Methods for Treating Hyperphenylalaninemia

Abstract

The present invention features a method for lowering blood phenylalanine levels in a subject by administering sepiapterin or a pharmaceutically acceptable salt thereof.

Description

[Background technology]

[0001] Phenylketonuria (PKU) is an inborn error of metabolism caused primarily by mutations in the phenylalanine hydroxylase (PAH) gene. Genetic mutations in PAH result in reduced catalytic activity leading to hyperphenylalaninemia (HPA). There are many mutations in the PAH gene (>400), resulting in phenotypic differences in the amount of enzyme produced and / or enzyme activity. Subjects with severe PKU have a complete or severe deficiency in PAH enzyme activity and typically have very high blood phenylalanine (Phe) concentrations (>1200 micromoles / liter). Partial deficiencies in PAH activity result in lower degrees of blood phenylalanine elevation (e.g., about 360–1200 micromoles / liter). High concentrations of phenylalanine are toxic to the brain. If left untreated, severe and irreversible intellectual disability can occur. With the introduction of newborn screening almost worldwide, PKU can now be diagnosed at birth. It has been reported in all ethnic groups and is estimated to occur in approximately 1 in 10,000 people.

[0002] Currently, there is no cure for PKU. Initial treatment consists of rapid implementation of a phenylalanine dietary restriction supplemented with specially designed medical foods. Adherence to the dietary restriction can be difficult for older children, adolescents, and adults. Tetrahydrobiopterin (BH4) supplementation has demonstrated clinically meaningful reductions in phenylalanine plasma concentrations in approximately 30% of PKU patients. BH4 is an essential cofactor for PAH in the conversion of Phe to tyrosine (Tyr). BH4 supplementation has been shown to result in increased intracellular BH4, improved PAH function, and reduced plasma phenylalanine. Kuvan® (sapropterin dihydrochloride), a synthetic formulation of BH4, has been approved as a drug in several countries for use in the treatment of hyperphenylalaninemia (HPA) in tetrahydrobiopterin-responsive adult and pediatric PKU patients. However, control with dietary phenylalanine restriction and / or BH4 supplementation is suboptimal, and the majority of patients maintain elevated phenylalanine plasma concentrations. Another treatment for PKU, pegvaliase-pqpz, is approved only for adult patients and involves daily injections of a phenylalanine-metabolizing enzyme. However, this approach has potential complications from side effects such as anaphylaxis, and many patients continue to have elevated phenylalanine levels despite treatment. Thus, additional methods to treat PKU are needed. Summary of the Invention

[0003] The present invention features a method for reducing blood phenylalanine levels in a subject in need thereof by administering sepiapterin or a pharma- ceutically acceptable salt thereof. The inventors have found that sepiapterin is effective in reducing blood phenylalanine levels. In particular, the inventors have found that sepiapterin is effective in reducing blood phenylalanine levels to levels similar to those of healthy subjects and / or in reducing blood phenylalanine levels in subjects who have failed prior treatment with sapropterin dihydrochloride and / or pegvalase pqpz.

[0004] In one aspect, the invention features a method of reducing phenylalanine levels (e.g., blood phenylalanine concentrations) in a subject (e.g., a subject with hyperphenylalaninemia, a subject with hyperphenylalaninemia due to phenylketonuria, a subject with phenylketonuria, a subject with tetrahydrobiopterin-responsive phenylketonuria, or a subject with sepiapterin-responsive phenylketonuria), the method including administering an effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof.

[0005] In one aspect, the invention features a method of treating hyperphenylalaninemia in a subject in need thereof (e.g., a subject with hyperphenylalaninemia due to phenylketonuria, such as tetrahydrobiopterin-responsive phenylketonuria, or sepiapterin-responsive phenylketonuria), the method includes administering an effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof.

[0006] In one aspect, the invention features a method of treating phenylketonuria in a subject in need thereof (e.g., a subject with tetrahydrobiopterin-responsive phenylketonuria, or sepiapterin-responsive phenylketonuria, classical phenylketonuria, and / or non-classical phenylketonuria), the method includes administering an effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof.

[0007] In some embodiments of any of the methods described herein, the subject has a blood phenylalanine concentration (e.g., unregulated phenylalanine blood concentration) of greater than 120 micromoles / liter (e.g., greater than 200 micromoles / liter, greater than 300 micromoles / liter, greater than 360 micromoles / liter, greater than 400 micromoles / liter, greater than 450 micromoles / liter, greater than 500 micromoles / liter, greater than 550 micromoles / liter, greater than 600 micromoles / liter, greater than 650 micromoles / liter, greater than 700 micromoles / liter, greater than 800 micromoles / liter, greater than 900 micromoles / liter, greater than 1000 micromoles / liter, greater than 1100 micromoles / liter, or greater than 1200 micromoles / liter). In some embodiments of any of the methods described herein, the subject has a blood phenylalanine concentration (e.g., an unregulated phenylalanine blood concentration) of 120-360 micromoles / liter, 360-600 micromoles / liter, 600-1200 micromoles / liter, or greater than 1200 micromoles / liter.

[0008] In some embodiments of any of the methods described herein, the subject has a blood phenylalanine concentration (e.g., an unregulated phenylalanine blood concentration) of greater than 400 micromoles / liter (e.g., greater than 450 micromoles / liter, greater than 500 micromoles / liter, greater than 550 micromoles / liter, greater than 600 micromoles / liter, greater than 650 micromoles / liter, greater than 700 micromoles / liter, greater than 800 micromoles / liter, greater than 900 micromoles / liter, greater than 1000 micromoles / liter, greater than 1100 micromoles / liter, or greater than 1200 micromoles / liter) under existing management, such as a phenylalanine restricted diet, and / or treatment with sapropterin or a pharma- ceutically acceptable salt thereof (e.g., sapropterin dihydrochloride). In some embodiments, treatment with sapropterin comprises administration of 5-20 mg / kg of sapropterin or a pharma- ceutically acceptable salt thereof.

[0009] In some embodiments of any of the methods described herein, the subject did not respond to treatment with sapropterin or a pharma- ceutically acceptable salt thereof (e.g., sapropterin dihydrochloride). In some embodiments, administration of sapropterin or a pharma- ceutically acceptable salt thereof (e.g., sapropterin dihydrochloride) reduces the subject's plasma concentration by less than 15-30%. In some embodiments, administration of sapropterin or a pharma- ceutically acceptable salt thereof (e.g., sapropterin dihydrochloride) reduces the subject's plasma concentration by less than 30%. In some embodiments, administration of sapropterin or a pharma- ceutically acceptable salt thereof (e.g., sapropterin dihydrochloride) reduces the subject's plasma concentration by less than 20%. In some embodiments, administration of sapropterin or a pharma- ceutically acceptable salt thereof (e.g., sapropterin dihydrochloride) reduces the subject's plasma concentration by less than 15%. For example, administration of at least about 10 mg / kg (e.g., at least about 15 mg / kg, at least about 20 mg / kg) of sapropterin or a pharma- ceutically acceptable salt thereof (e.g., sapropterin dihydrochloride) for at least 8 days (e.g., at least 14 days, at least 21 days, at least 28 days, at least 30 days) reduced the subject's plasma concentration by less than 30%. In some embodiments, after administration of sapropterin or a pharma- ceutically acceptable salt thereof (e.g., sapropterin dihydrochloride), the subject's plasma concentration is greater than 120 micromoles / liter (e.g., greater than 200 micromoles / liter, greater than 300 micromoles / liter, greater than 360 micromoles / liter, greater than 400 micromoles / liter, greater than 450 micromoles / liter, greater than 500 micromoles / liter, greater than 550 micromoles / liter, greater than 600 micromoles / liter, greater than 650 micromoles / liter, greater than 700 micromoles / liter, greater than 800 micromoles / liter, greater than 900 micromoles / liter, greater than 1000 micromoles / liter, greater than 1100 micromoles / liter, or greater than 1200 micromoles / liter).For example, after administration of at least about 10 mg / kg (e.g., at least about 15 mg / kg, at least about 20 mg / kg) of sapropterin or a pharma- ceutically acceptable salt thereof (e.g., sapropterin dihydrochloride) for at least 8 days (e.g., at least 14 days, at least 21 days, at least 28 days, at least 30 days), the subject's plasma concentration is greater than 120 micromoles / liter (e.g., greater than 200 micromoles / liter, greater than 300 micromoles / liter, greater than 360 micromoles / liter, greater than 400 micromoles / liter, greater than 450 micromoles / liter, greater than 500 micromoles / liter, greater than 550 micromoles / liter, greater than 600 micromoles / liter, greater than 650 micromoles / liter, greater than 700 micromoles / liter, greater than 800 micromoles / liter, greater than 900 micromoles / liter, greater than 1000 micromoles / liter, greater than 1100 micromoles / liter, or greater than 1200 micromoles / liter).

[0010] In some embodiments of any of the methods described herein, the subject did not respond to treatment with pegvaliase-pqpz. In some embodiments, administration of pegvaliase-pqpz reduced the subject's plasma concentration by less than 20%. For example, the subject's plasma concentration reduced by less than 30%, or less than 20%, when administered pegvaliase-pqpz at least about 20 mg once daily (e.g., at least about 30 mg once daily, at least about 40 mg once daily) for at least 16 weeks (e.g., at least 18 weeks, at least 20 weeks, at least 22 weeks, at least 24 weeks). In some embodiments, after administration of pegvaliase-pqpz, the subject's plasma concentration is greater than 120 micromoles / liter (e.g., greater than 200 micromoles / liter, greater than 300 micromoles / liter, greater than 360 micromoles / liter, greater than 400 micromoles / liter, greater than 450 micromoles / liter, greater than 500 micromoles / liter, greater than 550 micromoles / liter, greater than 600 micromoles / liter, greater than 650 micromoles / liter, greater than 700 micromoles / liter, greater than 800 micromoles / liter, greater than 900 micromoles / liter, greater than 1000 micromoles / liter, greater than 1100 micromoles / liter, or greater than 1200 micromoles / liter).For example, after administration of at least about 20 mg (e.g., at least about 30 mg, at least about 40 mg) of pegvariase-pqpz for at least 16 weeks (e.g., at least 18 weeks, at least 20 weeks, at least 22 weeks, at least 24 weeks), the subject's plasma concentration was greater than 120 micromoles / liter (e.g., greater than 200 micromoles / liter, greater than 300 micromoles / liter, greater than 360 micromoles / liter, greater than 400 micromoles / liter, greater than 450 micromoles / liter, greater than 500 micromoles / liter, greater than 550 micromoles / liter, greater than 600 micromoles / liter, greater than 650 micromoles / liter, greater than 700 micromoles / liter, greater than 800 micromoles / liter, greater than 900 micromoles / liter, greater than 1000 micromoles / liter, greater than 1100 micromoles / liter, or greater than 1200 micromoles / liter).

[0011] In some embodiments of any of the methods described herein, the subject discontinued treatment with pegvalase pqpz due to adverse reactions (e.g., anaphylaxis, injection site reactions, arthralgia, hypersensitivity reactions, headache, generalized skin reactions lasting at least 14 days, pruritus, nausea, abdominal pain, oropharyngeal soreness, vomiting, cough, diarrhea, and / or fatigue) and / or tolerability.

[0012] In some embodiments of any of the methods described herein, administering sepiapterin or a pharma- tically acceptable salt thereof reduces the subject's blood phenylalanine concentration to less than 600 micromoles / liter. In some embodiments of any of the methods described herein, administering sepiapterin or a pharma- tically acceptable salt thereof reduces the subject's blood phenylalanine concentration to less than 360 micromoles / liter. In some embodiments of any of the methods described herein, administering sepiapterin or a pharma- tically acceptable salt thereof reduces the subject's blood phenylalanine concentration to less than 120 micromoles / liter. In some embodiments of any of the methods described herein, administering sepiapterin or a pharma- tically acceptable salt thereof reduces the subject's blood phenylalanine concentration to between 120 and 360 micromoles / liter. In some embodiments of any of the methods described herein, administering sepiapterin or a pharma- tically acceptable salt thereof reduces the subject's blood phenylalanine concentration to between 360 and 600 micromoles / liter.

[0013] In some embodiments of any of the methods described herein, the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is sufficient to reduce the subject's blood phenylalanine concentration to less than 600 micromoles / liter. In some embodiments of any of the methods described herein, the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is sufficient to reduce the subject's blood phenylalanine concentration to less than 360 micromoles / liter. In some embodiments of any of the methods described herein, the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is sufficient to reduce the subject's blood phenylalanine concentration to less than 120 micromoles / liter. In some embodiments of any of the methods described herein, the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is sufficient to reduce the subject's blood phenylalanine concentration to between 120 and 360 micromoles / liter. In some embodiments of any of the methods described herein, the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is an amount sufficient to reduce the subject's blood phenylalanine concentration to between 360 and 600 micromoles per liter.

[0014] In some embodiments of any of the methods described herein, administering sepiapterin or a pharma- ceutically acceptable salt thereof reduces the subject's blood phenylalanine concentration by at least 10% (e.g., at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or about 10% to about 30%, about 20% to about 40%, about 30% to about 50%, about 40% to about 60%, about 50% to about 70%, about 60% to about 80%, or about 70% to about 90%). The amount of reduction may be measured after at least one week of administration (e.g., two weeks, three weeks, four weeks, five weeks, six weeks).

[0015] In some embodiments of any of the methods described herein, the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is an amount sufficient to reduce the blood phenylalanine concentration in a subject by at least 10% (e.g., at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or about 10% to about 30%, about 20% to about 40%, about 30% to about 50%, about 40% to about 60%, about 50% to about 70%, about 60% to about 80%, or about 70% to about 90%).

[0016] In some embodiments of any of the methods described herein, administration of sepiapterin or a pharma- ceutically acceptable salt thereof provides a therapeutically effective dose of at least about 1000 mg / day (e.g., at least about 1100 mg / day, at least about 1200 mg / day, at least about 1300 mg / day, at least about 1400 mg / day, at least about 1500 mg / day, at least about 1600 mg / day, at least about 1700 mg / day, at least about 1800 mg / day, at least about 1900 mg / day, at least about 2000 mg / day, at least about 2100 mg / day, at least about 2200 mg / day, at least about 2300 mg / day, at least about 2400 mg / day, at least about 2500 mg / day, at least about 2600 mg / day, at least about 2700 mg / day, at least about 2800 mg / day, at least about 2900 mg / day, at least about 3000 mg / day, at least about 3100 mg / day, at least about 3200 mg / day, at least about 3300 mg / day, at least about 3400 mg / day, at least about 3500 mg / day, at least about 3600 mg / day, at least about 3700 mg / day, at least about 3800 mg / day, at least about 3900 mg / day, at least about 4000 mg / day, at least about 4100 mg / day, at least about 4200 mg / day, at least about 4300 mg / day, at least about 4400 mg / day, at least about 4500 mg / day, at least about 4600 mg / day, at least about 4700 mg / day, at least about 4800 mg / day, day, or at least about 2000 mg / day, or 1000 mg / day to 1400 mg / day, 1200 mg / day to 1600 mg / day, 1300 mg / day to 1700 mg / day, 1600 mg / day to 2000 mg / day, 1800 mg / day to 2400 mg / day, 2000 mg / day to 3000 mg / day, 3000 mg / day to 4000 mg / day, 4000 to 5000 mg / day), which reduces the subject's blood phenylalanine concentration to less than 600 micromoles / liter. In some embodiments of any of the methods described herein, administration of sepiapterin or a pharma- ceutically acceptable salt thereof provides a therapeutically effective dose of at least about 1000 mg / day (e.g., at least about 1100 mg / day, at least about 1200 mg / day, at least about 1300 mg / day, at least about 1400 mg / day, at least about 1500 mg / day, at least about 1600 mg / day, at least about 1700 mg / day, at least about 1800 mg / day, at least about 1900 mg / day, at least about 2000 mg / day, at least about 2100 mg / day, at least about 2200 mg / day, at least about 2300 mg / day, at least about 2400 mg / day, at least about 2500 mg / day, at least about 2600 mg / day, at least about 2700 mg / day, at least about 2800 mg / day, at least about 2900 mg / day, at least about 3000 mg / day, at least about 3100 mg / day, at least about 3200 mg / day, at least about 3300 mg / day, at least about 3400 mg / day, at least about 3500 mg / day, at least about 3600 mg / day, at least about 3700 mg / day, at least about 3800 mg / day, at least about 3900 mg / day, at least about 4000 mg / day, at least about 4100 mg / day, at least about 4200 mg / day, at least about 4300 mg / day, at least about 4400 mg / day, at least about 4500 mg / day, at least about 4600 mg / day, at least about 4700 mg / day, at least about 4800 mg / day, day, or at least about 2000 mg / day, or 1000 mg / day to 1400 mg / day, 1200 mg / day to 1600 mg / day, 1300 mg / day to 1700 mg / day, 1600 mg / day to 2000 mg / day, 1800 mg / day to 2400 mg / day, 2000 mg / day to 3000 mg / day, 3000 mg / day to 4000 mg / day, 4000 to 5000 mg / day), which reduces the subject's blood phenylalanine concentration to less than 360 micromoles / liter.In some embodiments of any of the methods described herein, administration of sepiapterin or a pharma- ceutically acceptable salt thereof provides a therapeutically effective dose of at least about 1000 mg / day (e.g., at least about 1100 mg / day, at least about 1200 mg / day, at least about 1300 mg / day, at least about 1400 mg / day, at least about 1500 mg / day, at least about 1600 mg / day, at least about 1700 mg / day, at least about 1800 mg / day, at least about 1900 mg / day, at least about 2000 mg / day, at least about 2100 mg / day, at least about 2200 mg / day, at least about 2300 mg / day, at least about 2400 mg / day, at least about 2500 mg / day, at least about 2600 mg / day, at least about 2700 mg / day, at least about 2800 mg / day, at least about 2900 mg / day, at least about 3000 mg / day, at least about 3100 mg / day, at least about 3200 mg / day, at least about 3300 mg / day, at least about 3400 mg / day, at least about 3500 mg / day, at least about 3600 mg / day, at least about 3700 mg / day, at least about 3800 mg / day, at least about 3900 mg / day, at least about 4000 mg / day, at least about 4100 mg / day, at least about 4200 mg / day, at least about 4300 mg / day, at least about 4400 mg / day, at least about 4500 mg / day, at least about 4600 mg / day, at least about 4700 mg / day, at least about 4800 mg / day, day, or at least about 2000 mg / day, or 1000 mg / day to 1400 mg / day, 1200 mg / day to 1600 mg / day, 1300 mg / day to 1700 mg / day, 1600 mg / day to 2000 mg / day, 1800 mg / day to 2400 mg / day, 2000 mg / day to 3000 mg / day, 3000 mg / day to 4000 mg / day, 4000 to 5000 mg / day), which reduces the subject's blood phenylalanine concentration to less than 120 micromoles / liter. In some embodiments of any of the methods described herein, administration of sepiapterin or a pharma- ceutically acceptable salt thereof provides a therapeutically effective dose of at least about 1000 mg / day (e.g., at least about 1100 mg / day, at least about 1200 mg / day, at least about 1300 mg / day, at least about 1400 mg / day, at least about 1500 mg / day, at least about 1600 mg / day, at least about 1700 mg / day, at least about 1800 mg / day, at least about 1900 mg / day, or at least about 2000 mg / day, or 1000 mg / day to 1400 mg / day, 1200 mg / day to 1600 mg / day, 1300 mg / day to 1700 mg / day, 1600 mg / day to 2000 mg / day, 1800 mg / day to 2400 mg / day, 2000 mg / day to 3000 mg / day, 3000 mg / day to 4000 mg / day, 4000 mg / day to 5000 mg / day), which reduces the subject's blood phenylalanine concentration to between 120 and 360 micromoles / liter.In some embodiments of any of the methods described herein, administration of sepiapterin or a pharma- ceutically acceptable salt thereof provides a therapeutically effective dose of at least about 1000 mg / day (e.g., at least about 1100 mg / day, at least about 1200 mg / day, at least about 1300 mg / day, at least about 1400 mg / day, at least about 1500 mg / day, at least about 1600 mg / day, at least about 1700 mg / day, at least about 1800 mg / day, at least about 1900 mg / day, or at least about 2000 mg / day, or 1000 mg / day to 1400 mg / day, 1200 mg / day to 1600 mg / day, 1300 mg / day to 1700 mg / day, 1600 mg / day to 2000 mg / day, 1800 mg / day to 2400 mg / day, 2000 mg / day to 3000 mg / day, 3000 mg / day to 4000 mg / day, 4000 mg / day to 5000 mg / day), which reduces the subject's blood phenylalanine concentration to between 360 and 600 micromoles / liter.

[0017] In some embodiments of any of the methods described herein, administration of sepiapterin or a pharma- ceutically acceptable salt thereof reduces the subject's blood phenylalanine concentration to less than 600 micromoles / liter at a natural protein intake of greater than 10 g / day (e.g., greater than 20 g / day, greater than 30 g / day, greater than 40 g / day, greater than 50 g / day, greater than 60 g / day, greater than 70 g / day, greater than 80 g / day, or from about 10 g / day to about 30 g / day, from about 20 g / day to about 40 g / day, from about 30 g / day to about 50 g / day, from about 40 g / day to about 60 g / day, from about 50 g / day to about 70 g / day, from about 60 g / day to about 80 g / day). In some embodiments of any of the above-described methods, administration of sepiapterin or any pharma- ceutically acceptable salt thereof reduces the subject's blood phenylalanine concentration to less than 360 micromoles / liter at a natural protein intake of greater than 10 g / day (e.g., greater than 20 g / day, greater than 30 g / day, greater than 40 g / day, greater than 50 g / day, greater than 60 g / day, greater than 70 g / day, greater than 80 g / day, or from about 10 g / day to about 30 g / day, from about 20 g / day to about 40 g / day, from about 30 g / day to about 50 g / day, from about 40 g / day to about 60 g / day, from about 50 g / day to about 70 g / day, from about 60 g / day to about 80 g / day). In some embodiments of any of the methods described herein, administration of sepiapterin or any pharma- ceutically acceptable salt thereof reduces the subject's blood phenylalanine concentration to less than 120 micromoles / liter at a natural protein intake of greater than 10 g / day (e.g., greater than 20 g / day, greater than 30 g / day, greater than 40 g / day, greater than 50 g / day, greater than 60 g / day, greater than 70 g / day, greater than 80 g / day, or from about 10 g / day to about 30 g / day, from about 20 g / day to about 40 g / day, from about 30 g / day to about 50 g / day, from about 40 g / day to about 60 g / day, from about 50 g / day to about 70 g / day, from about 60 g / day to about 80 g / day).In some embodiments of any of the methods described herein, administration of sepiapterin or any pharma- ceutically acceptable salt thereof reduces the subject's blood phenylalanine concentration to between 120 and 360 micromoles / liter at a natural protein intake of greater than 10 g / day (e.g., greater than 20 g / day, greater than 30 g / day, greater than 40 g / day, greater than 50 g / day, greater than 60 g / day, greater than 70 g / day, greater than 80 g / day, or between about 10 g / day and about 30 g / day, between about 20 g / day and about 40 g / day, between about 30 g / day and about 50 g / day, between about 40 g / day and about 60 g / day, between about 50 g / day and about 70 g / day, between about 60 g / day and about 80 g / day). In some embodiments of any of the methods described herein, administration of sepiapterin or any pharma- ceutically acceptable salt thereof reduces the subject's blood phenylalanine concentration to between 360 and 600 micromoles / liter at a natural protein intake of greater than 10 g / day (e.g., greater than 20 g / day, greater than 30 g / day, greater than 40 g / day, greater than 50 g / day, greater than 60 g / day, greater than 70 g / day, greater than 80 g / day, or between about 10 g / day and about 30 g / day, between about 20 g / day and about 40 g / day, between about 30 g / day and about 50 g / day, between about 40 g / day and about 60 g / day, between about 50 g / day and about 70 g / day, between about 60 g / day and about 80 g / day).

[0018] In some embodiments of any of the methods described herein, administration of sepiapterin or a pharma- ceutically acceptable salt thereof results in a decrease in the subject's blood phenylalanine concentration by at least 20% (e.g., at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%) from the blood phenylalanine concentration prior to administration of sepiapterin or a pharma- ceutically acceptable salt thereof.

[0019] In some embodiments of any of the methods described herein, administration of sepiapterin or a pharma- ceutically acceptable salt thereof results in a BH4 concentration of at least 50 ng / ml (e.g., at least 60 ng / ml, at least 100 ng / ml, at least 200 ng / ml, at least 400 ng / ml, at least 600 ng / ml, at least 1000 ng / ml, or at least 2000 ng / ml, or between 50 ng / ml and 100 ng / ml, between 60 ng / ml and 400 ng / ml, and between 200 ng / ml and 600 ng / ml, between 400 ng / ml and 1000 ng / ml, or between 600 ng / ml and 1500 ng / ml) in the subject's plasma within 10 hours after administration.

[0020] In some embodiments of any of the methods described herein, the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is at least about 1000 mg / day (e.g., at least about 1100 mg / day, at least about 1200 mg / day, at least about 1300 mg / day, at least about 1400 mg / day, at least about 1500 mg / day, at least about 1600 mg / day, at least about 1700 mg / day, at least about 1800 mg / day, at least about 1900 mg / day, or at least about 2000 mg / day). In some embodiments, a phenylalanine intake of about 2000 mg / day, or 1000 mg / day to 1400 mg / day, 1200 mg / day to 1600 mg / day, 1300 mg / day to 1700 mg / day, 1600 mg / day to 2000 mg / day, 1800 mg / day to 2400 mg / day, 2000 mg / day to 3000 mg / day, 3000 mg / day to 4000 mg / day, or 4000 mg / day to 5000 mg / day) is sufficient to reduce a subject's blood phenylalanine concentration to less than 600 micromoles / liter. In some embodiments of any of the methods described herein, the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is at least about 1000 mg / day (e.g., at least about 1100 mg / day, at least about 1200 mg / day, at least about 1300 mg / day, at least about 1400 mg / day, at least about 1500 mg / day, at least about 1600 mg / day, at least about 1700 mg / day, at least about 1800 mg / day, at least about 1900 mg / day, or at least about 2000 mg / day). In some embodiments, a phenylalanine intake of about 2000 mg / day, or 1000 mg / day to 1400 mg / day, 1200 mg / day to 1600 mg / day, 1300 mg / day to 1700 mg / day, 1600 mg / day to 2000 mg / day, 1800 mg / day to 2400 mg / day, 2000 mg / day to 3000 mg / day, 3000 mg / day to 4000 mg / day, or 4000 mg / day to 5000 mg / day) is sufficient to reduce a subject's blood phenylalanine concentration to less than 360 micromoles / liter.In some embodiments of any of the methods described herein, the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is at least about 1000 mg / day (e.g., at least about 1100 mg / day, at least about 1200 mg / day, at least about 1300 mg / day, at least about 1400 mg / day, at least about 1500 mg / day, at least about 1600 mg / day, at least about 1700 mg / day, at least about 1800 mg / day, at least about 1900 mg / day, or at least about 2000 mg / day). In some embodiments, a phenylalanine intake of about 2000 mg / day, or 1000 mg / day to 1400 mg / day, 1200 mg / day to 1600 mg / day, 1300 mg / day to 1700 mg / day, 1600 mg / day to 2000 mg / day, 1800 mg / day to 2400 mg / day, 2000 mg / day to 3000 mg / day, 3000 mg / day to 4000 mg / day, or 4000 mg / day to 5000 mg / day) is sufficient to reduce a subject's blood phenylalanine concentration to less than 120 micromoles / liter. In some embodiments of any of the methods described herein, the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is at least about 1000 mg / day (e.g., at least about 1100 mg / day, at least about 1200 mg / day, at least about 1300 mg / day, at least about 1400 mg / day, at least about 1500 mg / day, at least about 1600 mg / day, at least about 1700 mg / day, at least about 1800 mg / day, at least about 1900 mg / day, or at least A phenylalanine intake of about 2000 mg / day, or 1000 mg / day to 1400 mg / day, 1200 mg / day to 1600 mg / day, 1300 mg / day to 1700 mg / day, 1600 mg / day to 2000 mg / day, 1800 mg / day to 2400 mg / day, 2000 mg / day to 3000 mg / day, 3000 mg / day to 4000 mg / day, or 4000 to 5000 mg / day) is sufficient to lower a subject's blood phenylalanine concentration to between 120 and 360 micromoles / liter.In some embodiments of any of the methods described herein, the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is at least about 1000 mg / day (e.g., at least about 1100 mg / day, at least about 1200 mg / day, at least about 1300 mg / day, at least about 1400 mg / day, at least about 1500 mg / day, at least about 1600 mg / day, at least about 1700 mg / day, at least about 1800 mg / day, at least about 1900 mg / day, or at least A phenylalanine intake of about 2000 mg / day, or 1000 mg / day to 1400 mg / day, 1200 mg / day to 1600 mg / day, 1300 mg / day to 1700 mg / day, 1600 mg / day to 2000 mg / day, 1800 mg / day to 2400 mg / day, 2000 mg / day to 3000 mg / day, 3000 mg / day to 4000 mg / day, or 4000 to 5000 mg / day) is sufficient to lower a subject's blood phenylalanine concentration to between 360 and 600 micromoles / liter.

[0021] In some embodiments of any of the methods described herein, the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is an amount sufficient to reduce a subject's blood phenylalanine concentration to less than 600 micromoles / liter at a natural protein intake of greater than 10 g / day (e.g., greater than 20 g / day, greater than 30 g / day, greater than 40 g / day, greater than 50 g / day, greater than 60 g / day, greater than 70 g / day, greater than 80 g / day, or from about 10 g / day to about 30 g / day, from about 20 g / day to about 40 g / day, from about 30 g / day to about 50 g / day, from about 40 g / day to about 60 g / day, from about 50 g / day to about 70 g / day, from about 60 g / day to about 80 g / day). In some embodiments of any of the methods described herein, the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is an amount sufficient to reduce a subject's blood phenylalanine concentration to less than 360 micromoles / liter at a natural protein intake of greater than 10 g / day (e.g., greater than 20 g / day, greater than 30 g / day, greater than 40 g / day, greater than 50 g / day, greater than 60 g / day, greater than 70 g / day, greater than 80 g / day, or from about 10 g / day to about 30 g / day, from about 20 g / day to about 40 g / day, from about 30 g / day to about 50 g / day, from about 40 g / day to about 60 g / day, from about 50 g / day to about 70 g / day, from about 60 g / day to about 80 g / day). In some embodiments of any of the methods described herein, the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is an amount sufficient to reduce a subject's blood phenylalanine concentration to less than 120 micromoles / liter at a natural protein intake of greater than 10 g / day (e.g., greater than 20 g / day, greater than 30 g / day, greater than 40 g / day, greater than 50 g / day, greater than 60 g / day, greater than 70 g / day, greater than 80 g / day, or from about 10 g / day to about 30 g / day, from about 20 g / day to about 40 g / day, from about 30 g / day to about 50 g / day, from about 40 g / day to about 60 g / day, from about 50 g / day to about 70 g / day, from about 60 g / day to about 80 g / day).In some embodiments of any of the methods described herein, the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is an amount sufficient to reduce a subject's blood phenylalanine concentration to between 120 and 360 micromoles / liter at a natural protein intake of greater than 10 g / day (e.g., greater than 20 g / day, greater than 30 g / day, greater than 40 g / day, greater than 50 g / day, greater than 60 g / day, greater than 70 g / day, greater than 80 g / day, or between about 10 g / day and about 30 g / day, between about 20 g / day and about 40 g / day, between about 30 g / day and about 50 g / day, between about 40 g / day and about 60 g / day, between about 50 g / day and about 70 g / day, between about 60 g / day and about 80 g / day). In some embodiments of any of the methods described herein, the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is an amount sufficient to reduce a subject's blood phenylalanine concentration to between 360 and 600 micromoles / liter at a natural protein intake of greater than 10 g / day (e.g., greater than 20 g / day, greater than 30 g / day, greater than 40 g / day, greater than 50 g / day, greater than 60 g / day, greater than 70 g / day, greater than 80 g / day, or between about 10 g / day and about 30 g / day, between about 20 g / day and about 40 g / day, between about 30 g / day and about 50 g / day, between about 40 g / day and about 60 g / day, between about 50 g / day and about 70 g / day, between about 60 g / day and about 80 g / day).

[0022] In some embodiments of any of the methods described herein, the effective amount is an amount sufficient to reduce the subject's blood phenylalanine concentration by at least 20% (e.g., at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%) from the blood phenylalanine concentration before administration of sepiapterin or a pharma- ceutically acceptable salt thereof.

[0023] In some embodiments of any of the methods described herein, the effective amount is an amount sufficient (e.g., 2.5 mg / kg to 100 mg / kg per administration) to produce a BH4 concentration of at least 50 ng / ml (e.g., at least 60 ng / ml, at least 100 ng / ml, at least 200 ng / ml, at least 400 ng / ml, at least 600 ng / ml, at least 1000 ng / ml, or at least 2000 ng / ml, or from 50 ng / ml to 100 ng / ml, 60 ng / ml to 400 ng / ml, 200 ng / ml to 600 ng / ml, 400 ng / ml to 1000 ng / ml, or 600 ng / ml to 1500 ng / ml) in the subject's plasma within 10 hours of administration of sepiapterin or a pharma- ceutically acceptable salt thereof.

[0024] In some embodiments of any of the methods described herein, the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is about 2.5 mg / kg to about 100 mg / kg (e.g., about 20 mg / kg to about 60 mg / kg, or about 20 mg / kg, about 30 mg / kg, about 40 mg / kg, about 50 mg / kg, or about 60 mg / kg) per administration.

[0025] In some embodiments of any of the methods described herein, an effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is administered with the food. In some embodiments of any of the methods described herein, the effective amount is an amount (e.g., 2.5 mg / kg to 100 mg / kg per dose) sufficient to produce a BH4 concentration of at least 50 ng / ml (e.g., at least 60 ng / ml, at least 100 ng / ml, at least 200 ng / ml, at least 400 ng / ml, at least 600 ng / ml, at least 1000 ng / ml, or at least 2000 ng / ml, or between 50 ng / ml and 100 ng / ml, between 60 ng / ml and 400 ng / ml, between 200 ng / ml and 600 ng / ml, between 400 ng / ml and 1000 ng / ml, or between 600 ng / ml and 1500 ng / ml) in the subject's plasma within 10 hours of administration with the food. In some embodiments, an effective amount includes a dose that is at least 5% (e.g., at least 60 ng / ml, at least 100 ng / ml, at least 200 ng / ml, at least 400 ng / ml, at least 600 ng / ml, at least 1000 ng / ml, or at least 2000 ng / ml, or between 50 ng / ml and 100 ng / ml, between 60 ng / ml and 400 ng / ml, between 200 ng / ml and 600 ng / ml, between 400 ng / ml and 1000 ng / ml, or between 600 ng / ml and 1500 ng / ml) lower than a dose sufficient to provide a maximum BH4 plasma concentration (Cmax) of at least 50 ng / ml in the plasma of a subject within 10 hours of administration of sepiapterin or a pharma- ceutically acceptable salt thereof without food.

[0026] In some embodiments of any of the methods described herein, administration to the subject occurs less than 30 minutes before ingesting the food or after ingesting the food, e.g., immediately before or up to 1 hour after ingesting the food. In some embodiments, administration to the subject is substantially simultaneous with the food. In some embodiments of any of the methods described herein, the food is a high protein food. In some embodiments of any of the methods described herein, the food is a high fat food (e.g., at least 25, 30, 40, or 50% of the calories are from fat). In some embodiments of any of the methods described herein, the food is a high protein and high fat food. In some embodiments, the food is a high calorie food (e.g., the food contains at least 100 calories, e.g., at least 200 calories, at least 300 calories, at least 400 calories, at least 500 calories, e.g., 500-1500 or 800-1000 calories). In some embodiments of any of the methods described herein, the food is a meal, e.g., breakfast, lunch, or dinner.

[0027] In some embodiments of any of the methods described herein, administration with food (e.g., less than 30 minutes before ingesting food or after ingesting food, e.g., between just before and 1 hour after ingesting food) results in an increase in the Cmax of BH4 (e.g., at least 5%, at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 100%, at least 110%, at least 120%, at least 130%, at least 140%, or at least 150%) compared to administration without food (e.g., more than 2 hours after ingesting food and up to 30 minutes before ingesting food).

[0028] In some embodiments of any of the methods described herein, administration with food (e.g., less than 30 minutes before ingesting food, or after ingesting food, e.g., immediately before ingesting food and up to 1 hour after ingesting food) may increase the extent of BH4 production and resulting plasma exposure (AUC 0-last ) (e.g., at least 5%, at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 100%, at least 110%, at least 120%, at least 130%, at least 140%, or at least 150%).

[0029] In some embodiments of any of the methods described herein, the sepiapterin or a pharma- ceutically acceptable salt thereof is provided in a composition separate from the food ingested (e.g., the sepiapterin is not incorporated into a food product). In some embodiments of any of the methods described herein, the ingestion of the food is performed prior to administration of the sepiapterin or a pharma- ceutically acceptable salt thereof (e.g., the ingestion of the food is performed between 1 hour prior to and just prior to administration of the sepiapterin). In some embodiments of any of the methods described herein, the ingestion of the food is performed after administration of the sepiapterin or a pharma- ceutically acceptable salt thereof (e.g., the ingestion of the food is performed between immediately after administration and 30 minutes after administration).

[0030] In some embodiments of any of the methods described herein, the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is administered in two equal doses (e.g., two doses at different times of the day). In some embodiments of any of the methods described herein, the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is administered in two 60 mg / kg doses (e.g., one 60 mg / kg dose in the morning and one 60 mg / kg dose in the evening). In some embodiments of any of the methods described herein, the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is administered in two 40 mg / kg doses (e.g., one 40 mg / kg dose in the morning and one 40 mg / kg dose in the evening). In some embodiments of any of the methods described herein, the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is administered in two 30 mg / kg doses (e.g., one 30 mg / kg dose in the morning and one 30 mg / kg dose in the evening). In some embodiments of any of the methods described herein, the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is administered in two 20 mg / kg doses (e.g., one 20 mg / kg dose in the morning and one 20 mg / kg dose in the evening). In some embodiments of any of the methods described herein, the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is administered in two 10 mg / kg doses (e.g., one 10 mg / kg dose in the morning and one 10 mg / kg dose in the evening).

[0031] In some embodiments of any of the methods described herein, the risk of an adverse event (e.g., headache, rhinorrhea, sore throat, diarrhea, vomiting, cough, and / or stuffy nose) is reduced compared to a subject administered at least 10 mg / kg (e.g., at least 15 mg / kg, at least 20 mg / kg) of sapropterin or a pharma- ceutically acceptable salt thereof (e.g., sapropterin dihydrochloride). In some embodiments of any of the methods described herein, the risk of an adverse event (e.g., headache, rhinorrhea, sore throat, diarrhea, vomiting, cough, and / or stuffy nose) is reduced compared to a subject administered at least 20 mg / kg of sapropterin or a pharma- ceutically acceptable salt thereof (e.g., sapropterin dihydrochloride).

[0032] In some embodiments of any of the methods described herein, the subject is a child (e.g., the subject is under 18 years old, under 17 years old, under 16 years old, under 15 years old, under 14 years old, under 13 years old, under 12 years old, under 11 years old, under 10 years old, under 9 years old, under 8 years old, under 7 years old, under 6 years old, under 5 years old, under 4 years old, under 3 years old, under 2 years old, under 1 year old). In some embodiments of any of the methods described herein, the subject is an adult (e.g., the subject is over 18 years old).

[0033] In some embodiments of any of the methods described herein, the subject has been diagnosed with tetrahydrobiopterin-responsive phenylketonuria. In some embodiments of any of the methods described herein, the subject has been diagnosed with hyperphenylalaninemia due to sepiapterin-responsive PKU.

[0034] In embodiments of any of the methods described herein, the subject is on a phenylalanine restricted diet (e.g., the subject is on a diet that includes a milk substitute or blend, such as Phenyl-Free 2, and measured amounts of fruits, vegetables, breads, pastas, and cereals). In some embodiments of any of the methods described herein, the subject has a median phenylalanine intake of less than 3000 mg / day (e.g., less than 2500 mg / day, less than 2000 mg / day, less than 1500 mg / day, less than 1000 mg / day, less than 500 mg / day).

[0035] In some embodiments of any of the methods described herein, the subject is not on a phenylalanine-restricted diet (e.g., the subject has a median phenylalanine intake of greater than 1000 mg / day (e.g., greater than 1500 mg / day, greater than 2000 mg / day, greater than 2500 mg / day, greater than 3000 mg / day)).

[0036] In some embodiments of any of the methods described herein, less than 25% (e.g., less than 20%, less than 15%, less than 10%, or less than 5%) of the protein in the subject's diet is native protein. In some embodiments of any of the methods described herein, more than 25% (e.g., more than 30%, more than 40%, more than 50%, more than 60%, more than 70%, more than 80%, more than 90%, more than 95%, more than 99%) of the protein in the subject's diet is native protein. In some embodiments of any of the methods described herein, the subject has a native protein intake of less than 10 g / day (e.g., less than 5 g / day, or between about 5 g / day and about 10 g / day). In some embodiments of any of the methods described herein, the subject has a natural protein intake of greater than 10 g / day (e.g., greater than 20 g / day, greater than 30 g / day, greater than 40 g / day, greater than 50 g / day, greater than 60 g / day, greater than 70 g / day, greater than 80 g / day, or about 10 g / day to about 30 g / day, about 20 g / day to about 40 g / day, about 30 g / day to about 50 g / day, about 40 g / day to about 60 g / day, about 50 g / day to about 70 g / day, about 60 g / day to about 80 g / day).

[0037] In some embodiments of any of the methods described herein, the sepiapterin or a pharma- ceutically acceptable salt thereof is formulated as an oral powder for suspension. In some embodiments of any of the methods described herein, the sepiapterin or a pharma- ceutically acceptable salt thereof is administered as a suspension in a flavored suspension vehicle (e.g., MEDISCA® Oral Mix).

[0038] In some embodiments of any of the methods described herein, the administering step results in an increase in neurocognitive function in the subject (e.g., increased executive function, reduced anxiety, reduced Attention Deficit / Hyperactivity Disorder symptoms, and / or reduced instances of brain fog). In some embodiments, the administering step results in an increase in neurocognitive function in the subject (e.g., increased executive function, reduced anxiety, reduced Attention Deficit / Hyperactivity Disorder symptoms, and / or reduced instances of brain fog). In some embodiments, the administering step results in an increase in neurocognitive function in the subject (e.g., increased executive function, reduced anxiety, reduced Attention Deficit / Hyperactivity Disorder symptoms, and / or reduced instances of brain fog). In some embodiments, the administering step results in an increase in neurocognitive function in the subject (e.g., increased executive function, reduced anxiety, reduced Attention Deficit / Hyperactivity Disorder symptoms, and / or reduced instances of brain fog). In some embodiments, the administering step results in an increase in neurocognitive function in the subject (e.g., increased executive function, reduced anxiety, reduced instances of anxiety The administering step results in an increase in executive function (e.g., as measured by an assessment such as the Neurocognitive Assessment (KBNA), Kaufman Short Neuropsychological Assessment, Mental Clutter Scale, Paced Auditory Serial Addition Test (PASAT), Phenylketonuria-Quality of Life (PKU-QOL), Rey-Osterlie's Complex Figures, Rough Figural Fluency Test, Stroop Task, Task of Executive Control, Test of Variables of Attention (TOVA), Tower of London Test, Trail Making Test (TMT) or Trails A & B, Wisconsin Card Sorting Test (WCST), or Symbol Digit Modalities Test). In some embodiments of any of the methods described herein, the administering step results in an increase in executive function (e.g., as measured by a Cambridge Neuropsychological Test Automated Battery (CANTAB) assessment (e.g., by measuring reaction speed, spatial span, spatial working memory, rapid digital information processing, sustained attention, and / or a stop-signal task)).In some embodiments of any of the methods described herein, the administering step results in improved attention and / or mood (e.g., as measured by the ADHD-RS 5 scale (or an inattention rating thereof) and / or the Profile of Mood States (POMS) scale).

[0039] In some embodiments of any of the methods described herein, the effective amount is an amount sufficient to cause an increase in neurocognitive function in the subject (e.g., an increase in executive function, a decrease in anxiety, a decrease in Attention Deficit / Hyperactivity Disorder symptoms, and / or a decrease in instances of brain fog). In some embodiments, the effective amount is an amount sufficient to cause an increase in neurocognitive function in the subject (e.g., an increase in executive function, a decrease in anxiety, a decrease in Attention Deficit / Hyperactivity Disorder symptoms, and / or a decrease in instances of brain fog). In some embodiments, the effective amount is an amount sufficient to cause an increase in neurocognitive function in the subject (e.g., an increase in executive function, a decrease in anxiety, a decrease in Attention Deficit / Hyperactivity Disorder symptoms, and / or a decrease in instances of brain fog). In some embodiments, the effective amount is an amount sufficient to cause an increase in neurocognitive function in the subject (e.g., an increase in executive function, a decrease in anxiety, a decrease in Attention Deficit / Hyperactivity Disorder symptoms, and / or a decrease in instances of brain fog). An effective amount is an amount sufficient to result in an increase in executive function (e.g., as measured by an assessment such as the Neurocognitive Assessment (KBNA), Kaufman Short Neuropsychological Assessment, Mental Clutter Scale, Paced Auditory Serial Addition Test (PASAT), Phenylketonuria-Quality of Life (PKU-QOL), Rey-Osterlie's Complex Figures, Rough Figural Fluency Test, Stroop Task, Task of Executive Control, Test of Variables of Attention (TOVA), Tower of London Test, Trail Making Test (TMT) or Trails A & B, Wisconsin Card Sorting Test (WCST), or Symbol Digit Modalities Test). In some embodiments, an effective amount is an amount sufficient to result in an increase in executive function (e.g., as measured by a Cambridge Neuropsychological Test Automated Battery (CANTAB) assessment (e.g., by measuring reaction speed, spatial span, spatial working memory, rapid digital information processing, sustained attention, and / or a stop-signal task)).In some embodiments of any of the methods described herein, an effective amount is an amount sufficient to produce an improvement in attention and / or mood (e.g., as measured by the ADHD-RS 5 scale (or an inattention rating thereof) and / or the Profile of Mood States (POMS) scale).

[0040] In some embodiments of any of the methods described herein, the administering step results in an increase in sleep quality and / or a decrease in symptoms associated with sleep deprivation. In some embodiments of any of the methods described herein, the effective amount is an amount sufficient to result in an increase in sleep quality and / or a decrease in symptoms associated with sleep deprivation.

[0041] definition In this application, unless otherwise clear from the context, (i) the term "a" may be understood to mean "at least one," (ii) the term "or" may be understood to mean "and / or," (iii) the terms "comprising" and "including" may be understood to include the listed components or steps, whether presented by themselves or with one or more additional components or steps, and (iv) the term "about" may be understood to allow for standard variations as understood by one of ordinary skill in the art, and (v) when ranges are provided, the endpoints are included.

[0042] Descriptions of compounds, compositions, formulations, and methods of treatment described herein should be understood to include the embodiments "comprising," "consisting," and "consisting essentially of." In some embodiments, for all compositions described herein and all methods of using compositions described herein, the compositions can include or "consist essentially of" the recited components or steps. When a composition is described as "consisting essentially of" a recited component, the composition includes the recited components and may include other components that do not substantially affect the condition being treated, but does not include any other components that substantially affect the condition being treated other than the components specifically recited; or, if the composition does include additional components that substantially affect the condition being treated other than the recited components, the composition does not include the additional components in a concentration or amount sufficient to substantially affect the condition being treated. When a method is described as "consisting essentially of" a recited step, the method includes the recited steps and may include other steps that do not substantially affect the condition being treated, but does not include any other steps that substantially affect the condition being treated other than the steps specifically recited. As a specific, non-limiting example, when a composition is described as "consisting essentially of" a certain component, the composition can further include any amount of pharma- ceutically acceptable carriers, vehicles, or diluents, as well as other such components that do not substantially affect the condition being treated.

[0043] Unless otherwise clear from the context, all references to sepiapterin contained herein refer to sepiapterin or to a pharma- ceutically acceptable salt of sepiapterin.

[0044] The term "about" as used herein refers to a value that is in the range of ±10% of the value following the term "about". As used herein, referring to a value or parameter as "about" includes (and describes) variations directed to the value or parameter itself. For example, a statement that refers to "about X" includes the statement of "X". As used herein, the term "administration" refers to administration of a composition (e.g., a compound described herein or a preparation comprising same) to a subject or system. Administration to an animal subject (e.g., administration to a human) may be by any suitable route. For example, in some embodiments, administration includes bronchial (including bronchial instillation), buccal, enteral, interdermal, intra-arterial, intradermal, intragastric, intramedullary, intramuscular, intranasal, intraperitoneal, intrathecal, intravenous, intraventricular, mucosal, nasal, oral, rectal, subcutaneous, sublingual, topical, intratracheal (including intratracheal instillation), transdermal, intravaginal, and intravitreal.

[0045] "Measuring the level of phenylalanine" means detecting phenylalanine by methods known in the art, either directly or indirectly. "Measuring directly" means performing a process (e.g., performing an assay or test on a sample, or "analyzing a sample" as that term is defined herein) to obtain a physical entity or value. "Measuring indirectly" refers to receiving a physical entity or value from another party or source (e.g., a third party laboratory that directly obtains the physical entity or value). Methods for measuring phenylalanine levels typically include, but are not limited to, fluorescent assays in which a by-product of the reaction of phenylalanine reacts with a fluorescent probe.

[0046] An "effective amount" of a compound may vary depending on factors such as the individual's condition, age, sex, and weight, as well as the ability of the compound to elicit a desired response. A therapeutically effective amount encompasses an amount in which any toxic or detrimental effects of the compound are outweighed by the therapeutically beneficial effects. An effective amount also encompasses an amount sufficient to provide a benefit, e.g., a clinical benefit.

[0047] As used herein, the term "failed to respond to prior therapy" refers to a subject whose condition has remained stable, worsened, or improved less than the accepted standard for a particular therapy despite treatment with that therapy (e.g., as described in the FDA-approved labeling for that therapy).

[0048] The term "food" as used herein refers to solid food having sufficient bulk and fat content so as not to be rapidly dissolved and absorbed in the stomach. For example, meals such as breakfast, lunch, or dinner. The term "with food" as used herein refers to administration of the composition from about 30 minutes before eating (e.g., food) to about 2 hours after eating. The terms "without food," "fasting," or "fasting" further refer to the state of not consuming solid food for at least about 2 hours, up to about 30 minutes before consuming solid food.

[0049] The term "hyperphenylalaninemia" as used herein refers to a medical condition in which a subject has elevated levels of phenylalanine in the blood compared to a healthy subject. For example, in some embodiments, a subject is considered to have hyperphenylalaninemia if the concentration of phenylalanine in the subject's blood is greater than 60 micromoles / liter. In some embodiments, a subject is considered to have hyperphenylalaninemia if the concentration of phenylalanine in the subject's blood is greater than 120 micromoles / liter (e.g., from 120 micromoles / liter to 2400 micromoles / liter).

[0050] "Level" refers to the level of phenylalanine as compared to a reference. The reference can be any useful reference as defined herein, for example, the level in the subject prior to administration, i.e., the baseline level. A "decreased level" or "increased level" of phenylalanine refers to a decrease or increase in phenylalanine levels as compared to a reference (e.g., about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, about 100%, about 150%, about 200%, about 300%, about 400%, about 500%, or greater decrease or increase); as compared to a reference. "A decrease or increase of greater than about 10%, about 15%, about 20%, about 50%, about 75%, about 100%, or about 200% of the amount of the antibody; a decrease or increase of about 0.01-fold, about 0.02-fold, about 0.1-fold, about 0.3-fold, about 0.5-fold, about 0.8-fold, or less; or an increase of about 1.2-fold, about 1.4-fold, about 1.5-fold, about 1.8-fold, about 2-fold, about 3.0-fold, about 3.5-fold, about 4.5-fold, about 5.0-fold, about 10-fold, about 15-fold, about 20-fold, about 30-fold, about 40-fold, about 50-fold, about 100-fold, about 1000-fold, or more). Phenylalanine levels may be expressed as mass / vol (e.g., g / dL, mg / mL, μg / mL, ng / mL), concentration / vol (e.g., nanomoles / liter, micromoles / liter, micromoles / milliliter), or as a percentage of total amino acids in the sample.

[0051] By "native protein" is meant a protein derived from a natural source (eg, an animal, plant, or fungus) that contains Phe.

[0052] The term "pharmaceutical composition" as used herein refers to a composition comprising a compound described herein formulated with a pharma- ceutical acceptable excipient. Pharmaceutical compositions can be formulated, for example, for oral administration (e.g., tablet, capsule, caplet, gel cap, suspension, solution, or syrup) of unit dosage form, for topical administration (e.g., as a cream, gel, lotion, or ointment), for intravenous administration (e.g., as a sterile solution in a solvent system that is particulate-free and suitable for intravenous use), or in any other pharma- ceutical acceptable formulation.

[0053] The term "pharmaceutical acceptable salt" as used herein means any salt that is suitable for use in contact with human and animal tissues without undue toxicity, irritation, or allergic reaction within the scope of sound medical judgment, and is commensurate with a reasonable benefit / risk ratio. Pharmaceutically acceptable salts are well known in the art. For example, pharmaceutical acceptable salts are described in Berge et al., J. Pharmaceutical Sciences 66:1-19, 1977, and Pharmaceutical Salts: Properties, Selection, and Use, (Eds. PH Stahl and CG Wermuth), Wiley-VCH, 2008. Salts can be prepared in situ during the final isolation and purification of the compounds described herein, or can be prepared separately by reacting the free base group with a suitable organic acid.

[0054] In many cases, compounds are prepared or used as pharma- ceutically acceptable salts prepared as addition products of pharma-ceutically acceptable acids.Suitable pharma-ceutically acceptable acids and methods for preparing suitable salts are well known in the art.Salts may be prepared from pharma-ceutically acceptable non-toxic acids, including inorganic and organic acids.

[0055] Representative acid addition salts include acetate, adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, besylate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, fumarate, gentisate, glucoheptonate, glycerophosphate, glycolate, hemisulfate, heptonate, hexanate, hydrobromide, hydrochloride, hydroiodide, 2-amino-3-propanesulfate ... -hydroxy-ethanesulfonate, 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, thalate, thiocyanate, toluenesulfonate, undecanoate, and valerate salts.

[0056] "Reduction in blood phenylalanine concentration" means that the level of phenylalanine is reduced in the blood and / or plasma of a subject. The level of phenylalanine can be measured using any method known in the art, for example, the concentration of phenylalanine in a subject's blood can be measured by dried blood spot or plasma analysis.

[0057] "Sepiapterin-responsive phenylketonuria" means phenylketonuria in which a subject exhibits a decrease in blood phenylalanine from baseline after up to one month of administration of sepiapterin or a pharma- ceutically acceptable salt thereof.

[0058] As used herein, the term "substantially free" refers to a qualitative condition indicating the absence of all or nearly all extent or degree of a compound or type of compound of interest. One of ordinary skill in the art of biology will understand that biological and chemical phenomena are rarely determined to be zero with certainty, for example, due to inherent error in any measurement. Thus, the term "substantially free" is used herein to capture the potential lack of completeness inherent in many biological and chemical measurements.

[0059] The term "subject" or "patient" as used herein refers to any organism to which a compound or composition according to the invention can be administered, e.g., for experimental, diagnostic, prophylactic, and / or therapeutic purposes. Exemplary subjects include any animal (e.g., mammals such as mice, rats, rabbits, non-human primates, and humans). A subject may be a human or animal in need of treatment, currently undergoing treatment, or about to undergo treatment, or who is under the care of a trained professional for a particular disease or condition.

[0060] The terms "treat", "treated" or "treating" as used herein refer to both therapeutic treatment and prophylactic or suppressive measures, the purpose of which is to prevent or slow (alleviate) an undesirable physiological condition, disorder, or disease, or to obtain a beneficial or desired clinical outcome. Beneficial or desired clinical outcomes include, but are not limited to, alleviation of symptoms, reduction in the extent of a condition, disorder, or disease, a stable (i.e., not worsening) appearance of a condition, disorder, or disease, a delay in the onset or progression of a condition, disorder, or disease, an improvement (partial or total) in a condition, disorder, or disease state or remission, whether detectable or undetectable, an improvement in at least one measurable physical parameter, which need not necessarily be discernible by the subject, or an improvement or enhancement of a condition, disorder, or disease. Treatment includes eliciting a clinically significant response without an excessive level of side effects. Treatment also includes extending survival compared to the expected survival in the absence of treatment.

[0061] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by those of ordinary skill in the art to which this invention belongs. Methods and materials are described herein for use in the disclosure of this specification, and other suitable methods and materials known in the art can also be used. The materials, methods, and examples are illustrative only and are not intended to be limiting. All publications, patent applications, patents, sequences, database entries, and other references mentioned herein are incorporated by reference in their entirety. In case of conflict, the present specification, including definitions, will control.

[0062] The details of one or more embodiments of the invention are set forth in the description below. Other features, objects, and advantages of the invention will become apparent from the description and the claims. [Brief description of the drawings]

[0063] [Figure 1] FIG. 1 is a graph illustrating the mean absolute reduction in plasma phenylalanine concentrations following treatment with low dose PTC923 and PTC923. [Diagram 2] FIG. 2 is a chart illustrating the six treatment sequences into which patients were randomized. [Diagram 3] FIG. 3 is a graph showing the mean absolute change in blood phenylalanine concentrations from baseline following treatment with sapropterin, low dose PTC923, and PTC923. [Figure 4] FIG. 4 is a graph showing the mean absolute change from baseline in blood phenylalanine concentrations after 3 days of treatment with sapropterin, low dose PTC923, and PTC923. [Diagram 5] FIG. 5 is a graph showing the absolute change in blood phenylalanine concentration for each subject following treatment with sapropterin and PTC923. [Figure 6] FIG. 6 is a graph showing the mean absolute change in blood phenylalanine concentrations in classical PKU subjects after treatment with sapropterin and PTC923. [Figure 7] FIG. 7 is a graph showing the mean absolute change in blood phenylalanine concentrations in the sensitivity analysis population following treatment with sapropterin, low dose PTC923, and PTC923. [Figure 8] FIG. 8 is a graph showing the increased percentage of responders to treatment with PTC923 compared to treatment with sapropterin. Detailed Description

[0064] The inventors have surprisingly found that sepiapterin is effective in reducing blood phenylalanine levels. In particular, the inventors have found that sepiapterin is effective in reducing blood phenylalanine levels to levels similar to those of healthy individuals, and / or in subjects who have not responded to prior treatment with sapropterin dihydrochloride and / or pegvalase-pqpz. Thus, the present invention features a method for reducing blood phenylalanine levels in a subject by administering sepiapterin or a pharma- ceutically acceptable salt thereof.

[0065] Sepiapterin Sepiapterin enters the cell and is converted to 7,8-dihydrobiopterin by sepiapterin reductase, which is then reduced to BH4 by dihydrofolate reductase.

[0066] Sepiapterin has the following structure: JPEG0007661315000001.jpg3765 Sepiapterin

[0067] Sepiapterin or a pharma- ceutically acceptable salt thereof can be incorporated into a pharmaceutical composition. In some embodiments, the pharmaceutical composition of the present invention contains sepiapterin or a salt thereof in an amount of 20-30% by weight, for example, about 20%, 22%, 25%, 27%, or 30%. In some embodiments, the pharmaceutical composition contains sepiapterin in an amount of more than 20% by weight, for example, more than 25%, more than 30%, more than 40%, more than 50%, more than 60%, more than 70%, more than 80%, or more than 90% by weight. In some embodiments, the pharmaceutical composition contains sepiapterin in an amount of less than 20% by weight, for example, less than 20%, less than 15%, less than 10%, or less than 5% by weight.

[0068] In some embodiments, the invention features a pharmaceutical composition comprising sepiapterin or a salt thereof and less than 10% by weight of an antioxidant (e.g., about 9%, 7%, 5%, 3%, 1%, 0.5%, 0.25%, 0.1%, or no antioxidant). The antioxidant may be ascorbic acid. In some embodiments, the ratio of sepiapterin or a pharma- ceutically acceptable salt thereof to the antioxidant is 1:1 by weight, or greater than 1:1, e.g., 2:1, 5:1, 7:1, or 10:1 by weight. The pharmaceutical composition may comprise 20-30% by weight of sepiapterin or a pharma- ceutically acceptable salt thereof, e.g., about 20%, 22%, 25%, 27%, or 30% by weight. The pharmaceutical composition may further comprise a dispersing agent, e.g., croscarmellose sodium. The pharmaceutical composition may comprise 0.1-1.5% of the dispersing agent by total weight, for example, 0.1%, 0.5%, 1% or 1.5%. In some embodiments, the pharmaceutical composition comprises at least one anti-caking agent, for example, colloidal silicon dioxide or microcrystalline cellulose. The pharmaceutical composition may comprise 65-75% of the anti-caking agent by total weight, for example, about 65%, 67%, 70%, 73%, or 75%. In some embodiments, the pharmaceutical composition comprises both colloidal silicon dioxide and microcrystalline cellulose. In some embodiments, the pharmaceutical composition comprises 60-65% of microcrystalline cellulose by total weight, and 5-7% of colloidal silicon dioxide by total weight. In some embodiments, the sepiapterin crystalline form is formulated as particles less than 140 μm (e.g., about 120 μm, 110 μm, 100 μm, 90 μm, 80 μm, 70 μm, 60 μm, 50 μm, 40 μm, 30 μm, 20 μm, 10 μm, or 5 μm). In some embodiments, the pharmaceutical composition contains less than 1% by total weight of impurities such as lactoylpterin, e.g., less than 0.9%, less than 0.8%, less than 0.7%, less than 0.6%, less than 0.5%, less than 0.4%, less than 0.3%, or less than 0.2%.

[0069] In some embodiments, the sepiapterin is a salt of sepiapterin, for example, a salt with sulfuric acid, p-toluenesulfonic acid, methanesulfonic acid, benzenesulfonic acid, malonic acid, tartaric acid (e.g., L-tartaric acid), phosphoric acid, gentisic acid, fumaric acid, glycolic acid, acetic acid, or nicotinic acid.

[0070] In some embodiments, sepiapterin or a pharma- ceutically acceptable salt thereof is in a crystalline form. The crystalline form of crystalline sepiapterin free base or a salt of sepiapterin may occur as an anhydrate (e.g., without any bound water or solvent or hydration or solvation), or as a hydrate, partial hydrate (e.g., hemihydrate, sesquihydrate, etc.), dihydrate, trihydrate, etc., where the crystalline form binds water of hydration or solvent molecules associated with the sepiapterin crystalline form or salt thereof. In one embodiment, crystalline sepiapterin occurs as a monohydrate or hemihydrate.

[0071] In some embodiments, sepioapterin is present in a crystalline form, for example, as described in WO 2018 / 102314 and WO 2018 / 102315, which crystalline forms are incorporated by reference herein in their entireties.

[0072] In some embodiments, the sepiapterin crystalline form is characterized by a powder X-ray diffraction pattern obtained by irradiation with CuKα X-rays having peaks expressed as 2θ of at least 9.7°±0.5, e.g., 9.7°±0.2, 10.2°±0.5, e.g., 10.2°±0.2, and 11.3°±0.5, e.g., 11.3°±0.2. In another embodiment, the sepiapterin crystalline form is characterized by a powder X-ray diffraction pattern obtained by irradiation with CuKα X-rays having peaks expressed in degrees 2θ of at least 9.7°±0.5, such as 9.7°±0.2, 10.2°±0.5, such as 10.2°±0.2, 11.3°±0.5, such as 11.3°±0.2, 14.0°±0.5, such as 14.0°±0.2, 14.6°±0.5, such as 14.6°±0.2, 19.9°±0.5, such as 19.9°±0.2, 22.2°±0.5, such as 22.2°±0.2, 25.3°±0.5, such as 25.3°±0.2, and 32.4°±0.5, such as 32.4°±0.2. In an essentially pure form of this crystalline form, peaks may be observed at refractive angles 2θ as shown in Table 1. Alternatively or additionally, this crystalline form is characterized by a DSC curve exhibiting two endotherms at 71.6° C. and 233.4° C.

[0073] [Table 1]

[0074] In some embodiments, the crystalline form of sepiapterin has at least one peak (e.g., 1, 2, or 3 peaks) at a diffraction angle 2θ (°) of 8.4°±0.5, e.g., 8.4°±0.2, 16.9°±0.5, e.g., 16.9°±0.2, or 25.4°±0.5, e.g., 25.4°±0.2, as measured by X-ray diffraction using CuKα X-ray irradiation or calculated from X-ray diffraction. In some embodiments, the crystalline form of sepiapterin has at least one peak (e.g., 1, 2, 3, 4 or 5 peaks) at diffraction angles 2θ (°) of 8.4°±0.5, e.g., 8.4°±0.2, 14.9°±0.5, e.g., 14.9°±0.2, 16.9°±0.5, e.g., 16.9°±0.2, 25.4°±0.5, e.g., 25.4°±0.2, and 34.1°±0.5, e.g., 34.1°±0.2, as measured by X-ray diffraction using CuKα X-ray irradiation or calculated from X-ray diffraction. In an essentially pure material of this crystalline form, peaks may be observed at refractive angles 2θ as shown in Table 2. Alternatively or additionally, this crystalline form is characterized by a DSC curve showing a melting event at 195.2°C.

[0075] [Table 2]

[0076] In some embodiments, the crystalline form of sepiapterin has at least one peak (e.g., 1, 2, or 3 peaks) at a diffraction angle 2θ (°) of 5.7°±0.5, e.g., 5.7°±0.2, 7.8°±0.5, e.g., 7.8°±0.2, or 25.4°±0.5, e.g., 25.4°±0.2, as measured by X-ray diffraction using CuKα X-ray irradiation or calculated from X-ray diffraction. In some embodiments, the crystalline form of sepiapterin has a crystallinity of 5.7°±0.5, e.g., 5.7°±0.2, 7.8°±0.5, e.g., 7.8°±0.2, 9.1°±0.5, e.g., 9.1°±0.2, 11.5°±0.5, e.g., 11.5°±0.2, 15.3°±0.5, For example, it has at least one peak (e.g., 1, 2, 3, 4, 5, 6, 7, 8, or 9 peaks) at diffraction angles 2θ (°) of 15.3°±0.2, 16.0°±0.5, e.g., 16.0°±0.2, 20.1°±0.5, e.g., 20.1°±0.2, 25.4°±0.5, e.g., 25.4°±0.2, and 26.6°±0.5, e.g., 26.6°±0.2. In an essentially pure material of this crystalline form, peaks may be observed at refractive angles 2θ as shown in Table 3. Alternatively or additionally, this crystalline form is characterized by a DSC curve showing five endothermic peaks at 58.3°C, 101.8°C, 129.8°C, 156.5°C, and 168.3°C.

[0077] [Table 3]

[0078] In some embodiments, the crystalline form of sepiapterin has at least one peak (e.g., 1, 2, or 3 peaks) at a diffraction angle 2θ (°) of 8.9°±0.5, e.g., 8.9°±0.2, 10.3°±0.5, e.g., 10.3°±0.2, or 26.0°±0.5, e.g., 26.0°±0.2, as measured by X-ray diffraction using CuKα X-ray irradiation or calculated from X-ray diffraction. In some embodiments, the crystalline form of sepiapterin has a crystallinity of 8.9°±0.5, such as 8.9°±0.2, 10.3°±0.5, such as 10.3°±0.2, 10.9°±0.5, such as 10.9°±0.2, 17.8°±0.5, such as 17.8°±0.2, 24.9°±0. 5, e.g., 24.9°±0.2, 26.0°±0.5, e.g., 26.0°±0.2, 26.7°±0.5, e.g., 26.7°±0.2, 26.8°±0.5, e.g., 26.8°±0.2, and 28.3°±0.5, e.g., 28.3°±0.2, at least one peak (e.g., 1, 2, 3, 4, 5, 6, 7, 8, or 9 peaks) at diffraction angles 2θ (°). In an essentially pure material of this crystalline form, peaks may be observed at diffraction angles 2θ as shown in Table 4. Alternatively or additionally, this crystalline form is characterized by a DSC curve showing three endotherms at 42.7°C, 66.3°C, and 232.9°C.

[0079] [Table 4]

[0080] In some embodiments, the crystalline form of sepiapterin has at least one peak (e.g., 1, 2, or 3 peaks) at a diffraction angle 2θ (°) of 4.7°±0.5, e.g., 4.7°±0.2, 7.4°±0.5, e.g., 7.4°±0.2, or 26.2°±0.5, e.g., 26.2°±0.2, as measured by X-ray diffraction using CuKα X-ray irradiation or calculated from X-ray diffraction. In some embodiments, the crystalline form of sepiapterin has at least one peak (e.g., 1, 2, 3, 4, 5, 6, 7, or 8 peaks) at a diffraction angle 2θ (°) of 4.7°±0.5, such as 4.7°±0.2, 7.4°±0.5, such as 7.4°±0.2, 9.5°±0.5, such as 9.5°±0.2, 11.3°±0.5, such as 11.3°±0.2, 15.6°±0.5, such as 15.6°±0.2, 16.4°±0.5, such as 16.4°±0.2, 26.2°±0.5, such as 26.2°±0.2, or 27.2°±0.5, such as 27.2°±0.2, as measured by X-ray diffraction using CuKα X-ray irradiation or calculated from X-ray diffraction. In essentially pure material of this crystalline form, peaks may be observed at refractive angles 2θ as shown in Table 5. Alternatively or additionally, this crystalline form is characterized by a DSC curve exhibiting endothermic peaks at 82.8°C and 179.8°C.

[0081] [Table 5]

[0082] In some embodiments, the crystalline form of sepiapterin has at least one peak (e.g., 1, 2, 3, 4, 5 or 6 peaks) at a diffraction angle 2θ (°) of 6.0°±0.5, 6.0°±0.210.6°±0.5, 10.6°±0.2, 12.1°±0.5, such as 12.1°±0.2, 15.9°±0.5, such as 15.9°±0.2, 20.9°±0.5, such as 20.9°±0.2, or 24.6°±0.5, such as 24.6°±0.2, as measured by X-ray diffraction using CuKα X-ray irradiation or calculated from X-ray diffraction. In some embodiments, the crystalline form of sepiapterin has an angle of 6.0°±0.5, such as 6.0°±0.2, 10.6°±0.5, such as 10.6°±0.2, 12.1°±0.5, such as 12.1°±0.2, 15.9°±0.5, such as 15.9°±0.2, 18.1°±0.5, such as 18.1°±0.2, 20.9°±0.5, such as 20.9°±0.2, 22.1°±0.5, such as At least one peak (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, or 13 peaks) at a diffraction angle 2θ (°) of 22.1°±0.2, 24.6°±0.5, such as 24.6°±0.2, 26.1°±0.5, such as 26.1°±0.2, 28.1°±0.5, such as 28.1°±0.2, 28.9°±0.5, such as 28.9°±0.2, 32.1°±0.5, such as 32.1°±0.2, or 37.0°±0.5, such as 37.0°±0.2. In an essentially pure form of this crystalline form, peaks may be observed at refractive angles 2θ as shown in Table 6. Alternatively or additionally, this crystalline form is characterized by a DSC curve exhibiting two endothermic peaks at 112.9°C and 195.8°C.

[0083] [Table 6]

[0084] In some embodiments, the crystalline form of sepiapterin has at least one peak (e.g., 1, 2, or 3 peaks) at a diffraction angle 2θ (°) of 10.0°±0.5, e.g., 10.0°±0.2, 10.6°±0.5, e.g., 10.6°±0.2, or 25.7°±0.5, e.g., 25.7°±0.2, as measured by X-ray diffraction using CuKα X-ray irradiation or calculated from X-ray diffraction. In some embodiments, the crystalline form of sepiapterin has a crystallinity of 10.0°±0.5, such as 10.0°±0.2, 10.6°±0.5, such as 10.6°±0.2, 11.2°±0.5, such as 11.2°±0.2, 15.3°±0.5, such as 15.3°±0.2, 15.9°±0.5, such as 15.9°± 0.2, 22.8°±0.5, such as 22.8°±0.2, 24.4°±0.5, such as 24.4°±0.2, 25.0°±0.5, such as 25.0°±0.2, 25.7°±0.5, such as 25.7°±0.2, or 26.6°±0.5, such as 26.6°±0.2, have at least one peak (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 peaks) at a diffraction angle 2θ (°). In an essentially pure material of this crystalline form, peaks may be observed at refractive angles 2θ as shown in Table 7.

[0085] [Table 7]

[0086] In some embodiments, the crystalline form of sepiapterin hydrochloride has at least one peak (e.g., 1, 2, or 3 peaks) at a diffraction angle 2θ (°) of 7.8°±0.5, e.g., 7.8°±0.2, 12.9°±0.5, e.g., 12.9°±0.2, or 26.2°±0.5, e.g., 26.2°±0.2, as measured by X-ray diffraction using CuKα X-ray irradiation or calculated from X-ray diffraction. In some embodiments, the most intense peak in the X-ray diffraction diagram of the crystalline form of sepiapterin hydrochloride is observed at a refractive angle 2θ of 7.8°±0.5, e.g., 7.8°±0.2. The essentially pure material of the crystal of sepiapterin hydrochloride may have peaks observed at refractive angles 2θ as shown in Table 8. Alternatively or additionally, the crystalline hydrochloride salt of sepiapterin is characterized by a DSC curve that exhibits an endotherm at 225.9°C.

[0087] [Table 8]

[0088] In some embodiments, the crystalline form 1 methanesulfonate of sepiapterin is characterized by peaks in an X-ray diffraction pattern observed at a refraction angle 2θ of at least 7.8°±0.5, such as 7.8°±0.5, such as 7.8°±0.2, 23.5°±0.5, such as 23.5°±0.2, and 29.0°±0.5, such as 29.0°±0.2. In some embodiments, the most intense peak in the X-ray diffraction pattern is observed at a refraction angle 2θ of 23.5°±0.5, such as 23.5°±0.2. In an essentially pure material of the crystalline form 1 methanesulfonate of sepiapterin, peaks may be observed at refraction angles 2θ as shown in Table 9. Alternatively or additionally, the crystalline form 1 methanesulfonate of sepiapterin is characterized by a DSC curve exhibiting two endotherms at 186.0°C and 229.1°C.

[0089] [Table 9]

[0090] In some embodiments, the crystalline form 2 methanesulfonate of sepiapterin is characterized by X-ray diffraction pattern peaks observed at refraction angles 2θ of at least 7.8°±0.5, such as 7.9°±0.5, such as 7.9°±0.2, 23.4°±0.5, such as 23.4°±0.2, and 28.9°±0.5, such as 28.9°±0.2. In some embodiments, the most intense peaks in the X-ray diffraction pattern are observed at a refraction angle 2θ of 23.5°±0.5, such as 23.5°±0.2. In an essentially pure material of the crystalline form 2 methanesulfonate of sepiapterin, peaks may be observed at refraction angles 2θ as shown in Table 10. Alternatively or additionally, the crystalline form of sepiapterin di-methanesulfonate is characterized by a DSC curve exhibiting three endotherms at 75.5°C, 182.6°C and 234.9°C.

[0091] [Table 10]

[0092] In some embodiments, the crystalline form 3 methanesulfonate of sepiapterin is characterized by peaks in an X-ray diffraction pattern observed at refraction angles 2θ of at least 7.8°±0.5, such as 21.7°±0.5, such as 21.7°±0.2, 26.0°±0.5, such as 26.0°±0.2, and 28.9°±0.5, such as 28.9°±0.2. In some embodiments, the most intense peak in the X-ray diffraction pattern is observed at a refraction angle 2θ of 26.0°±0.5, such as 26.0°±0.2. In an essentially pure material of the crystalline form 3 methanesulfonate of sepiapterin, peaks may be observed at refraction angles 2θ as shown in Table 11. Alternatively or additionally, the crystalline form of sepiapterin tri-methanesulfonate is characterized by a DSC curve exhibiting two endotherms at 195.1°C and 240.1°C.

[0093] [Table 11]

[0094] In some embodiments, the crystalline nicotinate salt of sepiapterin is characterized by peaks in an X-ray diffraction pattern observed at refraction angles 2θ of at least 9.5°±0.5, e.g., 9.5°±0.2, 9.9°±0.5, e.g., 9.9°±0.2, and 24.5°±0.5, e.g., 24.5°±0.2. In some embodiments, the most intense peaks in the X-ray diffraction pattern are observed at a refraction angle 2θ of 24.5°±0.5, e.g., 24.5°±0.2. In an essentially pure material of the crystalline nicotinate salt of sepiapterin, peaks may be observed at refraction angles 2θ as shown in Table 12. Alternatively or additionally, the crystalline nicotinate salt of sepiapterin is characterized by a DSC curve exhibiting an endotherm at 221.9°C.

[0095] [Table 12]

[0096] In some embodiments, the crystalline p-toluenesulfonate salt of sepiapterin is characterized by peaks in the X-ray diffraction pattern observed at refraction angles 2θ of at least 6.5°±0.5, e.g., 6.5°±0.2, 15.1°±0.5, e.g., 15.1°±0.2, and 23.4°±0.5, e.g., 23.4°±0.2. In some embodiments, the most intense peak in the X-ray diffraction pattern is observed at a refraction angle 2θ of 6.5°±0.5, e.g., 6.5°±0.2. In an essentially pure material of p-toluenesulfonate salt of sepiapterin, peaks may be observed at refraction angles 2θ as shown in Table 13. Alternatively or additionally, the crystalline p-toluenesulfonate salt of sepiapterin is characterized by a DSC curve exhibiting three endotherms at 77.2°C, 202.4°C, and 260.2°C.

[0097] [Table 13]

[0098] In some embodiments, the crystalline benzenesulfonate salt of sepiapterin is characterized by X-ray diffraction pattern peaks observed at refraction angles 2θ of at least 6.5°±0.5, e.g., 6.5°±0.2, 14.8°±0.5, e.g., 14.8°±0.2, and 19.6°±0.5, e.g., 19.6°±0.2. In some embodiments, the most intense peaks in the X-ray diffraction pattern are observed at refraction angles 2θ of 6.5°±0.5, e.g., 6.5°±0.2. In an essentially pure material of the benzenesulfonate salt of sepiapterin, peaks may be observed at refraction angles 2θ as shown in Table 14. Alternatively or additionally, the crystalline benzenesulfonate salt of sepiapterin is characterized by a DSC curve exhibiting two endotherms at 202.3°C and 265.5°C.

[0099] [Table 14]

[0100] In some embodiments, the crystalline phosphate salt of sepiapterin is characterized by X-ray diffraction pattern peaks observed at refraction angles 2θ of at least 16.6°±0.5, e.g., 16.6°±0.2, 22.2°±0.5, e.g., 22.2°±0.2, and 25.6°±0.5, e.g., 25.6°±0.2. In some embodiments, the most intense peaks in the X-ray diffraction pattern are observed at refraction angles 2θ of 25.6°±0.5, e.g., 25.6°±0.2. In an essentially pure material of the crystalline phosphate salt of sepiapterin, peaks may be observed at refraction angles 2θ as shown in Table 15. Alternatively or additionally, the crystalline phosphate salt of sepiapterin is characterized by a DSC curve exhibiting three endotherms at 125.9°C, 152.1°C, and 157.6°C.

[0101] [Table 15]

[0102] In some embodiments, the crystalline malonate salt of sepiapterin is characterized by X-ray diffractogram peaks observed at refraction angles 2θ of at least 6.9°±0.5, e.g., 6.9°±0.2, 22.7°±0.5, e.g., 22.7°±0.2, and 23.8°±0.5, e.g., 23.8°±0.2. In some embodiments, the most intense peaks in the X-ray diffractogram are observed at refraction angles 2θ of 6.9°±0.5, e.g., 6.9°±0.2. In an essentially pure material of the crystalline malonate salt of sepiapterin, peaks may be observed at refraction angles 2θ as shown in Table 16. Alternatively or additionally, the crystalline malonate salt of sepiapterin is characterized by a DSC curve exhibiting a melting event at 115.8°C.

[0103] [Table 16]

[0104] In some embodiments, the crystalline L-tartrate salt of sepiapterin is characterized by X-ray diffractogram peaks observed at refraction angles 2θ of at least 7.3°±0.5, e.g., 7.3°±0.2, 14.2°±0.5, e.g., 14.2°±0.2, and 21.8°±0.5, e.g., 21.8°±0.2. In some embodiments, the most intense peak in the X-ray diffractogram is observed at a refraction angle 2θ of 6.9°±0.5, e.g., 6.9°±0.2. In an essentially pure material of the crystalline L-tartrate salt of sepiapterin, peaks may be observed at refraction angles 2θ as shown in Table 17. Alternatively or additionally, the crystalline L-tartrate salt of sepiapterin is characterized by a DSC curve exhibiting two endotherms at 97.2°C and 160.6°C.

[0105] [Table 17]

[0106] In some embodiments, the crystalline gentisate salt of sepiapterin is characterized by X-ray diffraction pattern peaks observed at a refraction angle 2θ of at least 7.1°±0.5, e.g., 7.1°±0.2, 8.7°±0.5, e.g., 8.7°±0.2, and 26.7°±0.5, e.g., 26.7°±0.2. In some embodiments, the most intense peak in the X-ray diffraction pattern is observed at a refraction angle 2θ of 7.1°±0.5, e.g., 7.1°±0.2. In an essentially pure material of the crystalline gentisate salt of sepiapterin, peaks may be observed at refraction angles 2θ as shown in Table 18. Alternatively or additionally, the crystalline gentisate salt of sepiapterin is characterized by a DSC curve exhibiting three endotherms at 70.5°C, 128.2°C, and 184.7°C.

[0107] [Table 18]

[0108] In some embodiments, the crystalline fumarate salt of sepiapterin is characterized by peaks in the X-ray diffraction pattern observed at a refraction angle 2θ of at least 11.3°±0.5, e.g., 11.3°±0.2, 24.0°±0.5, e.g., 24.0°±0.2, and 28.2°±0.5, e.g., 28.2°±0.2. In some embodiments, the most intense peaks in the X-ray diffraction pattern are observed at a refraction angle 2θ of at least 24.0°±0.5, e.g., 24.0°±0.2. In an essentially pure material of the crystalline fumarate salt of sepiapterin, peaks may be observed at refraction angles 2θ as shown in Table 19. Alternatively or additionally, the crystalline fumarate salt of sepiapterin may be characterized by a DSC curve exhibiting two endotherms at 114.3°C and 229.7°C.

[0109] [Table 19]

[0110] In some embodiments, the crystalline glycolate salt of sepiapterin is characterized by peaks in an X-ray diffractogram observed at a refraction angle 2θ of at least 7.6°±0.5, e.g., 7.6°±0.2, 10.7°±0.5, e.g., 10.7°±0.2, and 24.0°±0.5, e.g., 24.0°±0.2. In some embodiments, the most intense peak in the X-ray diffractogram is observed at a refraction angle 2θ of 7.6°±0.5, e.g., 7.6°±0.2. In an essentially pure material of the crystalline glycolate salt of sepiapterin, peaks may be observed at refraction angles 2θ as shown in Table 20. Alternatively or additionally, the crystalline glycolate salt of sepiapterin is characterized by a DSC curve exhibiting two endotherms at 133.9°C and 147.7°C.

[0111] [Table 20]

[0112] In some embodiments, the crystalline acetate salt of sepiapterin is characterized by peaks in the X-ray diffraction pattern observed at refraction angles 2θ of at least 6.2°±0.5, e.g., 6.2°±0.2, 12.0°±0.5, e.g., 12.0°±0.2, and 18.1°±0.5, e.g., 18.1°±0.2. In some embodiments, the most intense peaks in the X-ray diffraction pattern are observed at refraction angles 2θ of at least 6.2°±0.5, e.g., 6.2°±0.2. In an essentially pure material of the crystalline acetate salt of sepiapterin, peaks may be observed at refraction angles 2θ as shown in Table 21. Alternatively or additionally, the crystalline acetate salt of sepiapterin is characterized by a DSC curve exhibiting two endotherms at 146.1°C and 175.4°C.

[0113] [Table 21]

[0114] In some embodiments, the crystalline form 1 sulfate of sepiapterin is characterized by peaks in an X-ray diffraction pattern observed at refraction angles 2θ of at least 5.1°±0.5, e.g., 5.1°±0.2, 7.8°±0.5, e.g., 7.8°±0.2, and 23.0°±0.5, e.g., 23.0°±0.2. In some embodiments, the most intense peak in the X-ray diffraction pattern is observed at a refraction angle 2θ of 5.1°±0.5, e.g., 5.1°±0.2. In an essentially pure material of crystalline form 1 sulfate of sepiapterin, peaks may be observed at refraction angles 2θ as shown in Table 22. Alternatively or additionally, the crystalline form 1 sulfate of sepiapterin is characterized by a DSC curve exhibiting three endotherms at 94.5°C, 158.3°C, and 209.9°C.

[0115] [Table 22]

[0116] In some embodiments, the crystalline disulfate salt of sepiapterin is characterized by X-ray diffractogram peaks observed at refraction angles 2θ of at least 7.8°±0.5, e.g., 7.8°±0.2, 8.8°±0.5, e.g., 8.8°±0.2, and 24.1°±0.5, e.g., 24.1°±0.2. In some embodiments, the most intense peak in the X-ray diffractogram is observed at a refraction angle 2θ of 8.8°±0.5, e.g., 8.8°±0.2. In an essentially pure material of the crystalline disulfate salt of sepiapterin, peaks may be observed at refraction angles 2θ as shown in Table 23.

[0117] [Table 23]

[0118] The present invention can employ pharmaceutical compositions comprising a pharma- ceutical acceptable excipient and an effective amount of sepiapterin or a pharma- ceutical acceptable salt thereof. Examples of pharmaceutical compositions of sepiapterin and its salts can be found in WO 2019 / 046849 and WO 2019 / 232120, which compositions are incorporated herein by reference in their entirety.

[0119] The pharma- ceutically acceptable excipients may be any of those conventionally used, limited only by physicochemical considerations such as solubility, route of administration, etc. Those skilled in the art will appreciate that in addition to the pharmaceutical compositions described below, sepiapterin can be formulated as an inclusion complex, such as a cyclodextrin inclusion complex, or as liposomes.

[0120] The pharma- ceutically acceptable excipients described herein, such as vehicles, adjuvants, excipients, or diluents, are well known to those skilled in the art and are readily available. Preferably, the pharma- ceutically acceptable excipients are chemically inert to sepiapterin and have no adverse side effects or toxicity under the conditions of use.

[0121] Formulations that increase gastric and / or foregut residence time Gastroretentive drug delivery is an approach where the drug formulation is designed to remain in the stomach for a longer period until drug release is complete.

[0122] Bioadhesive formulations utilize polymers that can adhere to a surface and provide controlled release of the drug. Bioadhesive polymers can be anionic (e.g., carboxymethylcellulose, alginic acid, polyacrylic acid, pectin, carrageenan, polycarbophil, or carbomer), cationic (e.g., chitosan, polylysine, or polybrene), or non-ionic (e.g., polyethylene glycol, polyvinylpyrrolidone, dextran, or hydroxypropylmethylcellulose).

[0123] High density formulations are designed to reside in the stomach at levels below the pyloric sphincter, thus avoiding emptying. Suitable excipients for high density formulations include iron powder, barium sulfate, zinc oxide, titanium dioxide, etc.

[0124] Bulking formulations are designed to expand in the stomach to be larger than the pyloric sphincter, thus avoiding hunger. For example, a formulation comprising a drug core, a swellable hydrocolloid, and an outer semipermeable polymer is suitable for a bulking formulation.

[0125] Superporous hydrogel formulations, like the expanding formulations, are designed to expand larger than the pyloric sphincter in the stomach. Superporous hydrogel formulations may include polymers such as croscarmellose sodium.

[0126] Floating formulations are designed to have a density lower than gastric fluids and may include compositions that include ion exchange resins, raft systems, inflatable chambers, effervescent mixtures, swellable hydrocolloids, or multiparticulate systems.

[0127] Antioxidants Sepiapterin tends to oxidize rapidly when exposed to air. Therefore, the pharmaceutical composition of the present invention may include an antioxidant. The antioxidant may minimize the oxidative degradation of sepiapterin. Examples of antioxidants include, but are not limited to, ascorbic acid, tocopherol, retinol, ascorbyl palmitate, N-acetylcysteine, glutathione, ethylenediaminetetraacetic acid, sodium bisulfite, sodium metabisulfite, thiourea, butylated hydroxytoluene, butylated hydroxyanisole, and vitamin E. In some embodiments, the pharmaceutical composition of the present invention includes ascorbic acid, tocopherol, retinol, ascorbyl palmitate, N-acetylcysteine, glutathione, butylated hydroxytoluene, and / or butylated hydroxyanisole as antioxidants.

[0128] In some embodiments, the pharmaceutical composition comprises less than 10% by weight of antioxidant, e.g., less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2%, less than 1%, or substantially no antioxidant. In some embodiments, the pharmaceutical composition comprises 2-9% by total weight of antioxidant, e.g., 2-4%, 3-5%, 4-6%, 5-7%, 6-8%, or 7-9%. In some embodiments, the pharmaceutical composition comprises 5-100% of the USP maximum daily dose of antioxidant, e.g., in some embodiments, the pharmaceutical composition comprises about 5%, 10%, 15%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% of the USP maximum daily dose of antioxidant. In some embodiments, the ratio of sepiapterin to antioxidant is at least 1:1 by weight, e.g., 2:1, 3:1, 4:1, 5:1, 6:1, 7:1, 8:1, 9:1, or 10:1.

[0129] Dispersants In some embodiments, the pharmaceutical composition of the present invention includes at least one dispersing agent. The dispersing agent may separate particles in the formulation, for example, releasing its medicinal agent upon contact with moisture. Examples of dispersing agents include, but are not limited to, cross-linked polyvinylpyrrolidone, carboxymethylcellulose (e.g., croscarmellose salts, e.g., croscarmellose sodium), starch (e.g., sodium starch glycolate), or alginic acid. In some embodiments, the dispersing agent in the pharmaceutical composition is carboxymethylcellulose, such as a pharma- ceutically acceptable salt of croscarmellose. In some embodiments, the pharmaceutical composition may include 0.1-1.5% (e.g., about 0.1%, 0.5%, 1%, or 1.5%) of dispersing agent by total weight. In some embodiments, the pharmaceutical composition includes less than 1.5% (e.g., less than 1%, less than 0.5%, or less than 0.1%) of dispersing agent by total weight.

[0130] Anti-caking Agent In some embodiments, the pharmaceutical composition of the present invention comprises at least one anti-caking agent. In some embodiments, the pharmaceutical composition of the present invention comprises at least two anti-caking agents. Exemplary anti-caking agents include colloidal silicon dioxide, microcrystalline cellulose, tricalcium phosphate, microcrystalline cellulose, magnesium stearate, sodium bicarbonate, sodium ferrocyanide, potassium ferrocyanide, calcium ferrocyanide, calcium phosphate, sodium silicate, colloidal silicon dioxide, calcium silicate, magnesium trisilicate, talcum powder, sodium aluminosilicate, potassium aluminosilicate, calcium aluminosilicate, bentonite, aluminum silicate, stearic acid, and polydimethylsiloxane. In some embodiments, the at least one anti-caking agent is colloidal silicon dioxide or microcrystalline cellulose. In some embodiments, the pharmaceutical composition may comprise 65-75% (e.g., about 65%, 67%, 70%, 73%, or 75%) of the anti-caking agent by total weight. In some embodiments, the pharmaceutical composition comprises both colloidal silicon dioxide and microcrystalline cellulose. In some embodiments, the pharmaceutical composition comprises 60-65% microcrystalline cellulose by total weight and 5-7% colloidal silicon dioxide by total weight.

[0131] Dosing Vehicle In some embodiments, the pharmaceutical compositions of the present invention are combined with a dosing vehicle prior to administration (e.g., a dosing vehicle having a viscosity of about 50 to 1750 centipoise (cP)). One type of suspending agent that can be used is a combination of glycerin and sucrose in water (e.g., MEDISCA 2.5% glycerin and 27% sucrose in water). (R) An appropriate amount of the composition can be added to the dosing vehicle mixture and stirred to suspend the composition immediately prior to administration.

[0132] Other suspending agents may also be used as administration vehicles. Exemplary suspending agents include agar, alginic acid, sodium carboxymethylcellulose, carrageenan, dextrin, gelatin, guar gum, hydroxyethylcellulose, hydroxypropylcellulose, hypromellose, methylcellulose, polyethylene glycol, povidone, tragacanth, xanthan gum, or other suspending agents known in the art.

[0133] dose Sepiapterin or a pharma- ceutically acceptable salt thereof can be used in any suitable dose. Appropriate doses and dosing regimens can be determined within the skill of the art. Generally, treatment is initiated with a dose smaller than the optimal dose. Thereafter, the dose is increased in small increments until the optimum effect under the circumstances is obtained. For convenience, the total daily dose can be administered in divided doses throughout the day, if desired. At appropriate doses and appropriate administration of a particular compound, the present invention provides a wide range of responses. Typically, the dosage is in the range of about 2.5 to about 150 mg / kg body weight / day of the subject being treated (e.g., 60 mg / kg / day). For example, in embodiments, sepiapterin, or a pharma- ceutically acceptable salt thereof, can be administered at about 20 mg / kg to about 150 mg / kg, about 20 mg / kg to about 60 mg / kg, about 40 mg / kg to about 100 mg / kg, about 100 mg / kg to about 150 mg / kg, about 60 mg / kg to about 120 mg / kg, about 80 mg / kg to about 100 mg / kg, about 40 mg / kg to about 60 mg / kg, about 2.5 mg / kg to about 20 mg / kg, about 2.5 mg / kg to about 10 mg / kg, or about 2.5 mg / kg to about 5 mg / kg of subject body weight per day, one or more times per day, to obtain the desired therapeutic effect.

[0134] In some embodiments, sepiapterin or a pharma- ceutically acceptable salt thereof can be formulated as a unit solid oral dosage formulation, such as particles, in these embodiments, each unit solid oral dosage formulation, e.g., a sachet, can contain any suitable amount of sepiapterin or a pharma- ceutically acceptable salt thereof. For example, each solid oral dosage formulation can contain about 2.5 mg, about 5 mg, about 10 mg, about 20 mg, about 30 mg, about 40 mg, about 50 mg, about 60 mg, about 70 mg, about 80 mg, about 90 mg, about 100 mg, about 125 mg, about 150 mg, about 175 mg, about 200 mg, about 225 mg, about 250 mg, about 275 mg, about 300 mg, about 325 mg, about 350 mg, about 375 mg, about 400 mg, about 425 mg, about 450 mg, about 475 mg, about 500 mg, about 750 mg, about 1 g, about 1.25 g, or about 1.5 g.

[0135] Sepiapterin or its pharma- ceutically acceptable salts can be used to prepare liquid preparations, such as in the form of a solution, suspension or emulsion. Preparations suitable for oral administration include, but are not limited to, (a) solids or granules, such as capsules, sachets, tablets, lozenges, and troches, each containing a predetermined amount of active ingredient; (b) powders; (c) liquids, such as an effective amount of the compound dissolved in a diluent, such as water, saline, or orange juice; (d) suspensions in a suitable liquid; and (e) suitable emulsions. Preferred are solid oral preparations, such as capsules, tablets, and powders. Capsule forms can be of the usual hard or soft shelled gelatin type, for example, containing surfactants, lubricants, and inert fillers, such as lactose, sucrose, calcium phosphate, and cornstarch. Tablet forms may contain one or more of lactose, sucrose, mannitol, corn starch, potato starch, alginic acid, microcrystalline cellulose, acacia, gelatin, guar gum, colloidal silicon dioxide, croscarmellose sodium, talc, magnesium stearate, calcium stearate, zinc stearate, stearic acid, and other excipients, coloring agents, diluents, buffers, disintegrants, wetting agents, preservatives, flavoring agents, and pharmacologically compatible excipients. Lozenge forms may contain the active ingredient in addition to the active ingredient in a flavor (usually sucrose and acacia or tragacanth), and similarly pastilles may contain the active ingredient in an inert base such as gelatin and glycerin, or sucrose and acacia, emulsions, gels, and the like, such excipients being known in the art.

[0136] Formulations suitable for oral and / or parenteral administration include aqueous and non-aqueous isotonic sterile injection solutions which may contain antioxidants, buffers, bacteriostats, and solutes that render the formulation isotonic with the blood of the intended recipient, and aqueous and non-aqueous sterile suspensions which may contain suspending agents, solubilizing agents, thickening agents, stabilizers, and preservatives. The compounds of the invention may be administered in physiologically acceptable diluents in pharmaceutical vehicles such as water, saline, aqueous dextrose and related sugar solutions, alcohols such as ethanol, benzyl alcohol, or hexadecyl alcohol, glycols such as propylene glycol or polyethylene glycol and polyethylene alcohol, glycerol ketals such as 2,2-dimethyl-1,3-dioxolane-4-methanol, ethers such as poly(ethylene glycol) 400, oils, fatty acids, fatty acid esters or glycerides, or acetylated fatty acid glycerides with or without the addition of pharma- ceutically acceptable surfactants such as soaps or detergents, suspending agents such as pectin, carbomers, methylcellulose, hydroxypropylmethylcellulose, or carboxymethylcellulose, or emulsifying agents, and other pharmaceutical adjuvants.

[0137] The present invention features a pharmaceutical composition in an orally acceptable formulation, comprising a therapeutically effective amount of sepiapterin and less than 10% antioxidant. In some embodiments, the pharmaceutical composition is a granular formulation dispersed in a pharma- ceutically acceptable excipient, e.g., the composition can be mixed in water and ingested by a subject (e.g., within 5 to 10 minutes). Suitable formulations for use in the present invention are described in Remington's Pharmaceutical Sciences, Mack Publishing Company, Philadelphia, PA 22nd ed., 2010. Except insofar as any conventional excipient is incompatible with the active ingredient, its use in the pharmaceutical composition of the present invention is contemplated. In addition, it will be understood that for animal (e.g., human) administration, the formulation should meet sterility, pyrogenicity, general safety and purity standards as required by FDA Office of Biological Standards.

[0138] Oral solid dosage forms Formulations for oral use include particles containing the active ingredient in a mixture with non-toxic pharma- ceutically acceptable excipients, and such formulations are known to those skilled in the art (e.g., U.S. Patent Nos.: 5,817,307, 5,824,300, 5,830,456, 5,846,526, 5,882,640, 5,910,304, 6,036,949, 6,036,949, 6,372,218, which are incorporated herein by reference). The excipient may be, for example, an inert diluent or filler (e.g., sucrose, sorbitol, sugar, mannitol, microcrystalline cellulose, starch including potato starch, calcium carbonate, sodium chloride, lactose, calcium phosphate, calcium sulfate, or sodium phosphate), a granulating / disintegrating agent (e.g., cellulose derivatives including microcrystalline cellulose, starch including potato starch, croscarmellose sodium, alginates or alginic acid), a binder (e.g., sucrose, glucose, sorbitol, acacia, alginic acid, sodium alginate, gelatin, starch, pregelatinized starch, microcrystalline cellulose, magnesium aluminum silicate, sodium carboxymethylcellulose, methylcellulose, hydroxypropylmethylcellulose, ethylcellulose, polyvinylpyrrolidone ... Lidone, or polyethylene glycol), and lubricants, glidants, anti-adherents (e.g., magnesium stearate, zinc stearate, stearic acid, silica, hydrogenated vegetable oil, or talc), and anti-caking agents (e.g., colloidal silicon dioxide, microcrystalline cellulose, tricalcium phosphate, microcrystalline cellulose, magnesium stearate, sodium bicarbonate, sodium ferrocyanide, potassium ferrocyanide, calcium ferrocyanide, calcium phosphate, sodium silicate, colloidal silicon dioxide, calcium silicate, magnesium trisilicate, talcum powder, sodium aluminosilicate, potassium aluminosilicate, calcium aluminosilicate, bentonite, aluminosilicate, stearic acid, polydimethylsiloxane). Other pharma- ceutically acceptable excipients may be colorants, flavorings, plasticizers, humectants, and buffers. In some embodiments, the excipients (e.g., flavorings) are packaged with the composition.In some embodiments, excipients (eg, flavorings) are packaged separately from the composition (eg, combined with the composition prior to administration).

[0139] The solid compositions of the invention may include a coating adapted to protect the composition from undesired chemical changes (e.g., chemical degradation prior to release of the active substance). The coating may be applied to the solid dosage form in a manner similar to that described in the Encyclopedia of Pharmaceutical Technology, supra.

[0140] The above ingredients may be used to prepare powders or granules in a conventional manner, for example, using a mixer, a fluidized bed apparatus, a melt-solidification apparatus, a rotor granulator, an extrusion / spheronization apparatus, or a spray drying apparatus.

[0141] Methods of Treatment Sepiapterin can function as a useful therapeutic agent for diseases associated with elevated blood phenylalanine levels, such as hyperphenylalaninemia or phenylketonuria.As described herein, various forms of these diseases, such as hyperphenylalaninemia caused by phenylketonuria, tetrahydrobiopterin-responsive phenylketonuria, sepiapterin-responsive phenylketonuria, classical phenylketonuria, or non-classical phenylketonuria, can be treated.Therefore, various forms of sepiapterin or its salts according to the present invention can be administered to a subject in an amount effective to obtain treatment or improvement of a disease, disorder, or condition.

[0142] Subjects treated by the methods herein typically have a blood phenylalanine concentration (e.g., unregulated phenylalanine blood concentration) greater than 120 micromoles / liter (e.g., greater than 200 micromoles / liter, greater than 300 micromoles / liter, greater than 360 micromoles / liter, greater than 400 micromoles / liter, greater than 450 micromoles / liter, greater than 500 micromoles / liter, greater than 550 micromoles / liter, greater than 600 micromoles / liter, greater than 650 micromoles / liter, greater than 700 micromoles / liter, greater than 800 micromoles / liter, greater than 900 micromoles / liter, greater than 1000 micromoles / liter, greater than 1100 micromoles / liter, or greater than 1200 micromoles / liter). Subjects may also be stratified by having a blood phenylalanine concentration (e.g., uncontrolled phenylalanine blood concentration) between 120 and 360 micromoles / liter, between 360 and 600 micromoles / liter, between 600 and 1200 micromoles / liter, or greater than 1200 micromoles / liter.

[0143] Subjects who have insufficient reduction in blood phenylalanine under existing therapy (e.g., phenylalanine-restricted diet, sapropterin, or pegvaliase-pqpz) can also be treated by the methods described herein. For example, subjects who are treated with a therapy other than sepiapterin may have a blood phenylalanine concentration greater than 120 micromoles / liter under existing management (e.g., greater than 200 micromoles / liter, greater than 300 micromoles / liter, greater than 360 micromoles / liter, greater than 400 micromoles / liter, greater than 450 micromoles / liter, greater than 500 micromoles / liter, greater than 550 micromoles / liter, greater than 600 micromoles / liter, greater than 650 micromoles / liter, greater than 700 micromoles / liter, greater than 800 micromoles / liter, greater than 900 micromoles / liter, greater than 1000 micromoles / liter, greater than 1100 micromoles / liter, or greater than 1200 micromoles / liter).

[0144] By administering sepiapterin or a pharma- ceutically acceptable salt thereof, the subject's blood phenylalanine concentration can be reduced to less than 600 micromoles / liter, for example, less than 360 micromoles / liter or less than 120 micromoles / liter, for example, 120 to 360 micromoles / liter or 360 to 600 micromoles / liter. Alternatively or additionally, by administering sepiapterin, for example, the subject's blood phenylalanine concentration before administration of sepiapterin can be reduced by at least 10% (for example, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or about 10% to about 30%, about 20% to about 40%, about 30% to about 50%, about 40% to about 60%, about 50% to about 70%, about 60% to about 80%, or about 70% to about 90%). The decrease can be measured after administration for at least 1 week (e.g., 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, at least about 2 weeks, at least about 3 weeks, at least about 4 weeks, at least about 5 weeks, at least about 6 weeks). Also, administration of sepiapterin can result in a BH4 concentration of at least 50 ng / ml (e.g., at least 60 ng / ml, at least 100 ng / ml, at least 200 ng / ml, at least 400 ng / ml, at least 600 ng / ml, at least 1000 ng / ml, or at least 2000 ng / ml, or 50 ng / ml to 100 ng / ml, 60 ng / ml to 400 ng / ml, 200 ng / ml to 600 ng / ml, 400 ng / ml to 1000 ng / ml, or 600 ng / ml to 1500 ng / ml) in the subject's plasma within 10 hours after administration.

[0145] The subject may be one who has not responded to treatment with sapropterin or a pharma- ceutically acceptable salt thereof (e.g., sapropterin dihydrochloride) or pegvaliase-pqpz. For example, administration of pegvaliase-pqpz may result in a decrease in the subject's plasma concentration by less than 20%. For example, administration of at least about 10 mg / kg (e.g., at least about 15 mg / kg, at least about 20 mg / kg) of sapropterin or a pharma- ceutically acceptable salt thereof (e.g., sapropterin dihydrochloride) results in a decrease in the subject's plasma concentration by less than 30%, or administration of at least about 20 mg of pegvaliase-pqpz once a day for at least 8 days (e.g., at least 14 days, at least 21 days, at least 28 days, at least 30 days) results in a decrease in the subject's plasma concentration by less than 20%. In some embodiments, administration of sapropterin or a pharma- ceutically acceptable salt thereof (e.g., sapropterin dihydrochloride), for example, administration of at least about 10 mg / kg (e.g., at least about 15 mg / kg, at least about 20 mg / kg), reduces the subject's plasma concentration by less than 15-30%. In some embodiments, administration of sapropterin or a pharma- ceutically acceptable salt thereof (e.g., sapropterin dihydrochloride), for example, administration of at least about 10 mg / kg (e.g., at least about 15 mg / kg, at least about 20 mg / kg), reduces the subject's plasma concentration by less than 30%. In some embodiments, administration of sapropterin or a pharma- ceutically acceptable salt thereof (e.g., sapropterin dihydrochloride), for example, administration of at least about 10 mg / kg (e.g., at least about 15 mg / kg, at least about 20 mg / kg), reduces the subject's plasma concentration by less than 20%. In some embodiments, administration of sapropterin or a pharma- ceutically acceptable salt thereof (e.g., sapropterin dihydrochloride), e.g., at least about 10 mg / kg (e.g., at least about 15 mg / kg, at least about 20 mg / kg), reduces the subject's plasma concentration by less than 15%.In some embodiments, sapropterin or a pharma- ceutically acceptable salt thereof (e.g., sapropterin dihydrochloride) is administered for at least 8 days (e.g., at least 14 days, at least 21 days, at least 28 days, at least 30 days) or pegvalase-pqpz is administered for at least about 20 mg / kg. After administration once daily for at least 16 weeks (e.g., at least 18 weeks, at least 20 weeks, at least 22 weeks, at least 24 weeks), the subject had a plasma concentration greater than 120 micromoles / liter (e.g., greater than 200 micromoles / liter, greater than 300 micromoles / liter, greater than 360 micromoles / liter, greater than 400 micromoles / liter, greater than 450 micromoles / liter, greater than 500 micromoles / liter, greater than 550 micromoles / liter, greater than 600 micromoles / liter, greater than 650 micromoles / liter, greater than 700 micromoles / liter, greater than 800 micromoles / liter, greater than 900 micromoles / liter, greater than 1000 micromoles / liter, greater than 1100 micromoles / liter, or greater than 1200 micromoles / liter).For example, sapropterin or a pharma- ceutically acceptable salt thereof (e.g., sapropterin dihydrochloride) at least 10 mg / kg (e.g., at least about 15 mg / kg, at least about 20 mg / kg) for at least 8 days (e.g., at least 14 days, at least 21 days, at least 28 days, at least 30 days), or pegvalase-pqpz at least about 20 mg / kg (e.g., at least 20 mg / kg) for at least 8 days (e.g., at least 14 days, at least 21 days, at least 28 days, at least 30 days). After administration once daily for at least 16 weeks (e.g., at least 18 weeks, at least 20 weeks, at least 22 weeks, at least 24 weeks), the subject had a plasma concentration greater than 120 micromoles / liter (e.g., greater than 200 micromoles / liter, greater than 300 micromoles / liter, greater than 360 micromoles / liter, greater than 400 micromoles / liter, greater than 450 micromoles / liter, greater than 500 micromoles / liter, greater than 550 micromoles / liter, greater than 600 micromoles / liter, greater than 650 micromoles / liter, greater than 700 micromoles / liter, greater than 800 micromoles / liter, greater than 900 micromoles / liter, greater than 1000 micromoles / liter, greater than 1100 micromoles / liter, or greater than 1200 micromoles / liter).

[0146] A subject may also be administered sepiapterin after discontinuing treatment with pegvalase pqpz due to adverse reactions (e.g., anaphylaxis, injection site reactions, arthralgia, hypersensitivity reactions, headache, generalized skin reactions lasting at least 14 days, pruritus, nausea, abdominal pain, oropharyngeal soreness, vomiting, cough, diarrhea, and / or fatigue) and / or tolerability. The risk of adverse events (e.g., headache, rhinorrhea, sore throat, diarrhea, vomiting, cough, and / or stuffy nose) may be reduced compared to a subject administered at least 10 mg / kg (e.g., at least 15 mg / kg, at least 20 mg / kg) of sapropterin or a pharma- ceutically acceptable salt thereof (e.g., sapropterin dihydrochloride).

[0147] In some embodiments, the subject is on a phenylalanine restricted diet (e.g., the subject is on a diet that includes a milk substitute or blend, such as Phenyl-Free 2, and measured amounts of fruits, vegetables, bread, pasta, and cereal). In some embodiments, the subject is not on a phenylalanine restricted diet (e.g., the subject is not on a diet that includes a milk substitute or blend, such as Phenyl-Free 2, and measured amounts of fruits, vegetables, bread, pasta, and cereal). In some embodiments, the subject has a median phenylalanine intake of less than 3000 mg / day (e.g., less than 2500 mg / day, less than 2000 mg / day, less than 1500 mg / day, less than 1000 mg / day, less than 500 mg / day). Treatment with sepiapterin, in conjunction with phenylalanine intake or natural protein intake, can allow for a reduction in blood phenylalanine levels (e.g., less than 600, 360, or 120 micromoles / liter). For example, the subject may be receiving at least about 1000 mg / day (e.g., at least about 1100 mg / day, at least about 1200 mg / day, at least about 1300 mg / day, at least about 1400 mg / day, at least about 1500 mg / day, at least about 1600 mg / day, at least about 1700 mg / day, at least about 1800 mg / day, at least about 1900 mg / day, or at least Approximately 2000 mg / day, or 1000 mg / day to 1400 mg / day, 1200 mg / day to 1600 mg / day, 1300 mg / day to 1700 mg / day, 1600 mg / day to 2000 mg / day, 1800 mg / day to 2400 mg / day, 2000 mg / day to 3000 mg / day, 3000 mg / day to 4000 mg / day, 4000 to 5000 mg / day) of phenylalanine may be taken. In some embodiments, less than 25% of the protein in the subject's diet is naturally occurring protein (e.g., less than 20%, less than 15%, less than 10%, or less than 5%), or more than 25% of the protein in the subject's diet is naturally occurring protein (e.g., more than 30%, more than 40%, more than 50%, more than 60%, more than 70%, more than 80%, more than 90%, more than 95%, more than 99%).In some embodiments, the subject has a natural protein intake of greater than 10 g / day (e.g., greater than 20 g / day, greater than 30 g / day, greater than 40 g / day, greater than 50 g / day, greater than 60 g / day, greater than 70 g / day, greater than 80 g / day, or about 10 g / day to about 30 g / day, about 20 g / day to about 40 g / day, about 30 g / day to about 50 g / day, about 40 g / day to about 60 g / day, about 50 g / day to about 70 g / day, about 60 g / day to about 80 g / day). In some embodiments of any of the methods described herein, the subject has a natural protein intake of less than 10 g / day (e.g., less than 5 g / day or about 5 g / day to about 10 g / day).

[0148] In some embodiments, the subject has PKU defined as any previous blood Phe measurement >360 μmol / L plus at least one measurement >450 μmol / L, or the average of the last three blood Phe measurements >450 μmol / L. In some embodiments, the subject does not have PKU defined as any previous blood Phe measurement >360 μmol / L plus at least one measurement >450 μmol / L, or the average of the last three blood Phe measurements >450 μmol / L. In some embodiments, the subject has classical PKU defined as any documented previous measurement of blood Phe of at least 1,200 μmol / L. In some embodiments, the subject does not have classical PKU defined as any documented previous measurement of blood Phe of at least 1,200 μmol / L.

[0149] In some embodiments, the subject is a child (e.g., the subject is under 18 years old, under 17 years old, under 16 years old, under 15 years old, under 14 years old, under 13 years old, under 12 years old, under 11 years old, under 10 years old, under 9 years old, under 8 years old, under 7 years old, under 6 years old, under 5 years old, under 4 years old, under 3 years old, under 2 years old, under 1 year old). In some embodiments, the subject is an adult (e.g., the subject is over 18 years old).

[0150] In some embodiments, the treatment increases the subject's neurocognitive function (e.g., increases executive function, decreases anxiety, decreases Attention Deficit / Hyperactivity Disorder symptoms, and / or decreases instances of brain fog). Alterations in executive function can be measured using a number of measures, including the Behavioral Assessment of Dysexecutive Syndrome (BADS), Behavioral Inventory of Executive Function (BRIEF or Mini-BRIEF), Berkeley Deficits in Executive Function Scale (BDEFS), Behavioral Disorders Scale (BDS), ASEBA Child Behavior Checklist (CBLC), Comprehensive Executive Function Inventory (CEFI), CogScreen, Continuous Performance Task (CPT), Controlled Oral Word Association Test (COWAT), d2 Test of Attention, Delis-Kaplan Executive Function System (D-KEFS), Digit Vigilance Test, Figural Fluency Test, Halstead Category Test, Hayling and Brixton Tests, Iowa Gambling Task, Kaplan Baycrest This may be measured by assessments such as the Neurocognitive Assessment (KBNA), Kaufman Short Neuropsychological Assessment, Mental Clutter Scale, Paced Auditory Serial Addition Test (PASAT), Phenylketonuria-Quality of Life (PKU-QOL), Rey-Osterlie's Complex Figures, Rough Figural Fluency Test, Stroop task, Task of Executive Control, Test of Variables of Attention (TOVA), Tower of London Test, Trail Making Test (TMT) or Trails A & B, Wisconsin Card Sorting Test (WCST), Symbol Digit Modalities Test, or the Cambridge Neuropsychological Test Automated Battery (CANTAB) assessment (e.g., by measuring reaction speed, spatial span, spatial working memory, rapid digital information processing, sustained attention, and / or a stop-signal task). Treatment may result in improved attention and / or mood (e.g., as measured by the ADHD-RS 5 scale (or its inattention rating) and / or the Profile of Mood States (POMS) scale). Treatment may result in increased sleep quality and / or a decrease in symptoms associated with poor sleep.

[0151] Sepiapterin may or may not be administered with food. Without being bound by theory, administering sepiapterin with food increases plasma exposure of BH4 (e.g., by slowing the rate of absorption of sepiapterin). If the administered sepiapterin is rapidly absorbed, e.g., by administration on an empty stomach, sepiapterin reductase and / or dihydrofolate reductase in cells may be increased by V max It is believed that the excess of sepiapterin is then excreted without being converted to BH4, resulting in lower plasma BH4 levels compared to administration of sepiapterin with food, which reduces or prolongs the rate of absorption of sepiapterin and results in reaction rates below, or slightly above, the Vmax of substrate saturation of sepiaptene reductase and / or dihydrofolate reductase. Administration of sepiapterin with food unexpectedly reduced the maximum BH4 plasma concentration (Cmax) and the area under the concentration-time curve from time zero to the final concentration of BH4 (AUC) compared to administration without food. 0-lastFor example, an effective amount of sepiapterin is an amount (e.g., 2.5 mg / kg to 100 mg / kg per dose) sufficient to produce a BH4 concentration of at least 50 ng / ml (e.g., at least 60 ng / ml, at least 100 ng / ml, at least 200 ng / ml, at least 400 ng / ml, at least 600 ng / ml, at least 1000 ng / ml, or at least 2000 ng / ml, or 50 ng / ml to 100 ng / ml, 60 ng / ml to 400 ng / ml, 200 ng / ml to 600 ng / ml, 400 ng / ml to 1000 ng / ml, or 600 ng / ml to 1500 ng / ml) in the plasma of the subject within 10 hours of administration with food. An effective amount may include a dose that is at least 5% (e.g., at least 60 ng / ml, at least 100 ng / ml, at least 200 ng / ml, at least 400 ng / ml, at least 600 ng / ml, at least 1000 ng / ml, or at least 2000 ng / ml, or between 50 ng / ml and 100 ng / ml, between 60 ng / ml and 400 ng / ml, between 200 ng / ml and 600 ng / ml, between 400 ng / ml and 1000 ng / ml, or between 600 ng / ml and 1500 ng / ml) lower than a dose sufficient to provide a maximum BH4 plasma concentration (Cmax) of at least 50 ng / ml in the plasma of a subject within 10 hours of administration of sepiapterin without food.

[0152] In some embodiments of any of the methods described herein, the food is a high protein food. In some embodiments of any of the methods described herein, the food is a high fat food (e.g., at least 25, 30, 40, or 50% of the calories are from fat). In some embodiments of any of the methods described herein, the food is a high protein and high fat food. In some embodiments, the food is a high calorie food (e.g., the food contains at least 100 calories, e.g., at least 200 calories, at least 300 calories, at least 400 calories, at least 500 calories, e.g., between 500 and 1500 or between 800 and 1000 calories). In some embodiments of any of the methods described herein, the food is a meal, e.g., breakfast, lunch, or dinner. The sepiapterin may be provided in a composition separate from the ingested food (e.g., the sepiapterin is not incorporated into a food product).

[0153] Administration to a subject may occur less than 30 minutes prior to or after ingestion of food, e.g., immediately prior to ingestion of food up to 1 hour after ingestion of food, e.g., substantially simultaneously with food. Administration with food (e.g., less than 30 minutes prior to ingestion of food, or after ingestion of food, e.g., immediately prior to ingestion of food up to 1 hour after ingestion of food) may result in a greater or lesser increase in the Cmax of BH4 or the extent of BH4 production and resulting plasma exposure (AUC 0-last ) increase (e.g., at least 5%, at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 100%, at least 110%, at least 120%, at least 130%, at least 140%, or at least 150%).

[0154] The actual dosage of the composition of the present invention administered to a subject can be determined by physical and physiological factors such as body weight, severity of the condition, type of disease being treated, previous or concurrent therapeutic interventions, characteristics of the subject, and route of administration. Depending on the dosage and route of administration, the preferred dosage and / or frequency of administration of an effective amount may vary depending on the subject's response. The physician responsible for administration will, in any event, determine the concentration of active ingredient in the composition and the appropriate dosage for the individual subject.

[0155] In some embodiments, the subject is administered about 2.5 mg / kg to 120 mg / kg (e.g., about 20 mg / kg to about 60 mg / kg, or about 20 mg / kg, about 30 mg / kg, about 40 mg / kg, about 50 mg / kg, about 60 mg / kg) per dose. The subject may be administered a pharmaceutical composition containing sepiapterin once a day, twice a day, or three times a day during treatment. In some embodiments, the subject continues other current treatments for high phenylalanine blood levels (e.g., phenylalanine restricted diet) except for BH4 supplementation (if taking BH4). In some embodiments, the subject may not be allowed to take drugs known to inhibit folate synthesis (e.g., methotrexate, pemetrexate, or trimetrexate). Sepiapterin or a pharma- ceutically acceptable salt thereof may be administered in two equivalent amounts (e.g., two doses at different times of the day), such as two 60 mg / kg doses (e.g., one 60 mg / kg dose in the morning and one 60 mg / kg dose in the evening), two 40 mg / kg doses (e.g., one 40 mg / kg dose in the morning and one 40 mg / kg dose in the evening), two 30 mg / kg doses (e.g., one 30 mg / kg dose in the morning and one 30 mg / kg dose in the evening), two 20 mg / kg doses (e.g., one 20 mg / kg dose in the morning and one 20 mg / kg dose in the evening), or two 10 mg / kg doses (e.g., one 10 mg / kg dose in the morning and one 10 mg / kg dose in the evening).

[0156] In some embodiments, subjects taking BH4 discontinue administration of BH4 (i.e., prior to or concurrent with BH4 washout, e.g., initiation of sepiapterin treatment). Blood samples for Phe concentration may be obtained during the BH4 washout period 7, 5, 3, and 1 days prior to treatment with a pharmaceutical composition of the invention, or at any time during the BH4 washout period until blood Phe concentration is >360 μmol / L. In some embodiments, pre-administration blood samples are tested for sepiapterin, Phe, BH4, and tyrosine (Tyr).

[0157] Equivalence and Scope Those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, many equivalents to the specific embodiments in accordance with the invention described herein. The scope of the invention is not intended to be limited to the above Description, but rather is as set forth in the appended claims.

[0158] Furthermore, it is understood that any particular embodiment of the present invention that falls under the prior art can be expressly excluded from any one or more of the claims.Such an embodiment can be excluded even if the exclusion is not expressly provided herein, since it is considered to be known to those skilled in the art.Any particular embodiment of the composition of the present invention (e.g., any compound, any method of production, any method of use, etc.) can be excluded from one or more claims for any reason, whether related to the existence of prior art or not. EXAMPLES

[0159] Example 1. Evaluation of the effect of food on administration of sepiapterin method Subjects were orally administered sepiapterin (10 mg / kg) twice, one week apart, in fasted and fed states. Subjects were asked to consume a standard high-fat (approximately 50% of the total calorie content of the diet) and high-calorie (approximately 800-1000 calories) meal 30 minutes prior to receiving the second oral dose of sepiapterin on day 8.

[0160] Sampling for PK analysis was performed pre-dose (within 30 minutes before dosing) on ​​days 1 and 8 and 0.5, 1, 2, 4, 8, 12, and 24 hours after dosing on days 1 and 8.

[0161] Serum concentrations of sepiapterin and BH4 were analyzed by MNG Labs.

[0162] Cerebrospinal fluid (CSF) from selected subjects is collected by lumbar puncture approximately 30 minutes after the maximum observed plasma BH4 concentration (Tmax) as measured by blood analysis on Day 1 (pre-dose) and Day 7 (i.e., after 7 days of daily dosing).

[0163] Cerebrospinal fluid (CSF) was analyzed by MNG Labs. Descriptive statistics are provided to characterize any changes in neurotransmitter metabolism between day 1 and day 7 sample results.

[0164] result Surprisingly, the Cmax of plasma BH4 was much higher in subjects who were fed prior to dosing compared to subjects who were fasted prior to dosing, as shown in Tables 24 and 25 below and in Figure 1. Furthermore, when sepiapterin was administered in the fed vs. fasted state, a decrease in plasma sepiapterin concentrations was observed, but an increase in BH4 concentrations was observed (Figure 1). The geometric mean ratios of plasma sepiapterin concentrations (fasted / fasted, 90% CI) were significantly higher than the AUC last is 1.29 (0.84-2.00), C max The corresponding ratios (90% CI) for plasma BH4 were AUC 0-inf is 0.58 (0.47-0.71), C max The AUC0- was 0.55 (0.45-0.68). inf and AUC last Overall exposure of BH4, as measured by, was increased 1.7-fold when sepiapterin was administered in the fed state compared to the fasted state.

[0165] [Table 24]

[0166] [Table 25]

[0167] Furthermore, as shown in Tables 26 and 27 below, surprisingly, the Cmax of sepiapterin in plasma was much lower in subjects who were fed prior to dosing compared to subjects who were fasted prior to dosing.

[0168] [Table 26]

[0169] [Table 27]

[0170] Example 2. Comparison of Adverse Events in Fed and Fasted Subjects method Twelve subjects received a single dose of sepiapterin (10 mg / kg) under fasted conditions followed 7 days later under fed conditions. Standard definitions of adverse events (AEs) were used. All-cause AEs were those occurring at any time, and treatment-emergent adverse events (TEAEs) were those occurring at the time of or after study drug administration. TEAEs related to study drug were based on the opinion of the investigator. Serious AEs were defined as life-threatening or resulting in death, hospitalization or prolongation of existing hospitalization, or persistent or significant disability / incapacity, or substantial disruption of the ability to perform normal life functions, or congenital anomaly / birth defect.

[0171] result As shown in Table 28, surprisingly, there was a reduced incidence of adverse events when sepiapterin was administered to fed subjects compared to fasted subjects.

[0172] [Table 28]

[0173] Example 3. Pharmacokinetic analysis of multiple doses of sepiapterin method Three cohorts of eight fed subjects were randomized in a 6:2 ratio to receive either sepiapterin or placebo once daily for 7 days. A sentinel dosing strategy was also used to administer the highest dose of sepiapterin.

[0174] result Plasma time concentrations of sepiapterin and BH4 were similar after 1 and 7 days of treatment with sepiapterin, and there was no accumulation of drug. Pharmacokinetic data are shown in Table 29 below.

[0175] [Table 29]

[0176] Example 4. Reduction of phenylalanine blood concentration in PKU patients by administration of sepiapterin After screening for eligibility, a 7-day run-in period was initiated. Patients then received either PTC923 20 mg / kg / day ("low dose"), or PTC923 60 mg / kg / day or sapropterin dihydrochloride orally once daily for 7 days in a randomized order. PTC923 (formerly CNSA-001) is an oral sepiapterin. Each patient was randomly assigned to one of six treatment sequences (Figure 2). Randomization with six different treatment sequences was intended to minimize carryover effects, treatment period effects, patient fatigue, and natural fluctuations in blood Phe levels. There was a 7-day washout period between each active treatment period. Patients were asked to maintain their usual pre-study diet, including intake of an amino acid mixture if prescribed, and were monitored by a dietician throughout the study period.

[0177] Blood samples for Phe were collected 7, 5, 3, and 1 days prior to randomization. During the treatment period, blood samples were collected pre-dose on day 1 and post-dose on days 3, 5, and 7. During the washout period, blood samples for Phe were collected on days 1, 3, 5, and 7. All samples were collected at the same time of day, either fasting or >3 hours postprandial. Blood Phe was measured at a central bioanalytical laboratory (Agilex Biolabs Pty Ltd) using a validated LC-MS-MS technique for dried blood sampling by Volumetric Absorptive Microsampling (VAMS®) using a Mitra® 4 sampler clamshell.

[0178] Twenty-four patients were randomized, all of whom completed the study and were included in both the efficacy and safety populations. The mean age of the patients was 26 years, and the mean weight was 69 kg (Table 30). Two-thirds of the patients were female (67%). All patients were sapropterin treatment naive. Eleven of the 24 patients (46%) had classical PKU, defined as any documented past measurement of blood Phe of at least 1,200 μmol / L. Supplements containing amino acids for management of PKU were noted to have been received by 13 patients (54%) before the study and 14 patients (58%) during the study.

[0179] [Table 30]

[0180] All patients entered the study on a low-protein, Phe-restricted diet and were instructed to maintain their daily protein intake throughout the study. Mean [SD] daily dietary Phe intake was similar for the 3 groups at baseline (PTC923 60 mg / kg, 2,550 [1,371] mg; PTC923 20 mg / kg, 2,265

[0946] mg; sapropterin dihydrochloride 20 mg / kg, 2,510 [1,395] mg). The mean change in daily dietary Phe intake by day 7 increased in all groups, by 19 [1,260] mg, 650 [1,262] mg, and 523 [1,132] mg, respectively.

[0181] The safety of PTC923 was monitored and adverse events (AEs) were coded using MedDRA® (Medical Dictionary for Regulatory Activities) version 21.0. Treatment-emergent adverse events (TEAEs) were defined as AEs occurring at or after the first dose of study drug. TEAEs were related to the current study treatment or the last study treatment prior to the event. Treatment-related TEAEs were those that, in the investigator's judgment, were possibly, probably, or definitely related to treatment. Serious AEs (SAEs) were defined as those resulting in death, life-threatening AE, hospitalization or prolongation of existing hospitalization, persistent or significant disability / incapacity, substantial disruption of the ability to perform normal life functions, or congenital anomaly / birth defect.

[0182] result PTC923 significantly reduced blood Phe from baseline at both doses (Table 31). PTC923 at 60 mg / kg was significantly more effective at lowering blood Phe than sapropterin dihydrochloride at 20 mg / kg (p=0.0098). The effect of PTC923 at 20 mg / kg on blood Phe was numerically greater than that of sapropterin dihydrochloride at 20 mg / kg, with the lower dose of PTC923 producing a mean absolute reduction in plasma phenylalanine of approximately 100 micromoles / liter and PTC923 producing a mean absolute reduction in plasma phenylalanine of more than 200 micromoles / liter, as shown in FIG. 1.

[0183] [Table 31]

[0184] Least squares mean (LSM) changes (SE) from baseline blood Phe levels were -206.4 (41.8) μmol / L (p<0.0001) for PTC923 60 mg / kg, -146.9 (41.8) μmol / L (p=0.0010) for PTC923 20 mg / kg, and -91.5 (41.7) μmol / L (p=0.0339) for sapropterin dihydrochloride (Figure 3). The effect of PTC923 appeared to be dose-related.

[0185] The proportion of patients randomized to PTC923 60 mg / kg who achieved blood Phe < 360 μmol / L was 12 / 24 (50%), 11 / 24 (46%), and 10 / 24 (42%) randomized to PTC923 20 mg / kg and sapropterin dihydrochloride, respectively. The proportion of patients randomized to either dose of PTC923 who achieved blood Phe < 360 μmol / L was 13 / 24 (54%).

[0186] Blood Phe declined rapidly with study treatment: the least squares mean changes (SE; p-values) from baseline to the first Phe measurement after treatment initiation (day 3) in blood Phe were -206.6 (36.6; p<0.0001) μmol / L for PTC923 60 mg / kg, -167.5 (36.6; p<0.0001) μmol / L for PTC923 20 mg / kg, and -72.3 (36.6; p=0.0543) μmol / L for sapropterin dihydrochloride. Mean changes in blood Phe decline were significantly greater at day 3 for both PTC923 doses versus sapropterin dihydrochloride (LSM differences -131.3 [SE 33.8; p=0.0007] and -95.2 [SE 33.7; p=0.0135] for PTC923 60 mg / kg and PTC923 20 mg / kg, respectively) (Figure 4).

[0187] As shown in FIG. 5, PTC923 provided superior reduction in blood phenylalanine levels compared to sapropterin dihydrochloride in this study.

[0188] The least squares mean changes (SE; p-values) in blood Phe in 11 classical PKU patients were -150.8 (63.1; p=0.0287) μmol / L for PTC923 60 mg / kg, -71.5 (61.8; p=0.2629) μmol / L for PTC923 20 mg / kg, and -2.8 (62.0; p=0.9640) μmol / L for sapropterin dihydrochloride (Table 31). Comparison of PTC923 60 mg / kg vs. sapropterin dihydrochloride approached statistical significance (LSM difference -148.0 [SE 63.0; p=0.0566] μmol / L) (Figure 6).

[0189] A sensitivity analysis was performed excluding treatment periods for patients with baseline blood Phe <300 μmol / L. The least squares mean changes (SE; p-values) from baseline in blood Phe in the sensitivity population were -226.9 (44.2; p<0.0001) μmol / L for PTC923 60 mg / kg, -167.8 (45.2; p=0.0007) μmol / L for PTC923 20 mg / kg, and -105.5 (43.7; p=0.0211) μmol / L for sapropterin dihydrochloride 20 mg / kg. The mean reduction in blood Phe was significantly greater with PTC923 60 mg / kg vs. sapropterin dihydrochloride (LSM difference -121.4 [SE 42.9] mol / L, p=0.0146) (Figure 7).

[0190] Cofactor responder analysis was also performed in 19 patients with baseline Phe ≥ 300 μmol / L for all treatment periods. Twelve of the 19 patients (in both treatment groups) demonstrated a ≥ 20% reduction in blood Phe. The least squares mean changes (SE; p-values) from baseline in blood Phe for this group of 12 responders were -322.2 (60.0; p<0.0001) μmol / L for PTC923 60 mg / kg, -234.8 (61.2; p=0.0011) μmol / L for PTC923 20 mg / kg, and -139.70 (58.6; p=0.0277) μmol / L for sapropterin dihydrochloride. The mean change in blood Phe reduction was significantly greater with PTC923 60 mg / kg versus sapropterin dihydrochloride (LSM difference -182.5 [SE 62.0] μmol / L, p=0.0158) (Figure 8). The number of PTC923-treated patients with a ≥20% reduction in blood Phe was >50% greater than the number of sapropterin-treated patients. The absolute mean percent change and absolute Phe reduction for patients treated with PTC923 60 mg / kg who achieved at least a 30% reduction in Phe from baseline were -72.5% and -485.3 μmol / L, respectively.

[0191] Similar numbers of patients (PTC923 60 mg / kg / PTC923 20 mg / kg / sapropterin dihydrochloride) reported any adverse events (AEs; 29% / 25% / 21%), serious AEs (0% / 4% / 0%), and treatment-related AEs (13% / 0% / 0%). Headache was the most common all-cause AE (17% / 4% / 4%) and was transient in all cases.

[0192] No other individual AEs occurred in ≥1 patient. Most AEs were mild in severity (19 / 22, 86% of all AEs). Two patients reported moderate to severe AEs (conjunctivitis, dysmenorrhea) and one patient reported a serious AE (gastrointestinal reflux). None of these three moderate to severe AEs were related to treatment. There were no SAEs and no patients discontinued due to AEs. Changes in vital signs and ECG parameters were generally small and comparable between treatment groups.

[0193] The primary objective of the study was to evaluate the efficacy of PTC923, measured as the mean blood Phe change from baseline. The efficacy population consisted of all patients who were randomized and had blood Phe measurements obtained pretreatment and on days 3, 5, and 7 of treatment from at least one treatment period. The safety population consisted of all patients who were randomized and received any dose of study treatment. Sample size calculations were based on an all-case placebo-controlled regulatory study of sapropterin with an observed mean reduction in blood Phe of 99 μmol / L and standard deviation of 220 μmol / L, using a two-sided α of 0.05 to estimate a true mean reduction in blood Phe of 300 μmol / L with 80% power.

[0194] Responsiveness to PTC923 was assessed by measuring the mean change from baseline in blood Phe (calculated as the average of measurements on days 3, 5, and 7 of each treatment period). The predose blood Phe concentration on day 1 of each treatment period served as the individual patient-specific baseline for that period.

[0195] Changes from baseline in plasma Phe for each treatment period were modeled using a linear mixed model for repeated measures (MMRM). Efficacy models included fixed effects for baseline for each treatment period, treatment group, sequence, and period, and random subject effects within each sequence, with a first-order autoregressive AR(1) covariance structure. No unplanned covariates were included.

[0196] Pairwise comparisons of changes from baseline in Phe were performed between each PTC923 and sapropterin treatment by calculating least squares means using Dunnett's adjustment for multiple comparisons. The estimated change in blood Phe levels, its standard error (SE), and associated p-value were calculated for each pairwise comparison. Dunnett's adjustment was used to account for multiple comparisons of test groups against the control group. This adjustment ensured that the overall type I error rate under the null hypothesis was maintained despite multiple test groups being compared to a single control group.

[0197] The overall mean change in blood Phe for each treatment was calculated and the standard deviation (SD) was determined.

[0198] A prespecified sensitivity analysis excluded treatment periods with baseline Phe < 300 μmol / L, and a responder analysis examined patients with a baseline of ≥ 300 μmol / L who responded to sapropterin and / or PTC923 treatment with a ≥ 20% decline.

[0199] Data were analyzed using SAS v.9.4.

[0200] Summary Surprisingly, PTC923 showed statistically significant superiority over sapropterin dihydrochloride in reducing blood phenylalanine levels, even in a study that included only 24 subjects. Furthermore, approximately 45% of subjects with classical PKU responded to PTC923, lowering phenylalanine levels by 20% or more. This is surprising, considering that historically, only about 10% of subjects with classical PKU responded to treatment with sapropterin dihydrochloride. PTC923 also showed a surprisingly high overall level of responders compared to sapropterin dihydrochloride. Compared to treatment with sapropterin dihydrochloride, 60% of subjects responded to PTC923, with 3-4 times more subjects normalizing (i.e., blood phenylalanine levels below 120 micromoles / liter on day 7). Indeed, subjects receiving PTC923 were 13.45 times more likely to achieve normal blood phenylalanine levels (i.e., less than 120 micromoles per liter) compared with subjects receiving sapropterin dihydrochloride. Notably, 46% of non-classical subjects treated with PTC923 achieved normalization compared with 15.4% of those treated with sapropterin dihydrochloride. Subjects receiving PTC923 also reported improved sleep quality and improved focus.

Claims

1. A pharmaceutical composition comprising sepiapterin or a pharma- ceutical acceptable salt thereof for use in treating phenylketonuria in a subject having a blood phenylalanine concentration of greater than 120 micromoles / liter, wherein an effective amount is administered to the subject, the subject having failed to respond to treatment with sapropterin or a pharma- ceutical acceptable salt thereof.

2. The pharmaceutical composition of claim 1, wherein the subject is on a phenylalanine restricted diet.

3. 3. The pharmaceutical composition of claim 1 or 2, wherein the subject has a blood phenylalanine concentration greater than 600 micromoles / liter.

4. The pharmaceutical composition of any one of claims 1 to 3, wherein the subject has not responded to treatment with pegvalase-pqpz.

5. The pharmaceutical composition of any one of claims 1 to 4, wherein the subject has discontinued treatment with pegvalase-pqpz due to adverse reactions and / or tolerability.

6. 6. The pharmaceutical composition of any one of claims 1 to 5, wherein said administration reduces the subject's blood phenylalanine concentration to less than 360 micromoles / liter.

7. 7. The pharmaceutical composition of claim 6, wherein said administration reduces the subject's blood phenylalanine concentration to less than 120 micromoles / liter.

8. The pharmaceutical composition according to any one of claims 1 to 7, wherein the administration reduces the subject's blood phenylalanine concentration by at least 35% compared to the blood phenylalanine concentration before administration of sepiapterin or a pharma- ceutical acceptable salt thereof.

9. 9. The pharmaceutical composition of any one of claims 1 to 8, wherein said administration reduces the subject's blood phenylalanine concentration to less than 360 micromoles / liter at a phenylalanine intake of at least about 1500 mg / day.

10. 10. The pharmaceutical composition of any one of claims 1 to 9, wherein said administration results in a BH4 concentration of at least 50 ng / ml in the plasma of said subject within 10 hours after administration of sepiapterin or a pharma- ceutical acceptable salt thereof.

11. The pharmaceutical composition according to any one of claims 1 to 10, wherein the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is about 20 mg / kg to about 60 mg / kg per administration.

12. The pharmaceutical composition according to any one of claims 1 to 11, wherein the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is about 20 mg / kg per dose.

13. The pharmaceutical composition according to any one of claims 1 to 11, wherein the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is about 40 mg / kg per dose.

14. The pharmaceutical composition according to any one of claims 1 to 11, wherein the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is about 60 mg / kg per dose.

15. The pharmaceutical composition according to any one of claims 1 to 14, wherein said effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is administered once a day.

16. The pharmaceutical composition according to any one of claims 1 to 14, wherein said effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is administered twice a day.

17. 17. The pharmaceutical composition of claim 16, wherein the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is administered in two equivalent amounts.

18. The pharmaceutical composition according to any one of claims 1 to 17, wherein the effective amount of sepiapterin or a pharma- ceutically acceptable salt thereof is administered together with food.

19. 20. The pharmaceutical composition of claim 18, wherein the administration to the subject is less than 30 minutes before or after ingesting food.

20. 20. The pharmaceutical composition of claim 18, wherein administration to the subject is substantially simultaneous with food.

21. The pharmaceutical composition according to any one of claims 18 to 20, wherein the food is a high-protein and / or high-fat food.

22. The pharmaceutical composition according to any one of claims 18 to 21, wherein the food is a high-calorie food.

23. 23. The pharmaceutical composition of any one of claims 1 to 22, wherein the risk of adverse events is reduced compared to a subject administered at least 10 mg / kg of sapropterin or a pharma- ceutically acceptable salt thereof.

24. 24. The pharmaceutical composition of any one of claims 1 to 23, wherein said administration results in an increase in neurocognitive function in said subject.

25. The pharmaceutical composition according to any one of claims 1 to 24, wherein the subject is a child.

26. 26. The pharmaceutical composition of claim 25, wherein the child is under 7 years of age.

27. The pharmaceutical composition of any one of claims 1 to 25, wherein the subject is over 7 years old.

28. The pharmaceutical composition of any one of claims 1 to 27, wherein the subject has been diagnosed with tetrahydrobiopterin-responsive phenylketonuria.

29. The pharmaceutical composition of any one of claims 1 to 28, wherein the subject has been diagnosed with sepiapterin-responsive phenylketonuria.

30. 30. The pharmaceutical composition of any one of claims 1 to 29, wherein the sepiapterin or a pharma- ceutically acceptable salt thereof is formulated as an oral powder for suspension.

31. 31. The pharmaceutical composition of any one of claims 1 to 30, wherein the sepiapterin or a pharma- ceutically acceptable salt thereof is administered as a suspension in a flavored suspension vehicle.

32. 32. The pharmaceutical composition of any one of claims 1 to 31, wherein the subject has a blood phenylalanine concentration greater than 1200 micromoles / liter.

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