1:1:2 to about 1:4:4. In a composition containing four active ingredients, the first through fourth active ingredients may be present in a ratio of, for example, about 1:1:1:2 to about 1:2:5:5. In a composition containing five active ingredients, the first through fifth active ingredients may be present in a ratio of, for example, about 1:1:2:2:2 to about 1:1:2:5:5:8.

[0202] In some embodiments, compositions containing two or more cysteamine precursors include one precursor selected for rapid in vivo cysteamine production (e.g., requiring simple disulfide bond reduction) and a second precursor selected for more moderate or slower in vivo conversion to cysteamine (e.g., requiring chemical reduction and at least one enzymatic degradation step). In some embodiments, pharmaceutical compositions containing two or more cysteamine precursors include those in which at least one precursor is a cysteamine mixed disulfide, which can yield cysteamine upon disulfide bond reduction. In further related embodiments, at least one additional component is a disulfide-containing pantetheine or a compound degradable to pantetheine in the gastrointestinal tract.

[0203] The composition can be formulated in a solid unit dosage form (e.g., tablet or capsule), with each dosage containing, for example, 50 to 800 mg of the active ingredient of the first component. For example, dosages can be about 50 mg to about 800 mg, about 50 mg to about 700 mg, about 50 mg to about 600 mg, about 50 mg to about 500 mg, about 75 mg to about 800 mg, about 75 mg to about 700 mg, about 75 mg to about 600 mg, about 75 mg to about 500 mg, about 100 mg to about 800 mg, about 100 mg to about 700 mg, about 100 mg to about 600 mg, It may contain about 100 mg to about 500 mg, about 250 mg to about 800 mg, about 250 mg to about 700 mg, about 250 mg to about 600 mg, about 250 mg to about 500 mg, about 400 mg to about 800 mg, about 400 mg to about 700 mg, about 400 mg to about 600 mg, about 450 mg to about 700 mg, or about 450 mg to about 600 mg of the active ingredient of the first component.

[0204] In alternative embodiments, the compositions can be formulated in liquid or powder unit dosage form, with each dosage unit containing about 250 mg to about 10,000 mg of cysteamine precursor. For example, dosages can range from about 250 mg to about 10,000 mg, about 250 mg to about 8,000 mg, about 250 mg to about 6,000 mg, about 250 mg to about 5,000 mg, about 500 mg to about 10,000 mg, about 500 mg to about 8,000 mg, about 500 mg to about 6,000 mg, about 500 mg to about 5,000 mg, about 750 mg to about 10,000 mg, about 750 mg to about 8,000 mg, about 750 mg to about 6,000 mg, about 750 mg to about 6,000 mg, about 750 mg to about 5,000 mg, about 750 mg to about 10,000 mg, about 750 mg to about 8,000 mg, about 750 mg to about 6,000 mg, about 750 mg to about 5 ... The active ingredient of the first component may be from about 5,000 mg, from about 1,250 mg to about 10,000 mg, from about 1,250 mg to about 8,000 mg, from about 1,250 mg to about 6,000 mg, from about 1,250 mg to about 5,000 mg, from about 2,000 mg to about 10,000 mg, from about 2,000 mg to about 8,000 mg, from about 2,000 mg to about 6,000 mg, from about 2,000 mg to about 5,000 mg, or from about 3,000 mg to about 6,000 mg.

[0205] In compositions having a first and second cysteamine precursor component, the amount of the second active component in the solid unit dosage form can vary, for example, from 50 to 700 mg. For example, dosages may include about 50 mg to about 700 mg, about 50 mg to about 600 mg, about 50 mg to about 500 mg, about 50 mg to about 450 mg, about 75 mg to about 700 mg, about 75 mg to about 600 mg, about 100 mg to about 700 mg; about 100 mg to about 600 mg, about 100 mg to about 500 mg, about 100 mg to about 400 mg, about 250 mg to about 700 mg, about 250 mg to about 600 mg, about 250 mg to about 500 mg, about 250 mg to about 400 mg, about 400 mg to about 700 mg, about 400 mg to about 600 mg, about 400 mg to about 500 mg, about 450 mg to about 700 mg, about 450 mg to about 600 mg, and about 450 mg to about 500 mg. In a composition having a cysteamine precursor as a first active ingredient and an enhancer of in vivo cysteamine production as a second active ingredient, the amount of the second active ingredient in a unit dosage form can vary, for example, from 0.1 mg to 400 mg.

[0206] In alternative embodiments comprising first and second cyseamine precursor components, the amount of the second active ingredient in a liquid or powder unit dosage form can vary, for example, from about 250 mg to about 6,000 mg. For example, dosages can range from about 250 mg to about 6,000 mg, about 250 mg to about 5,000 mg, about 250 mg to about 4,000 mg, about 250 mg to about 3,000 mg, about 250 mg to about 2,000 mg, about 500 mg to about 6,000 mg, about 500 mg to about 5,000 mg, about 500 mg to about 4,000 mg, about 500 mg to about 3,000 mg, about 750 mg to about 6,000 mg, about 750 mg to about 5,000 mg, or about 750 mg to about 4 The active ingredient of the second ingredient may be about 2,000 mg, about 750 mg to about 3,000 mg, about 1,250 mg to about 6,000 mg, about 1,250 mg to about 5,000 mg, about 1,250 mg to about 4,000 mg, about 1,250 mg to about 3,000 mg, about 2,000 mg to about 6,000 mg, about 2,000 mg to about 5,000 mg, about 2,000 mg to about 4,000 mg, about 2,000 mg to about 3,000 mg, or about 2,500 mg to about 5,000 mg.

[0207] In solid compositions containing a third, or third and fourth, cysteamine precursor component, a unit dosage can contain about 50 mg to about 400 mg of each of the third active ingredient and, if present, the fourth active ingredient. For example, dosages can contain about 50 mg to about 400 mg, about 50 mg to about 350 mg, about 50 mg to about 300 mg, about 50 mg to about 250 mg, about 75 mg to about 400 mg, about 75 mg to about 350 mg, about 75 mg to about 300 mg, about 75 mg to about 250 mg, about 100 mg to about 400 mg, about 100 mg to about 350 mg, about 100 mg to about 300 mg, about 100 mg to about 250 mg, about 250 mg to about 400 mg, about 250 mg to about 350 mg, or about 250 mg to about 300 mg. In a composition having five active ingredients, the unit dosage of the five ingredients can range from about 50 mg to about 300 mg. In a composition having a fourth active ingredient and, optionally, a promoter of in vivo cysteamine production as a third active ingredient, the amount of the fourth active ingredient and, optionally, the third active ingredient in a unit dosage form can vary, for example, from 0.1 mg to 400 mg.

[0208] In alternative embodiments comprising a third, or third and fourth cysteamine precursor components in a liquid or powder unit dosage form, the unit dosage of the third and optionally fourth active ingredients may vary, for example, from about 250 mg to about 4,000 mg. For example, dosage amounts may range from about 250 mg to about 4,000 mg, from about 250 mg to about 3,000 mg, from about 250 mg to about 2,000 mg, from about 250 mg to about 1,000 mg, from about 500 mg to about 4,000 mg, from about 500 mg to about 3,000 mg, from about 500 mg to about 2,000 mg, from about 500 mg to about 1,000 mg, from about 750 mg to about 4,000 mg, from about 750 mg to about 3,000 mg, from about 750 mg to about 2,000 mg, or from about 750 mg per dose. and optionally, a third and optionally a fourth active ingredient may be contained in an amount of from about 1,000 mg, about 1,000 mg to about 4,000 mg, about 1,000 mg to about 3,000 mg, about 1,000 mg to about 2,000 mg, about 1,000 mg to about 1,500 mg, about 1,500 mg to about 4,000 mg, about 1,500 mg to about 3,000 mg, about 1,500 mg to about 2,000 mg, about 2,000 mg to about 4,000 mg, or about 2,000 mg to about 3,000 mg.

[0209] Pharmaceutical compositions can be formulated to provide immediate, delayed, gastroretentive, sustained, or colonic release (collectively referred to as controlled release) of the active ingredient after administration to a patient by employing procedures known in the art.

[0210] To prepare solid compositions such as tablets, active ingredient(s) (for example, some cysteamine precursors) can be mixed with one or more pharmaceutical excipients to form a solid bulk formulation composition containing a homogeneous mixture of the compounds of the present invention.When these bulk formulation compositions are homogeneous, the active ingredient is typically evenly dispersed throughout the composition, allowing the composition to be easily subdivided into equally effective unit dosage forms such as tablets, capsules, or microparticles.The solid bulk formulation is then subdivided into the above-mentioned types of unit dosage forms.

[0211] Alternatively, two homogenous batches of active ingredient(s) mixed with one or more pharmaceutical excipients can be prepared, each using a different concentration of the active ingredient(s). The first mixture can then be used to form a core, and the second mixture can form a shell around the core to form a composition with variable drug release characteristics. If the higher-concentration batch is located in the core and the lower-concentration batch is located in the shell, once the shell has substantially dissolved or eroded, an initial moderate rate of drug release is followed by a faster rate of drug release. In some embodiments, the pharmaceutical composition contains a higher concentration of active ingredient(s) in the core than in the shell. The ratio of cysteamine precursor concentration in the core:shell can range, for example, from about 1.5:1 to 4:1. The type or concentration of excipients can also differ between the two batches to affect the drug release rate. In some embodiments, the polymer(s) or other matrix-forming components in the core release the active ingredient(s) more slowly than from the shell. In such embodiments, the higher concentration of cysteamine precursor(s) in the core is partially or completely balanced by a slower rate of drug release, extending the duration of cysteamine precursor release and thus the duration of cysteamine production in vivo, intestinal absorption, and elevated blood levels. One or more coatings may be applied to the core before the shell layer is applied, and additional coatings can be applied to the shell to allow for an efficient manufacturing process and / or to help provide desired pharmacological properties, including the timing and location of drug release within the gastrointestinal tract.

[0212] The pharmaceutical compositions of the present invention include those formulated to release mixtures of cysteamine precursors that differ in the number of degradation steps or mechanism(s) that lead to cysteamine production. Specifically, mixtures of two, three, four, or five cysteamine precursors, each separated by one, two, three, or more chemical and / or enzymatic degradation steps from releasing cysteamine, can be used. For example, one step can be disulfide bond reduction (in the case of cysteamine mixed disulfide) or pantetheinase cleavage (in the case of pantetheine). Two steps can be disulfide bond reduction followed by pantetheinase cleavage (in the case of pantetheine disulfide), or phosphatase cleavage followed by pantetheinase cleavage (in the case of 4-phosphopantetheine). Three steps can be disulfide bond reduction followed by pantetheinase cleavage (e.g., by phosphatase) before or after degradation to pantetheine, followed by pantetheinase cleavage (e.g., in the case of 4-phosphopantetheine disulfide). The four steps can be disulfide bond reduction, followed by two degradation steps to pantetheine (e.g., removal of the adenine nucleotide moiety by ectonucleotide diphosphatase, followed by removal of the 4' phosphate by a phosphatase), followed by pantetheinase cleavage (e.g., coenzyme A or dephospho-coenzyme A disulfide). The purpose of combining cysteamine precursors with different chemical and / or enzymatic degradation pathways to cysteamine is to extend the time during which cysteamine is produced and absorbed from the intestine, thereby extending the duration of therapeutically effective cysteamine blood levels. In some embodiments, the pharmaceutical compositions of the present invention comprise at least two cysteamine precursors, and in further embodiments, the pharmaceutical compositions contain three cysteamine precursors.

[0213] The pharmaceutical compositions of the present invention can be formulated for mixed release, meaning that one composition contains two drug release profiles. For example, an immediate-release formulation can be combined with a sustained-release formulation. (See, e.g., Compound F in Figure 14.) In such compositions, the first active ingredient can be formulated for immediate release, starting between about 5 minutes and about 30 minutes after ingestion. For example, the first active ingredient can be released 5, 10, 15, 20, 25, 30, or 45 minutes after ingestion of the composition. The first active ingredient is formulated so that cysteamine plasma concentrations within the therapeutic range are achieved between about 15 minutes and 3 hours, preferably 30 minutes and 2 hours, after ingestion. For example, therapeutic plasma cysteamine concentrations can be reached 0.5 hours, 1 hour, 2 hours, or 3 hours after ingestion of the composition. The type of cysteamine precursor used (e.g., thiol, cysteamine mixed disulfide, pantothecin disulfide, coenzyme A disulfide, N-acetylcysteamine disulfide, etc.) affects the length of time it takes to reach therapeutic blood levels of cysteamine and the duration for which therapeutic blood levels are maintained.

[0214] In compositions having two, three, and optionally four or five active ingredients (e.g., multiple cysteamine precursors and / or enhancers of in vivo cysteamine production and absorption), each of the second, third, and / or fourth, and / or fifth active ingredients is formulated to initiate controlled release from the composition between about 1 hour and about 8 hours after ingestion. Controlled-release compositions may include delayed-release and / or sustained-release formulations. For example, the second, third, and / or fourth active ingredients may be released from the composition between 1 hour, 1.5 hours, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 7 hours, or 8 hours after ingestion. The second, third, and / or fourth active ingredients are formulated so that plasma concentrations of cysteamine (reflecting the contributions of all active ingredients) are maintained within the therapeutic range beginning between about 30 minutes and 2 hours after ingestion and extending for 6 to 10 hours, more preferably 8 to 12 hours, or longer. For example, plasma cysteamine concentrations may be sustained within the therapeutic range for 6, 8, 10, 12, 15, 20, or 24 hours after ingestion of the active ingredients of the composition. Depending on the age and size of the patient, the disease being treated, and the patient's rate of cysteamine metabolism, two or more compositions may be required to deliver sufficient cysteamine precursors to achieve therapeutic blood levels over several hours.

[0215] As an alternative or complement to pharmaceutical compositions containing mixed formulations, in some embodiments, compositions consisting of a single formulation can be produced. That is, time-based formulations, such as immediate-release or sustained-release formulations, and anatomically targeted formulations, such as gastroretentive, delayed-release, and colon-targeted formulations, can be prepared for administration as separate compositions. Formulating a collection of pharmaceutical compositions with different drug release characteristics (whether time-based or anatomically / physiologically based) has certain advantages. For example, such compositions can be administered to different patients in different combinations and ratios to achieve blood cysteamine levels within the therapeutic range over time. That is, a treatment regimen consisting of one, two, three, or more compositions administered on a specific schedule can be tailored to an individual patient's cysteamine production, absorption, and metabolic capacities. Because these capacities are known to vary between patients, formulating multiple homogeneous compositions containing different cysteamine precursors and different drug release characteristics can be combined in different ratios for different patients, addressing the known limitations of existing cysteamine formulations.

[0216] Preferably, the combination of two or more pharmaceutical compositions is capable of maintaining cysteamine blood levels within the therapeutic range for at least 2-8 hours after ingestion, more preferably 1-8 hours, even more preferably 2-10 hours, and most preferably 1-10 hours, 1-12 hours, 1-14 hours, or more. Separately formulated pharmaceutical compositions containing different cysteamine precursors with different drug release profiles provide the dosing flexibility needed to individualize dosing regimens to achieve therapeutically effective cysteamine blood levels over the long term.

[0217] It is well documented that gastric emptying times and colonic transit times vary significantly (up to two-fold or more) among healthy individuals. The intestinal redox environment and levels of pantetheinase activity are also known to vary among individuals. These and other factors likely explain the wide interindividual variability in plasma cysteamine levels observed after cysteamine administration. For example, in a study of the pharmacokinetics of immediate-release cysteamine bitartrate in healthy volunteers, peak cysteamine blood levels (Cmax) following a 600 mg oral dose administered with a meal varied more than eight-fold, from 7 micromoles to 57.3 micromoles (Dohil R. and P. Rioux, Clinical Pharmacology in Drug Development 2:178 (2013)). In the same study, Cmax following 600 mg of delayed-release cysteamine bitartrate administered with a meal varied 12-fold, from 2.1 micromoles to 25.4 micromoles (Dohil R. and P. Rioux, Clinical Pharmacology in Drug Development 2:178 (2013)). Interpatient variability in cysteamine plasma levels was less extreme when cysteamine was administered to fasting patients, but still varied by up to four-fold. (When cysteamine is administered every 6 hours, as with Cystagon®, or every 12 hours, as with Procysbi®, it is difficult to completely avoid mealtimes).

[0218] Current cysteamine formulations and administration methods offer only one tool for addressing inter-subject variability: increasing or decreasing the dose. The cysteamine precursors, in vivo cysteamine production and absorption enhancers, drug formulations, and drug administration methods of the present invention provide multiple tools for achieving therapeutic blood cysteamine levels by tailoring the compound, dosage form, and administration regimen to the individual patient without incurring the unacceptable toxicity often associated with a high Cmax, or inadequate therapeutic effects associated with blood levels below the therapeutic threshold for long periods of time.

[0219] Another advantage of separately formulated compositions is that they can be administered at different times relative to meals.This is a useful option because different classes of cysteamine precursors and different types of preparations interact differently with meals.For example, in order to maximize gastric retention time, gastroretentive preparations should be administered with meals or immediately after meals, preferably with nutritious meals.On the other hand, immediate release preparations containing cysteamine mixed disulfides, which can be quickly converted into cysteamine by reducing disulfide bonds, should preferably not be administered with heavy meals.Although heavy meals hinder the absorption of cysteamine in some individuals, meals are compatible with certain cysteamine precursors, such as pantetheine disulfide, which produce little (if any) cysteamine in the stomach, and tend to be converted into cysteamine in the small intestine.

[0220] The personalized dosing regimen possible with the compounds and formulations of the present invention is particularly useful because, although the wide inter-individual variability in the intestinal absorption of cysteamine has been well documented, the relatively moderate intra-individual variability is also well documented.That is, a given subject will absorb and metabolize a dose of cysteamine in substantially the same way when administered on multiple occasions under similar circumstances.Therefore, a personalized dosing regimen that produces blood cysteamine levels within the therapeutic range for a particular patient should produce relatively stable and predictable results over time.

[0221] Sustained-release formulations can be designed to release drugs over widely varying periods of time using methods known in the art. (Wen, H. and Park, K., eds.: Oral Controlled Release Formulation Design and Drug Delivery: Theory to Practice, Wiley, 2010; Wells, JI and Rubinstein, MH, eds.: Pharmaceutical Technology: Controlled Drug Release, Vol. I and Vol. II, Ellis and Horwood, 1991; and Gibson, M., ed.: Pharmaceutical Preformulation and Formulation: A Practical Guide from Candidate Drug Selection to Commercial Dosage Form, 2nd ed., Informa, 2009.)

[0222] 14, 15, and 16 provide examples of pharmaceutical compositions of the present invention and are intended to illustrate aspects such as active ingredients (cysteamine precursors, enhancers of conversion of cysteamine precursors to cysteamine, and enhancers of intestinal absorption of cysteamine), dosage ranges (when all active ingredients are combined), formulation types (including mixed formulations), composition combinations, and methods of administration (e.g., with food or with a meal). The active ingredients include cysteamine precursors, as well as enhancers of in vivo cysteamine production and enhancers of intestinal absorption of cysteamine.

[0223] Formulations for oral administration Pharmaceutical compositions contemplated by the present invention include those formulated for oral administration ("oral dosage forms"). Oral dosage forms may be in the form of, for example, tablets, capsules, liquid solutions or suspensions, powders, or liquid or solid crystals or granules, which contain the active ingredient(s) in a mixture with non-toxic pharmaceutically acceptable excipients. When formulated as a liquid, powder, crystals, or granules, the dosage may be packaged in a manner that clearly defines the unit dose. For example, powders or granules or particulates may be packaged in sachets. Liquids may be packaged in glass or plastic containers.

[0224] Excipients are selected to provide acceptable organoleptic properties, control drug release characteristics, facilitate efficient manufacturing, and ensure long-term stability of the pharmaceutical composition, among other considerations known to those skilled in the art of pharmacology, pharmaceuticals, and pharmaceutical manufacturing. Excipients may be, for example, inert diluents or fillers (e.g., sucrose, sorbitol, sugars, mannitol, microcrystalline cellulose, starches including potato starch, calcium carbonate, sodium chloride, lactose, calcium phosphate, calcium sulfate, or sodium phosphate); granulating and disintegrating agents (e.g., cellulose derivatives including microcrystalline cellulose, starches including potato starch, croscarmellose sodium, alginates, or alginic acid); binders (e.g., sucrose, glucose, sorbitol, acacia, alginic acid, sodium alginate, gelatin, starch, pregelatinized starch, microcrystalline cellulose, magnesium aluminum silicate, sodium carboxymethylcellulose, hydroxypropylmethylcellulose, ethylcellulose, polyvinylpyrrolidone, or polyethylene glycol); and lubricants, glidants, and anti-adherents (e.g., magnesium stearate, zinc stearate, stearic acid, silica, hydrogenated vegetable oil, or talc). Other pharmaceutically acceptable excipients may be colorants, flavoring agents, plasticizers, humectants, preservatives, buffers, stabilizers, etc. Many of these excipients are sold by multiple excipient manufacturers in various chemical forms and / or can be used at different concentrations and / or in various combinations with other excipients to ensure differences in performance characteristics. Certain excipients may fulfill multiple purposes in a formulation.

[0225] The formulation for oral administration may also be presented as chewable tablets, hard gelatin capsules in which the active ingredient is mixed with an inert solid diluent (e.g., potato starch, lactose, microcrystalline cellulose, calcium carbonate, calcium phosphate, or kaolin), or soft gelatin capsules in which the active ingredient is mixed with water or an oil medium, such as peanut oil, liquid paraffin, or olive oil. Powders, granules, and pellets may be prepared using the ingredients described above under tablets and capsules in a conventional manner, for example, using a mixer, a fluidized bed device, or a spray-drying device.

[0226] One category of useful formulations primarily controls the rate of drug release (e.g., immediate- and sustained-release formulations), although the location of drug release is important. A second category of useful formulations primarily controls the anatomical site of drug release (e.g., gastroretentive formulations for drug release in the stomach and colon-targeted formulations in the large intestine), although the timing of release is important. Enteric-coated formulations, designed to remain intact in the acidic gastric environment and often designed to dissolve in the more alkaline small intestine (a type of anatomical target), are often referred to as delayed-release formulations, emphasizing the time-control element. However, colon-targeted formulations may also have an enteric coating to prevent dissolution in the stomach, highlighting the complex relationship between anatomical targeting and control of drug release rate. Furthermore, there is extensive overlap between the excipients used in time-based and anatomically or physiologically targeted formulations. These types of formulations can be combined in various ways to create multiple compositions with distinct drug release profiles in both time and space. Such compositions, in turn, can be combined in different amounts and ratios to individualize treatment regimens to accommodate biochemical and physiological variations among patients, as well as variations in disease type, severity, and activity.

[0227] Gastric retention preparations A gastroretentive formulation may be used to release a cysteamine precursor or its salt from the composition of the present invention in the stomach and to control the release of the active ingredient(s) of the composition in the stomach over an extended period of time. In other words, since the point of a gastroretentive formulation is long-term gastric retention, the accompanying excipients should provide sustained release of the active ingredient throughout the expected period that the gastroretentive dosage form is present in the stomach, and optionally for a longer period, including the time it takes to pass through the small intestine and into the colon. Gastric retention of the active ingredient of the present invention may be achieved by various mechanisms, such as mucoadhesion, flotation, sedimentation, swelling, and distension, and / or by co-administration of a pharmacological agent that delays gastric emptying. The excipients used in the gastroretentive formulation, as well as the size and shape of the pharmaceutical composition, vary depending on the mechanism of gastric retention.

[0228] Mucoadhesive / bioadhesive gastroretentive formulations Mucoadhesion refers to the attachment of a polymer utilized in a formulation to the gastrointestinal mucus layer until it is naturally removed from the surface as a result of ongoing mucus production. Bioadhesion, sometimes used interchangeably with mucoadhesion, also encompasses the attachment of a polymer or other component of a pharmaceutical composition to molecules on the surface of gastrointestinal epithelial cells. The purpose of mucoadhesion and bioadhesion is to increase the time that a pharmaceutical composition is in close proximity to gastrointestinal epithelial cells, including cell types capable of cysteamine precursor cleavage (i.e., cells expressing pantetheinase on their surface) and cysteamine uptake and transport into the circulation (e.g., cells expressing organic cation transporters). Mucoadhesive polymers can be used in formulating large dosage forms such as tablets or capsules, as well as small dosage forms such as microparticles or microspheres. Various physiological factors, such as peristalsis, mucin type, mucin turnover rate, gastrointestinal pH, fasting / fed state, and type of food in the fed state, affect the degree and persistence of mucoadhesion. The mechanism of mucoadhesion is thought to be due to the formation of electrostatic and hydrogen bonds at the interface between the polymer and the mucus. Generally, mucoadhesion is achieved by polymers that have affinity for the gastrointestinal mucosa, and are selected from synthetic or natural bioadhesive substances, such as polyacrylic acid, methacrylic acid and its derivatives or both, polybrene, polylysine, polycarbophil, carbomer, alginate, chitosan, cholestyramine, gum, lectin, polyethylene oxide, sucralfate, tragacanth, dextrin (e.g., hydroxypropyl beta-cyclodextrin), polyethylene glycol (PEG), gliadin, cellulose and cellulose derivatives, such as hydroxypropylmethylcellulose (HPMC), or mixtures thereof.For example, cross-linked acrylic and methacrylic acid copolymers available under the trade names CARBOPOL (e.g., Carbopol 974P and 971P) and POLYCARBOPHIL are used in mucoadhesive formulations. (Hombach J. and A. Bernkop-Schnurch. Handbook of Experimental Pharmacology 197:251 (2010)).Other bioadhesive cationic polymers include acidic gelatin, polygalactosamine, poly-amino acids such as polylysine, polyornithine, polyquaternaries, prolamines, polyimines, diethylaminoethyldextran (DEAE), DEAE-imine, polyvinylpyridine, polythiodiethylaminomethylethylene (PTDAE), polyhistidine, DEAE-methacrylate, DEAE-acrylamide, poly-p-aminostyrene, polyoxetane, Eudragit RL, Eudragit RS, GAFQUAT, polyamidoamine, cationic starch, DEAE-dextran, DEAE-cellulose, and copolymethacrylates (including copolymers of HPMA), N-(2-hydroxypropyl)-methacrylamide (see, e.g., U.S. Pat. No. 6,207,197).

[0229] Mucoadhesion is most effective when applied to small particles (e.g., microparticles). Mucoadhesive formulations can be combined with one or more other gastroretentive formulation methods, including floating formulations, swelling / expanding formulations, or any type of sustained release formulation.

[0230] Floating gastroretentive formulation Floating as a gastric retention mechanism is effective in formulating active ingredients (e.g., cysteamine precursors) with a bulk density lower than that of gastric juice and / or chyme (partially digested food in the stomach) to maintain buoyancy in the stomach. Generally, a density of less than 1 gram per cubic centimeter is desirable, and more preferably a density of less than 0.9 grams per cubic centimeter. Buoyancy can be achieved by (i) using low-density substances, including lipids; (ii) preforming air bubble(s) in the center of the composition; or (iii) using effervescent excipients to generate bubbles in vivo. The latter type of pharmaceutical composition must be designed so that the gas generated by the effervescent excipient remains in the composition and thereby contributes to its buoyancy. For example, the effervescent excipient can be embedded in a polymer matrix to trap bubbles in the composition. The latter type of buoyant formulations are generally prepared using a swellable polymer or polysaccharide and an effervescent couple (e.g., sodium bicarbonate and citric or tartaric acid), or a matrix containing a chamber of trapped air or a liquid that generates gas upon contact with liquid stomach contents at body temperature. Floating gastroretentive formulations have been extensively reviewed (e.g., Kotreka, UK Critical Reviews in Therapeutic Drug Carrier Systems, 28:47 (2011)).

[0231] Floating pharmaceutical compositions designed for gastric retention have been known in the art for some time. For example, U.S. Patent Nos. 4,126,672, 4,140,755, and 4,167,558, each incorporated herein by reference, describe a "hydrodynamically balanced" drug delivery system (HBS) in tablet form with a density lower than that of gastric fluid (i.e., less than 1 gram per cubic centimeter). As a result, the composition floats on the gastric fluid or chyme, thereby avoiding release through the pylorus during gastric muscle contractions. The drug is continuously released from a cellulose-derived hydrocolloid, such as methylcellulose, hydroxyalkylcellulose (e.g., hydroxypropylcellulose, hydroxypropylmethylcellulose, hydroxyethylcellulose), or sodium carboxymethylcellulose, which, upon contact with gastric fluid, forms a water-impermeable barrier on the surface of the composition, which gradually erodes and slowly releases the drug. A bilayer floating tablet having an outer layer formulated for immediate release and an inner layer formulated for sustained release is also disclosed in U.S. Pat. No. 4,140,755, which is incorporated herein by reference.

[0232] Similar hydrodynamically balanced floating formulations for sustained delivery of L-dopa and decarboxylase inhibitors have also been described (see U.S. Pat. No. 4,424,235). Hydrocolloids such as gum arabic, gum tragacanth, locust bean gum, guar gum, karaya gum, agar, pectin, carrageen, soluble and insoluble alginates, carboxypolymethylene, gelatin, casein, zein, and bentonite may be useful in preparing the floating formulations of the present invention. The floating formulations may contain up to about 60% of a fatty substance or mixture of fatty substances selected from beeswax, cetyl alcohol, stearyl alcohol, glyceryl monostearate, hydrogenated castor oil, and hydrogenated cottonseed oil (oils have a lower density than gastric fluids). The floating formulations may promote sustained release of cysteamine precursors, providing elevated plasma cysteamine levels over a longer period of time. Long-term elevated plasma cysteamine levels allow for less frequent administration.

[0233] The floating composition of the present invention may contain a gas generating agent. Methods for formulating floating compositions using gas generating compounds are known in the art. For example, floating minicapsules containing sodium bicarbonate are described in U.S. Patent No. 4,106,120. Similar floating granules based on gas generation are described in U.S. Patent No. 4,844,905. Floating capsules are described in U.S. Patent No. 5,198,229.

[0234] The buoyant composition may optionally contain an acid source and a gas-generating carbonate or bicarbonate agent, which together act as an effervescent couple, producing carbon dioxide gas that provides buoyancy to the formulation. The effervescent couple, consisting of a soluble organic acid and an alkali metal carbonate, produces carbon dioxide when the mixture comes into contact with water, or when the alkaline component comes into contact with an acidic liquid (e.g., gastric juice). Typical examples of acids that can be used include citric acid, tartaric acid, malic acid, fumaric acid, or adipic acid. Typical examples of gas-generating alkalis that can be used include sodium bicarbonate, sodium carbonate, sodium glycine carbonate, sodium sesquicarbonate, potassium carbonate, potassium bicarbonate, calcium carbonate, ammonium bicarbonate, sodium bisulfite, sodium metabisulfite, etc. The gas-generating agent interacts with the acid source, triggered by contact with water or hydrochloric acid in gastric juice, to produce carbon dioxide or sulfur dioxide, which is trapped within the matrix of the composition and improves its flotation properties. In one embodiment, the gas-generating agent is sodium bicarbonate, and the acid source is citric acid.

[0235] Floating behavior is important because if a composition is not lighter than gastric fluid and / or chyme immediately after reaching the stomach, it may be rapidly expelled through the pylorus. Some compositions, such as those containing preformed gas bubbles or low-density substances such as lipids, have a lower density than gastric fluid and chyme upon ingestion. For buoyant compositions (i.e., effervescent formulations) that must achieve a density lower than that of gastric fluid and / or chyme after reaching the stomach, a density of less than 1 gram per cubic centimeter is preferably achieved within 30 minutes, more preferably within 15 minutes, and most preferably within 10 minutes after contact with gastric fluid. The duration of floatation is also important and should be consistent with the duration of drug release. That is, if a composition is designed to release a drug for 6 hours or more, it must also be able to float for 6 hours. Preferably, a buoyant composition maintains a density of less than 1 for at least 5 hours, more preferably 7.5 hours, and even more preferably 10 hours or more.

[0236] Large doses of cysteamine precursor (e.g., 2-10 grams) may be necessary to effectively treat some cysteamine-sensitive diseases and / or achieve adequate blood levels in adult subjects. Because the amount of any active agent that can be included in a standard dosage form (e.g., tablet, capsule) is limited by a patient's ability to swallow large compositions, and because administering multiple tablets or capsules can be inconvenient or uncomfortable (impossible for patients with swallowing disorders), alternative dosage forms that do not limit the amount of active agent in a unit dosage form are useful. Powders, granules, and liquids are examples of dosage forms that are not size-restricted but can be delivered in unit dosage amounts in appropriate packaging (e.g., sachets or vials). In some embodiments of the present invention, the floating, gastroretentive compositions of the present invention are administered in liquid form. In further embodiments, the liquid composition includes an alginate salt. In other embodiments, the active pharmaceutical ingredient is delivered in the form of a powder or granules that can be sprinkled on food.

[0237] One type of liquid gastroretentive, floating drug delivery system utilizes alginate as an excipient. Alginate is a linear block polysaccharide copolymer made from beta-D-mannuronic acid and alpha-L-guluronic acid residues connected by 1,4-glycosidic bonds. It is used for a wide range of purposes in pharmaceutical compositions, including as a sustained-release polymer (see Murata et al., Eur J Pharm Biopharm 50:221 (2000)). Gaviscon is the brand name for a floating liquid alginate formulation containing an antacid. Because it has been used to treat gastroesophageal reflux for decades, the safety of chronic alginate ingestion is well established. Floating formulations of alginate with small molecule drugs have been described (see Katayama et al., Biol Pharm Bull. 22:55 (1999) and Itoh et al., Drug Dev Ind Pharm. 36:449 (2010)). Floating formulations that form a layer on the surface of gastric contents are sometimes referred to as raft-forming formulations. Raft-forming floating / gelling sustained-release compositions are described by Prajapati et al., J Control Release 168:151 (2013), and Nagarwal et al., Curr Drug Deliv. 5:282 (2008).

[0238] U.S. Patent No. 4,717,713, incorporated herein by reference, discloses a liquid (drinkable) formulation that forms a semi-solid gel-like matrix in the stomach upon contact with stomach contents, thereby resulting in controlled release of the drug from the gelatinous matrix. Gel-forming vehicles are disclosed, including xanthan gum, sodium alginate, complex coacervate pairs such as gelatin or other polymers and carrageenan, and thermally gelling methylcellulose, all or a subset of which can be combined in various ratios to affect the dissolution and / or diffusion rate of the suspended pharmaceutically active agent(s). Other excipients used include carbonate compounds such as calcium carbonate, which are effective both as gelation promoters and as gas-generating agents to suspend the gel. Xyloglucan and gellan gum can also be used as gelling agents or in combinations of gelling agents.

[0239] Liquid (drinkable) floating formulations can include particulates that can be provided as liquid suspensions (concentrates or ready to use) or as powders that can be added to liquids (e.g., water, juice, or other beverages). Floating gastroretentive compositions can also be delivered in the form of a powder that is sprinkled on or otherwise mixed with food.

[0240] The floating gastroretentive formulations can include mucoadhesive polymers or other mucoadhesive components (see U.S. Patent Nos. 6,207,197 and 8,778,396), and can utilize polymers such as polyethylene oxide, polyvinyl alcohol, sodium alginate, ethyl cellulose, poly(lactic)coglycolic acid (PLGA), polylactic acid, polymethacrylate, polycaprolactone, polyester, polyacrylic acid, and polyamide.

[0241] Swelling and expansion of gastroretentive compositions Swelling and distension are mechanisms of gastric retention in which, upon contact with gastric fluid, the composition swells to an extent that it prevents it from exiting the stomach through the pylorus. As a result, the composition remains in the stomach for an extended period of time, for example, until the surface of the composition erodes to a size smaller than the diameter of the pylorus, or until food is substantially emptied from the stomach, at which point strong muscle contractions (sometimes called "housekeeper waves") sweep through the stomach and remove its contents. In the swollen or expanded state, the composition exceeds a diameter of approximately 14-16 mm, thereby excluding it from passing through the pyloric sphincter. Preferably, the composition exceeds a diameter of 16-18 mm. Swelling may be combined with buoyancy, which keeps the formulation away from the pylorus, especially in the fed state.

[0242] The concept of a drug product that swells upon contact with gastric juice and consequently remains in the stomach has been known since the 1960s. U.S. Patent No. 3,574,820 discloses a tablet that swells upon contact with gastric juice to a size that prevents it from passing through the pylorus and thus causes it to be retained in the stomach. Similarly, U.S. Patent No. 5,007,790 describes a tablet or capsule made of a hydrophilic, water-swellable crosslinked polymer that rapidly swells to promote gastric retention while allowing slow dissolution of drug molecules mixed with the polymer.

[0243] U.S. Patent Publication No. 2003 / 0104053, incorporated herein by reference, discloses a unit dosage tablet for pharmaceutical delivery in which the active ingredient is dispersed in a solid unit matrix formed from a combination of poly(ethylene oxide) and hydroxypropyl methylcellulose. This combination is said to offer unique advantages in terms of release rate control and reproducibility, while allowing both tablet swelling, which results in gastric retention, and tablet disintegration, which removes the tablet from the gastrointestinal tract after drug release has occurred. U.S. Patent No. 6,340,475, incorporated herein by reference and assigned to DepoMed, highlights a unit oral dosage form developed by incorporating an active ingredient into a polymer matrix composed of a hydrophilic polymer that swells upon absorption of water to a size large enough to promote retention of the dosage form in the stomach during the fed mode. The polymer matrix is ​​formed of a polymer selected from the group consisting of poly(ethylene oxide), cellulose, cross-linked polyacrylic acid, xanthan gum, and alkyl-substituted celluloses such as hydroxymethyl-cellulose, hydroxyethyl-cellulose, hydroxypropyl-cellulose, hydroxypropylmethyl-cellulose, carboxymethyl-cellulose, and microcrystalline cellulose.

[0244] Furthermore, gum-based swellable gastroretentive systems have also been developed by DepoMed researchers. U.S. Patent No. 6,635,280, incorporated herein by reference, discloses a controlled-release oral dosage form for highly water-soluble drugs, comprising one or more polymers that form a solid polymer matrix upon absorption of water, swelling to a size large enough to promote retention of the dosage form in the stomach during the fed mode. The polymer matrix may be formed from a polymer selected from poly(ethylene oxide), cellulose, alkyl-substituted cellulose, cross-linked polyacrylic acid, and xanthan gum. U.S. Patent No. 6,488,962, incorporated herein by reference, discloses an optimal tablet shape that prevents the tablet from passing through the pylorus while remaining convenient for swallowing. Tablets are made using water-swellable polymers, including cellulose polymers and their derivatives, polysaccharides and their derivatives, polyalkylene oxides, polyethylene glycol, chitosan, poly(vinyl alcohol), xanthan gum, maleic anhydride copolymers, poly(vinylpyrrolidone), starch and starch-based polymers, maltodextrin, poly(2-ethyl-2-oxazoline), poly(ethyleneimine), polyurethane hydrogels, cross-linked polyacrylic acid and its derivatives, and copolymers of the polymers listed above, including block copolymers and graft polymers.

[0245] U.S. Patent No. 6,723,340, incorporated herein by reference, discloses an optimal polymer mixture for producing a swellable gastroretentive composition. The mixture provides optimal control of swelling and drug release parameters, as well as control of dissolution / erosion parameters, to ensure passage of the composition into the small intestine with substantially complete drug release. A preferred polymer mixture includes a combination of poly(ethylene oxide) and hydroxypropyl methylcellulose. Preferred molecular weight and viscosity ranges are provided for the polymer mixture.

[0246] The methods described in the above patent publications have been used to formulate four USFDA-approved swellable gastroretentive formulations that are described in multiple publications (e.g., reviewed in Berner et al., Expert Opin Drug Deliv. 3:541 (2006)).

[0247] U.S. Patent Publication No. 2008 / 0220060, incorporated herein by reference, discloses a gastroretentive formulation containing an active ingredient granulated with a mixture of a weak gelling agent, a strong gelling agent, and a gas-generating agent. The strong gelling agent is selected from the group consisting of methylcellulose, hydroxypropylmethylcellulose, hydroxypropylcellulose (excluding low-substituted hydroxypropylcellulose), hydroxyethylcellulose, ethylcellulose, sodium carboxymethylcellulose, xanthan gum, guar gum, carrageenan gum, locust bean gum, sodium alginate, agar-agar, gelatin, modified starch, copolymers of carboxyvinyl polymers, copolymers of acrylates, copolymers of oxyethylene and oxypropylene, and mixtures thereof. This patent also describes a manufacturing method. U.S. Patent No. 7,674,480 discloses a swellable gastroretentive formulation that uses a mixture containing a superdisintegrant, tannic acid, and one or more hydrogels to produce very rapid swelling. U.S. Patent Publication No. 2004 / 0219186, incorporated herein by reference, provides an expandable gastroretentive device comprising a gel formed from a polysaccharide based on xanthan gum, locust bean gum, or a combination thereof. U.S. Patent Publication No. 2006 / 0177497, incorporated herein by reference, discloses a gellan gum-based oral controlled release dosage form as a platform technology for gastroretentive administration. The dosage form further comprises a hydrophilic polymer such as guar gum, hydroxypropylmethylcellulose, carboxymethylcellulose sodium salt, or xanthan gum.

[0248] U.S. Patent No. 6,660,300 discloses a biphasic swellable gastroretentive formulation suitable for delivering water-soluble drugs, in which swelling and drug release are achieved by separate compartments of the composition, and the internal solid granular phase comprises the drug and one or more hydrophilic polymers, one or more hydrophobic polymers, and / or one or more hydrophobic materials (such as waxes, fatty alcohols, and / or fatty acid esters). The external solid continuous phase (in which the drug-containing internal phase granules are embedded) is formed using one or more hydrophobic polymers and / or one or more hydrophobic materials (such as waxes, fatty alcohols, and / or fatty acid esters). Tablets and capsules are disclosed.

[0249] Other excipients useful in swellable or expandable matrix formulations include (i) water-swellable polymer matrices, and (ii) the following: polyalkylene oxides, particularly poly(ethylene oxide), polyethylene glycol, and poly(ethylene oxide)-poly(propylene oxide) copolymers; cellulose polymers; acrylic and methacrylic acid polymers, copolymers and esters thereof, preferably formed from acrylic acid, methacrylic acid, methyl acrylate, ethyl acrylate, methyl methacrylate, ethyl methacrylate, and copolymers thereof, either with each other or with additional acrylate species such as aminoethyl acrylate; maleic anhydride copolymers; polymaleic acid; poly(acrylamides), such as polyacrylamide itself, poly(methacrylamide), poly(dimethylacrylamide), and poly(N-isopropyl-acrylamide); poly(olefinic alcohols), such as poly(vinyl alcohol), poly(N-vinyl lactams), such as poly(vinylpyrrolidone), poly(N-vinyl caprolactam), and the like. copolymers thereof; polyols such as glycerol, polyglycerol (especially highly branched polyglycerol), propylene glycol, and trimethylene glycol substituted with one or more polyalkylene oxides, for example, mono-, di-, and tri-polyoxyethylated glycerol, mono- and di-polyoxyethylated propylene glycol, and mono- and di-polyoxyethylated trimethylene glycol; polyoxyethylated sorbitol and polyoxyethylated glucose; polyoxazolines including poly(methyloxazoline) and poly(ethyloxazoline); polyvinylamine; polyvinyl acetates, including polyvinyl acetate itself and ethylene-vinyl acetate copolymers, polyvinyl acetate phthalate, and the like; polyimines such as polyethyleneimine; starch and starch-based polymers; polyurethane hydrogels; chitosan; polysaccharide gums; gelin; and hydrophilic polymers selected from shellac, shellac acetyl alcohol, and shellac n-butyl stearate.The gastroretentive formulations may also include any combination of buoyant formulations, mucoadhesive formulations, expandable matrix formulations, modified shape formulations, and / or magnetic formulations.

[0250] In some embodiments, the pharmaceutical compositions of the present invention are gastroretentive compositions that are retained in the stomach as a result of swelling to a size that inhibits passage through the pylorus, hi further embodiments, the gastroretentive composition retains in the stomach by both swelling and floating mechanisms.

[0251] Expandable, shape-changing gastroretentive preparation Pharmaceutical compositions that expand, decompress, or otherwise change size and / or shape upon contact with liquid gastric contents have also been described and are suitable delivery vehicles for the compounds and formulations of the present invention. Such compositions use a similar principle to swelling / expanding gastroretentive formulations in that they change the shape of the stomach to a size and / or geometry that makes it difficult for the stomach to pass through the pylorus. Methods and materials for making expanding, stretching, or other shape-changing gastroretentive compositions are known in the art. For example, U.S. Pat. No. 3,844,285 describes various such devices intended for veterinary use in ruminants, although the basic principles also apply to human gastroretentive formulations. U.S. Pat. No. 4,207,890 describes a controlled-release drug delivery system consisting of a "disintegrated, expandable, nonporous polymeric envelope containing an effective expansion amount of a swelling agent therein," which swells and expands upon contact with gastric fluid, resulting in retention in the stomach in an expanded state. The composition is administered inside the capsule in its disintegrated form. Expanding and shape-changing gastroretentive compositions have been reviewed (e.g., Klausner et al., Journal of Controlled Release 90:143 (2003)).

[0252] An exemplary unfolding gastroretentive technology, called the "accordion pill," has been developed by Intec Pharma (Jerusalem, Israel). Multilayer planar structures of various shapes (at least one layer containing a drug) are folded into an accordion or stepped shape and packaged in a capsule, as described in Kagan, L., Journal of Controlled Release 113:208 (2006). Further features of the accordion pill and related technology, including the pharmaceutical excipients preferably used in its construction, are disclosed in U.S. Pat. No. 6,685,962, incorporated herein by reference. Upon contact with gastric contents, the capsule dissolves, releasing the folded composition, which rapidly unfolds and subsequently remains in the stomach for up to 12 hours when administered with a normal meal.

[0253] Other gastroretentive technologies include superporous hydrogels and ion-exchange resin systems. Superporous hydrogels rapidly swell (within 1 minute of liquid contact) due to rapid water uptake through numerous interconnected pores. The compositions can swell to over 100 times their original size while retaining sufficient mechanical strength to withstand the forces of gastric contraction through co-formulation with hydrophilic polymers such as croscarmellose sodium (e.g., Ac-Di-Sol). Ion-exchange resin beads can be loaded with negatively charged drugs and suspended using a gas-generating agent (e.g., bicarbonate, which reacts with chloride ions in gastric juice to produce carbon dioxide gas). The beads are encapsulated in a semipermeable membrane that traps gas, resulting in long-term suspension of the beads.

[0254] The gastroretentive formulation may also comprise any combination of mucoadhesive, floating, raft-forming, swelling, unfolding / shape-changing, superporous hydrogel or ion-exchange resin formulations. Such combinations are known to those skilled in the art. For example, U.S. Patent No. 8,778,396 ("Multi-unit gastroretentive pharmaceutical dosage form comprising microparticles"), the entire contents of which are incorporated herein by reference, describes a composite mucoadhesive, floating gastroretentive formulation composed of microparticles.

[0255] The composition of the present invention can comprise, but is not limited to, the hydrophilic polymer with swelling and / or mucoadhesive properties to further promote gastric retention.The hydrophilic polymer with swelling and / or mucoadhesive properties suitable for being incorporated into the composition of the present invention includes, but is not limited to, polyalkylene oxide; cellulose polymer; acrylic acid and methacrylic acid polymer and their ester, maleic anhydride polymer; polymaleic acid; poly(acrylamide); poly(olefin alcohol); poly(N-vinyl lactam); polyol; polyoxyethylated sugar; polyoxazoline; polyvinylamine; polyvinyl acetate; polyimine; starch and starch-based polymer; polyurethane hydrogel; chitosan; polysaccharide gum; zein; shellac-based polymer; polyethylene oxide, hydroxypropyl cellulose, hydroxypropylmethylcellulose, hydroxyethylcellulose, sodium carboxymethylcellulose, calcium carboxymethylcellulose, methylcellulose, polyacrylic acid, maltodextrin, pregelatinized starch and polyvinyl alcohol, copolymers and mixtures thereof.

[0256] The release of the active ingredient from the composition can be achieved by the use of suitable retardants, including excipients well known in the pharmaceutical art for their release-retarding properties. Examples of such release retardants include, but are not limited to, polymeric release retardants, non-polymeric release retardants, or any combination thereof.

[0257] The polymer release retardant used for the purpose of the present invention includes, but is not limited to, cellulose derivatives; polyhydric alcohols; sugars, gums, and their derivatives; vinyl derivatives, polymers, copolymers, or mixtures thereof; maleic acid copolymers; polyalkylene oxides or copolymers thereof; acrylic acid polymers and acrylic acid derivatives; or any combination thereof.Cellulose derivatives include, but are not limited to, ethyl cellulose, methyl cellulose, hydroxypropyl methyl cellulose (HPMC), hydroxypropyl cellulose (HPC), hydroxyethyl cellulose, hydroxymethyl cellulose, hydroxyethyl methyl cellulose, carboxymethyl cellulose (CMC), or any combination thereof.Polyhydric alcohols include, but are not limited to, polyethylene glycol (PEG) or polypropylene glycol, or any combination thereof. Sugars, gums, and their derivatives include, but are not limited to, dextrin, polydextrin, dextran, pectin and pectin derivatives, alginic acid, sodium alginate, starch, hydroxypropyl starch, guar gum, locust bean gum, xanthan gum, karaya gum, tragacanth gum, carrageenan, acacia gum, gum arabic, fenugreek fiber, or gellan gum, or any combination thereof. Vinyl derivatives, polymers, copolymers, or mixtures thereof include, but are not limited to, polyvinyl acetate, polyvinyl alcohol, a mixture of polyvinyl acetate (8 parts w / w) and polyvinylpyrrolidone (2 parts w / w) (Kollidon SR), copolymers of vinylpyrrolidone, vinyl acetate copolymers, polyvinylpyrrolidone (PVP), or combinations thereof. Polyalkylene oxides or copolymers thereof include, but are not limited to, polyethylene oxide, polypropylene oxide, poly(oxyethylene)-poly(oxypropylene) block copolymers (poloxamers), or combinations thereof.Maleic acid copolymers include, but are not limited to, vinyl acetate maleic anhydride copolymers, butyl acrylate styrene maleic anhydride copolymers, etc., or any combination thereof. Acrylic acid polymers and acrylic acid derivatives include, but are not limited to, carbomers, methacrylic acid, polymethacrylic acid, polyacrylates, polymethacrylates, etc., or any combination thereof. Polymethacrylates include, but are not limited to, a) copolymers formed from monomers selected from methacrylic acid, methacrylic acid esters, acrylic acid, and acrylic acid esters, c) copolymers formed from monomers selected from ethyl acrylate, methyl methacrylate, and trimethylammonioethyl methacrylate chloride, etc., or any combination thereof. Non-polymeric release retardants for use in the present invention include, but are not limited to, fats, oils, waxes, fatty acids, fatty acid esters, long-chain monohydric alcohols, and esters thereof, or combinations thereof. In embodiments, non-polymeric release retardants for use in the present invention include, but are not limited to, Cutina (hydrogenated castor oil), Hydrobase (hydrogenated soybean oil), Castorwax (hydrogenated castor oil), Croduret (hydrogenated castor oil), Carbowax, Compritol (glyceryl behenate), Sterotex (hydrogenated cottonseed oil), Lubritab (hydrogenated cottonseed oil), Apifil (yellow wax), Akofine (hydrogenated cottonseed oil), Softtisan (hydrogenated palm oil), Hydrocote (hydrogenated soybean oil), Corona (lanolin), Gelucire (macrogolglyceride lauric), Precirol (glyceryl palmitostearate), Emulcire (cetyl alcohol), Pullulol diisostearic (polyglyceryl diisostearate), and Geleol (glyceryl stearate), and mixtures thereof.

[0258] The gastroretentive compositions of the present invention may be in the form of, but are not limited to, a monolithic or multi-layered dosage form or an inlay system. In one embodiment of the present invention, the gastroretentive composition is in the form of a bilayer or trilayered solid dosage form. In an exemplary embodiment, a solid pharmaceutical composition in the form of an expandable bilayer system for oral administration is adapted to deliver an active pharmaceutical ingredient from a first layer immediately after reaching the gastrointestinal tract and deliver an additional pharmaceutical agent, which may be the same or different, from a second layer in a modified manner over a specific period of time. The second layer may be formulated to expand within the composition, thereby extending the retention of the composition in the stomach.

[0259] In a further exemplary embodiment, a solid pharmaceutical composition for oral administration comprises two layers, one layer containing the active ingredient together with a suitable release-retardant, and the other layer containing a swelling agent in combination with other excipients. In another embodiment of the invention, a solid pharmaceutical composition for oral administration contains an inlay system, a special dosage form comprising a first tablet containing the active ingredient(s) disposed inside a second tablet containing excipients that ensure gastric retention. In this system, the tablet containing the active ingredient is small and is coated on all but at least one side with a blend of excipients, including a swellable polymer or a flotation system, or both, to ensure gastric retention.

[0260] In yet another embodiment of the present invention, the dosage form may be optionally coated. Surface coatings may be used for organoleptic purposes (especially thiols or disulfides with odors or unpleasant tastes), drug labeling purposes (e.g., color-coding systems for dosage forms), cosmetic purposes, dimensionally stabilizing the compressed dosage form, or delaying drug release. The surface coating may be any conventional coating suitable for enteral use. Coatings may be performed using any conventional technique using conventional ingredients. Surface coatings may be obtained using fast-dissolving films using conventional polymers, such as, but not limited to, hydroxypropylmethylcellulose, hydroxypropylcellulose, carboxymethylcellulose, polyvinyl alcohol, polymethacrylate, etc. Coating excipients and methods for using them are well known in the art. See, for example, McGinity, James W. and Linda A. Felton, Aqueous Polymeric Coatings for Pharmaceutical Dosage Forms, 3rd Edition, Informa Healthcare, 2008.

[0261] Furthermore, in another embodiment of the present invention, the composition may be in the form of a pellet, microsphere, microcapsule, microbead, microparticle, or nanoparticle, which has a prolonged intestinal transit time, for effective delivery of active agents requiring a longer intestinal residence time. The multiparticulate system may be (i) bioadhesive or mucoadhesive, thereby delaying gastrointestinal transit; (ii) float on the stomach contents, optionally forming a gel-like layer; (iii) coated with a pH-sensitive outer layer or layer that dissolves in the mildly acidic environment of the small intestine or in the neutral to slightly basic environment of the ileum (typically the intestinal segment with the highest pH); or (iv) formed using a drug-containing polymer that is not digested by human enzymes but is digested by enzymes produced by intestinal bacteria, leading to drug release in the distal ileum and colon. In an embodiment, the composition of the present invention is in the form of a multiparticulate and gastroretentive. Such multiparticulate systems may be prepared by methods including, but not limited to, pelleting, granulation, spray drying, spray congealing, etc.

[0262] Suitable polymeric release-controlling agents can be used in the compositions of the present invention. In one embodiment, the polymeric release-controlling agent is pH-independent or pH-dependent, or any combination thereof. In another embodiment, the polymeric release-controlling agent used in the compositions of the present invention can be swelling or non-swelling. In a further embodiment, the polymeric release-controlling agent that can be used in the compositions of the present invention includes, but is not limited to, cellulose derivatives, sugars or polysaccharides, poly(oxyethylene)-poly(oxypropylene) block copolymers (poloxamers), vinyl derivatives or polymers or copolymers thereof, polyalkylene oxides and derivatives thereof, maleic acid copolymers, acrylic acid derivatives, etc., or any combination thereof.

[0263] Controlled-release compositions for oral use may be constructed to release the active drug by controlling the dissolution and / or diffusion of the active drug substance. To achieve controlled release and thereby optimize the plasma concentration versus time profile, any of a number of strategies can be implemented. In one example, controlled release is achieved by the appropriate selection of various formulation parameters and components, including, for example, various types of controlled-release compositions and coatings. Thus, the drug is formulated with appropriate excipients into a pharmaceutical composition that releases the drug in a controlled manner upon administration. Examples include single or multiple unit tablet or capsule compositions, oily solutions, liquids, suspensions, emulsions, microcapsules, microspheres, nanoparticles, powders, and granules. In certain embodiments, the composition includes a biodegradable, pH-, and / or temperature-sensitive polymer coating.

[0264] Dissolution or diffusion controlled release can be achieved by suitable coating of the compound's tablet, capsule, pellet or granule formulation, or by incorporating the compound into a suitable matrix.The controlled release coating can include one or more of the above-mentioned coating materials, and / or for example, shellac, beeswax, glycowax, castor oil wax, carnauba wax, stearyl alcohol, glyceryl monostearate, glyceryl distearate, glycerol palmitostearate, ethyl cellulose, acrylic resin, di-polylactic acid, cellulose acetate butyrate, polyvinyl chloride, polyvinyl acetate, vinylpyrrolidone, polyethylene, polymethacrylate, methyl methacrylate, 2-hydroxymethacrylate, methacrylate hydrogel, 1,3-butylene glycol, ethylene glycol methacrylate, and / or polyethylene glycol. In controlled release matrix formulations, the matrix material may include, for example, hydrated methylcellulose, carnauba wax, and stearyl alcohol, Carbopol 934, silicone, glyceryl tristearate, methyl acrylate-methyl methacrylate, polyvinyl chloride, polyethylene, and / or halogenated fluorocarbons.

[0265] Alternatively, certain cysteamine precursors or enhancers of in vivo cysteamine production or absorption may be formulated and administered as medical foods. Medical foods are regulated by the US FDA as foods, not drugs. Methods for formulating medical foods are known in the art. For a description of methods for preparing and administering active compounds in foods or beverages, see, for example, US Patent Publication No. 2010 / 0261791. Nutracia, a medical food company based in the Netherlands, has more than 250 patent applications and patents describing methods for combining pharmacologically active agents with foods or beverages.

[0266] coating Pharmaceutical compositions formulated for oral delivery, such as tablets or capsules of the present invention, can be coated or otherwise compounded to provide dosage forms that offer the advantage of delayed or extended release. The coating can be adapted to release the active drug substance in a predetermined pattern (e.g., to achieve a controlled release formulation) or may be adapted not to release the active drug substance until after passage through the stomach, for example, by use of enteric coatings (e.g., polymers that are pH sensitive ("pH-controlled release"), polymers with slow or pH-dependent swelling rates, dissolution or erosion ("time-controlled release"), polymers that are enzymatically degraded ("enzyme-controlled release" or "biodegradable release"), and polymers that form a tough layer that is ruptured by increased pressure ("pressure-controlled release")). Exemplary enteric coatings that can be used in the pharmaceutical compositions described herein include sugar coatings, film coatings (e.g., based on hydroxypropylmethylcellulose, methylcellulose, methylhydroxyethylcellulose, hydroxypropylcellulose, carboxymethylcellulose, acrylate copolymers, polyethylene glycol, and / or polyvinylpyrrolidone), or coatings based on methacrylic acid copolymers, cellulose acetate phthalate, hydroxypropylmethylcellulose phthalate, hydroxypropylmethylcellulose acetate succinate, polyvinylacetate phthalate, shellac, and / or ethylcellulose. Additionally, time delay materials, such as, for example, glyceryl monostearate or glyceryl distearate, may be employed.

[0267] For example, the tablet or capsule can comprise an inner dosage and an outer dosage component, the latter being in the form of an envelope over the former, and the two components can be separated by an enteric layer which serves to resist disintegration in the stomach and permit the inner component to pass intact into the duodenum or to be delayed in release.

[0268] When an enteric coating is used, it is desirable that a substantial amount of the drug be released in the lower gastrointestinal tract. Alternatively, a leaky enteric coating can be used to provide a release profile intermediate between immediate-release and delayed-release formulations. For example, U.S. Patent Application No. 2008 / 0020041(A1) discloses a pharmaceutical formulation coated with an enteric material that releases at least a portion of the active ingredient upon contact with gastric fluid, with the remaining ingredients being released upon contact with intestinal fluid.

[0269] In addition to coatings that provide delayed or extended release, solid tablet compositions may include coatings adapted to protect the composition from unwanted chemical changes (e.g., chemical degradation prior to the release of the active drug substance). Coatings can be applied to solid dosage forms in a manner similar to those described in Encyclopedia of Pharmaceutical Technology, Vols. 5 and 6, Swarbrick and Boyland, eds., 2000.

[0270] In the case of controlled-release formulations, the active ingredient of the composition can be targeted for release in the small intestine. The formulation can include an enteric coating so that the composition is resistant to the low pH environment found in the stomach but is sensitive to the higher pH environment of the small intestine. To control the release of the active ingredient in the small intestine, a multiparticulate formulation can be used to prevent simultaneous release of the active ingredients. The multiparticulate composition can include multiple individual enteric-coated cores containing a hydrophobic phase containing a cysteamine precursor or its salt dispersed in a microcrystalline cellulose-based gel and a hydrophilic phase containing a hydrogel. The microcrystalline cellulose (MCC) acts as a release-controlling polymer for the cysteamine precursor or its salt while the core is dissolving or eroding in the intestine, preventing dose dumping and stabilizing the cysteamine precursor or its salt. Two or more multiparticulate compositions that differ in terms of excipients in the core or coating layer can be combined into a single pharmaceutical composition (e.g., capsule, powder, or liquid) to release the active ingredient (e.g., cysteamine precursor) over a longer period of time. Alternatively, the same effect can be achieved by using different concentrations of excipients in two or more batches of microparticles and then combining the microparticles from the different batches in a ratio (e.g., 1:1) selected to result in a targeted drug release profile.

[0271] The composition comprises about 15% w / w to about 70% w / w of a cysteamine precursor or salt thereof, about 25% w / w to about 75% w / w of microcrystalline cellulose, and about 2% w / w to about 15% w / w of methylcellulose, where % w / w is the % w / w of the enteric coated core.

[0272] In some cases, it may be advantageous to include a continuous proteinaceous subcoating layer that covers and separates the individual cores from their respective enteric coatings to further enhance the stability of the cysteamine precursor or salt thereof. The continuous proteinaceous subcoating is adapted to prevent mixing of the cysteamine precursor or salt thereof with the enteric coating. Some preferred proteinaceous subcoatings have the following attributes: the subcoating can include a gelatin film adhered to the core, and / or the subcoating can include a dried proteinaceous gel.

[0273] In certain embodiments, the enteric coated core releases about 20% or less of the cysteamine precursor or salt thereof within about 2 hours when placed in a 0.1 N HCl solution, and then releases about 85% or more of the cysteamine precursor or salt thereof within about 8 hours when placed in a substantially neutral pH environment.

[0274] Preferably, the enteric coated core is spheroidal and has a diameter of 3 mm or less.

[0275] In order to prevent the adhesion of separately administered compositions in the stomach, the composition of the present invention can be coated with an anti-adhesion agent.Anti-adhesion agents can also be used to prevent microparticles from sticking together.For example, the composition can be coated with a thin outermost layer of microcrystalline cellulose powder.Alternatively, adhesion can be prevented by coating with a polymer that is insoluble in gastric juice but is permeable and swellable.For example, 30% polyacrylate dispersion (e.g., Eudragit NE30D, Evonik Industries) has been shown to prevent the adhesion of floating mini-tablets in the stomach (see Rouge et al., European Journal of Pharmaceutics and Biopharmaceutics 43:165(1997)).

[0276] Commercially available forms of the listed excipients used in enteric coatings include, for example, various brands of polymethacrylates (a chemically homogeneous group of compounds including amino methacrylate copolymers, ammonio methacrylate copolymers, ethyl acrylate copolymer dispersions, methyl methacrylate copolymer dispersions, methacrylic acid copolymers, and methacrylic acid copolymer dispersions), which are sold as product lines by companies including, but not limited to, Ashland, BASF Fine Chemicals (Kollicoat product line), ColorCon (Acryl-EZE product line), Eastman Chemical (Eastacryl product line), and Evonik Industries (Eudragit product line).

[0277] Formulations for ileal and colonic drug release In some embodiments, cysteamine precursors can be delivered to the distal ileum and colon using ileum- and / or colon-targeted formulations. (The term "colon-targeted" is used herein to refer to both ileum- and colon-targeted formulations. Any composition that begins to release drug in the ileum may also release drug in the colon, and some drug released in the ileum may reach the colon.) Advantages of drug delivery with colon-targeted compositions include prolonged contact with the colonic epithelium and the presence of colonic bacteria available for site-specific delivery.

[0278] From a pharmacokinetic perspective, colonic absorption of cysteamine is desirable because, due to its extremely short half-life, it must be continuously produced (and absorbed) in the gastrointestinal tract to maintain blood levels within the therapeutic range. An ingested pharmaceutical composition (otherwise known as a gastroretentive composition) can reach the colon 3-5 hours after ingestion (on average, in most subjects) when taken in a fasted state, or 6-10 hours after ingestion (on average, in most subjects) with food. The only way to maintain blood cysteamine levels within the therapeutic range after the dosage form reaches the colon is to ensure that cysteamine is produced and absorbed in the colon. Some cysteamine precursors released in the small intestine may enter the colon intact and be degraded to cysteamine in the colon. However, to provide robust cysteamine production in the colon, cysteamine precursors should be formulated for release in the colon (or ileum), where they can be degraded to cysteamine and absorbed. The colon-targeted composition is not intended to be used alone as a treatment for cysteamine-sensitive disorders, but rather to complement formulations aimed at other areas of the gastrointestinal tract.

[0279] Two approaches to colon-targeted delivery have been widely developed and are described below.

[0280] The first approach involves utilizing enzymes produced in the colon by intestinal bacteria. Intestinal bacteria can digest various polymers that are not digested by human enzymes present in saliva, gastric juice, intestinal juice, or pancreatic juice. Pharmaceutical compositions containing such polymers cannot be digested, and therefore, the active ingredients mixed with the polymers cannot escape until they encounter enzymes produced by intestinal bacteria in the distal ileum (where bacterial density begins to increase) or colon (where there can be as many as one trillion bacteria per milliliter of colonic content).

[0281] Cysteamine precursors and / or other active ingredients (e.g., enhancers of in vivo cysteamine production or absorption) can be mixed with polymers that delay drug release and are digestible only (in the human gastrointestinal tract) by enzymes produced by intestinal bacteria. Polymers used for colon-targeted drug delivery based on selective degradation by intestinal bacteria include dextran hydrogels (Hovgaard, L. and H. Brondsted, J. Controlled ReI. 36:159 (1995)), cross-linked chondroitin (Rubinstein et al., Pharm. Res. 9:276 (1992)), and hydrogels containing azoaromatic moieties (Brondsted, H. and J. Kopoecek, Pharm. Res. 9:1540 (1992) and Yeh et al., J. Controlled ReI. 36:109 (1995)).

[0282] Covalent binding of drugs to carriers forms stable precursors in the stomach and small intestine, and releases the drug in the large intestine upon enzymatic cleavage by intestinal microflora; examples of these precursors include azo complexes, cyclodextrin complexes, glycoside complexes, glucuronate complexes, dextran complexes, polypeptides, and polymer complexes. The basic principle is that the covalent bond linking the drug and carrier must be indigestible by human enzymes but digestible by intestinal bacterial enzymes.

[0283] The second approach involves utilizing a higher pH in the ileum relative to other parts of the gastrointestinal tract. In healthy subjects, the pH of the gastrointestinal tract increases from the duodenum (approximately pH 5.5-6.6 from the proximal to distal duodenum) to the terminal ileum (approximately pH 7-7.5), then decreases in the cecum (pH approximately 6.4), and increases again from the right to the left side of the colon to a final value of approximately pH 7.

[0284] The composition can be coated with a pH-sensitive polymer that dissolves only at neutral to mildly alkaline pH (e.g., pH 6.5 or higher, pH 6.8 or higher, or pH 7 or higher). Beneath the pH-sensitive coating is a sustained-release formulation from which the drug is gradually released by diffusion, erosion, or a combination. This approach is described in U.S. Pat. No. 5,900,252, incorporated herein by reference.

[0285] Colonic targeting methods based on gut bacteria and pH can be combined. See, for example, Naeem et al., Colloids Surf B Biointerfaces S0927 (2014). This study describes coated nanoparticles formed using bacterially digestible polymers. Another technique combining pH and bacterial enzyme digestion for colonic delivery of drug-containing liquid-filled capsules is described in U.S. Patent Publication No. 2007 / 0243253, which discloses formulations utilizing polymers including starch, amylose, amylopectin, chitosan, chondroitin sulfate, cyclodextrin, dextran, pullulan, carrageenan, scleroglucan, chitin, curduran, and levan, along with a pH-sensitive coating that dissolves at about pH 5 or above.

[0286] Other approaches for colon-targeted drug delivery use (i) time-release systems, in which the outer coating begins to dissolve as the multi-coated formulation pass...

Claims

1. 1. A pharmaceutical for treating a cysteamine-sensitive disorder in a subject, the pharmaceutical comprising N-acetylcysteine-pantetheine disulfide or cysteamine-N-acetylcysteine ​​disulfide, or a pharmaceutically acceptable salt thereof, at a dose of 50 to 150 milligrams per kilogram of body weight (mg / kg), the pharmaceutical being orally administered to the subject one or more times daily.

2. 2. The method of claim 1, wherein the method is not formulated for administration within two hours of administering a reducing agent to the subject.

3. 3. The medicament of claim 1 or 2, formulated for administration to the subject once, twice, or three times daily.

4. The medicament according to any one of claims 1 to 3, wherein the dose is 2,000 to 8,000 milligrams of N-acetylcysteine-pantetheine disulfide or cysteamine-N-acetylcysteine ​​disulfide, or a pharmaceutically acceptable salt thereof.

5. The pharmaceutical composition according to any one of claims 1 to 4, which is formulated for immediate release.

6. 6. The pharmaceutical of claim 5, (i) formulated as a powder, in the form of a sachet, (ii) formulated as a tablet or capsule, or (iii) formulated as a liquid solution or suspension.

7. 7. The pharmaceutical composition of any one of claims 1 to 6, wherein the cysteamine-sensitive disorder is selected from cystinosis; neurodegenerative diseases; neurodevelopmental diseases; neuropsychiatric diseases; mitochondrial diseases; fibrotic diseases of the kidney, liver, or lung; parasitic infections; sickle cell anemia; cancer; ischemic diseases including ischemic heart disease or stroke; chronic obstructive pulmonary disease (COPD); cystic fibrosis (CF); bacterial infections; viral infections; non-alcoholic steatohepatitis (NASH); alcoholic steatohepatitis; and non-alcoholic fatty liver disease (NAFLD).

8. The pharmaceutical composition of claim 7, wherein the cysteamine-sensitive disorder is a neurodegenerative disease selected from the group consisting of Huntington's disease, neurodegeneration associated with brain iron accumulation, Parkinson's disease, and Alzheimer's disease.

9. The pharmaceutical composition according to claim 7, wherein the cysteamine-sensitive disorder is a neurodevelopmental disease selected from Rett syndrome and other MECP2 mutation-associated disorders.

10. The pharmaceutical composition of claim 7, wherein the cysteamine-sensitive disorder is a mitochondrial disease selected from Leigh syndrome, MELAS, MERFF, and Friedreich's ataxia.

11. The pharmaceutical composition of claim 7, wherein the cysteamine-sensitive disorder is a fibrotic disease selected from Alport's disease, focal segmental glomerulosclerosis (FSGS), alcoholic steatohepatitis (ASH), and pulmonary fibrosis.

12. The pharmaceutical composition according to claim 7, wherein the cysteamine-sensitive disorder is a bacterial infection or a viral infection.

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