Gannesolone for treatment of nodular sclerosis

Abstract

The present application relates to ganaolone for the treatment of nodular sclerosis, and in particular, to a method for the treatment of nodular sclerosis or epilepsy associated with nodular sclerosis comprising administering to a subject in need thereof a therapeutically effective amount of a pharmaceutically acceptable pregnenolone neurosteroid, such as ganaolone, or a pharmaceutically acceptable pregnenolone neurosteroid, such as ganaolone, or a pharmaceutically acceptable pregnenolone neurosteroid or a pharmaceutically acceptable pregnenolone neurosteroid or a pharmaceutically acceptable pregnenolone neurosteroid or a pharmaceutically acceptable pregnenolone neurosteroid. The composition can be used for relieving one or more symptoms of the nodular sclerosis or epilepsy related to the nodular sclerosis.

Description

[0001] This invention is a divisional application of the PCT patent application entered into China with Chinese patent application number 202080083001.0, invention name “Ganexolone for the treatment of tuberous sclerosis complex” and international application date December 7, 2020.

[0002] This application claims the benefit of U.S. Provisional Application No. 62 / 944,549, filed on December 6, 2019, the entire contents of which are incorporated herein by reference. Background Art

[0003] Tuberous sclerosis, also known as tuberous sclerosis complex (TSC), is a rare, multisystem genetic disorder characterized by the growth of numerous noncancerous (benign) tumors in many parts of the body (Northrup et al., (2013), Pediatr Neurol., 49(4):243-254). These tumors may occur in the skin, brain, kidneys, and other organs, and in many cases can cause serious health problems. Ibid.

[0004] Epilepsy is the most common neurological symptom in patients with TSC ("TSC-associated epilepsy") and is a significant cause of morbidity and mortality. Jülich and Sahin (2014), Pediatric Neurol., 50:290-296. Infantile spasms are the most common type of seizure presenting in infancy and represent the first manifestation of epilepsy in 50% of patients. Ibid. In older children and adults, focal impaired perception seizures (formerly called complex partial seizures) are the most common. Ibid. Other focal and generalized seizures may also occur. Ibid.

[0005] TSC-related epilepsy affects up to about 90% of patients, of which about 70% are resistant to treatment (Portocarrero et al., (2018), An Bras Dermatol., 93(3): 323-331). In addition, compared with patients with controlled epilepsy, patients with refractory epilepsy have a higher prevalence of intellectual disability (such as autism, mental retardation, and mood disorders), and disease manifestations persist throughout the life of affected individuals. Same as above. The negative impact of epilepsy on cognitive development and quality of life makes the prevention and management of epileptic seizures an important goal in the treatment of TSC (Vergeer et al., (2019), Epilepsia Open, 4: 581-592).

[0006] There is no cure for this disease. The first-line treatment for TSC-related infantile spasms is vigabatrin (Uliel-Sibony et al. (2020), Child's Nervous System, 36:2511-2517). However, vigabatrin is associated with serious adverse side effects. Ibid. For example, it causes irreversible retinal damage in approximately 21-34% of patients. Other potential side effects include brain abnormalities in the thalamus, basal ganglia, brainstem tegmentum, and cerebellar dentate nucleus, hyperkinetic movement disorders, and acute encephalopathy. Ibid. In addition, no other anti-seizure medication can significantly reduce the severity or incidence of TSC-related epilepsy.

[0007] Therefore, there is a significant unmet need for effective therapies to treat TSC. Summary of the invention

[0008] The present disclosure relates to methods for treating TSC and / or TSC-related epilepsy. As described above, TSC and / or TSC-related epilepsy is an extremely complex and challenging disease to treat. In 70% of cases, patients are resistant to treatment and epilepsy is refractory. Refractory epilepsy often causes significant behavioral and developmental consequences. As described and exemplified herein, the inventors believe that ganaxolone can provide an effective therapy for TSC and / or TSC-related epilepsy.

[0009] Given the complexity and difficulty of treating TSC and TSC-related epilepsy, the inventors believe that effective treatment requires high doses of drugs (e.g., ganaxolone). For example, in a clinical trial for focal seizures in adults, ganaxolone was administered at a daily dose of 1800 mg (administered twice a day at 900 mg), but the clinical endpoint was not achieved. The inventors have now unexpectedly discovered that ganaxolone administered at a lower daily dose but more frequently may provide an effective therapy for TSC and TSC-related epilepsy. Without wishing to be bound by any particular theory or mechanism, it is believed that more frequent administration of ganaxolone (e.g., three times a day) but at a lower daily dose than previously used may improve drug exposure by maintaining ganaxolone serum levels (e.g., trough levels) above a threshold level for a longer period of time, thereby providing effective treatment. For example, administration of oral ganaxolone three times daily (or more) and a total daily dose of no more than 1800 mg, or no more than 1700 mg, or no more than 1600 mg, or no more than 1500 mg, or no more than 63 mg / kg of ganaxolone per day can produce a ganaxolone plasma concentration of at least about 100 ng / ml over a 24-hour day for about 70% or more and provide effective seizure reduction. In addition, less ganaxolone is generally required to achieve increased ganaxolone trough concentrations with three times daily (or more frequent) administration, which is also beneficial to the subject being treated. In fact, trough levels of ganaxolone with twice daily dosing generally remain below 100 ng / ml over a 24-hour treatment period.

[0010] The inventors have also unexpectedly discovered that a subpopulation of patients with TSC-related epilepsy have low plasma concentrations of allopregnanolone-sulfate (Allo-S) (eg, less than 2,500 pg / ml) and may respond better to ganaxolone treatment.

[0011] Thus, the present disclosure relates to methods for effectively treating TSC and or TSC-related epilepsy. The methods disclosed herein include administering to a subject in need thereof a therapeutically effective amount of a neurosteroid, preferably ganaxolone, or a pharmaceutically acceptable salt thereof. Ganaxolone is preferably administered in an amount that provides a ganaxolone trough level (e.g., ganaxolone plasma concentration) of about 100 ng / ml or more over a 24-hour day by about 70% or more.

[0012] To achieve a ganaxolone plasma concentration of about 100 ng / ml or greater by about 70% or more within a 24-hour day, ganaxolone may be administered three times daily (or more) up to a maximum of about 1,800 mg per day. In subjects weighing less than 40 kg, ganaxolone may be administered three times daily (or more) up to a maximum of 63 mg / kg per day. Typically, up to about 1,500 mg per day of ganaxolone when administered three times a day may produce a plasma concentration of about 100 ng / ml or greater by about 70% or more within a 24-hour day. Ganaxolone may be administered three times a day at a dose of about 500 mg. A skilled clinician will appreciate that the amount of ganaxolone administered three times a day (e.g., orally) may be adjusted to achieve a desired ganaxolone trough level as long as the total amount does not exceed the maximum daily dose of ganaxolone.

[0013] Preferably, ganaxolone is administered orally (e.g., as an oral suspension or oral capsule). Without being bound by theory, the inventors believe that three doses may provide increased anti-seizure activity (i.e., reduced seizure frequency and / or seizure severity) due to increased plasma ganaxolone exposure. This is in contrast to previous treatment regimens that teach administration of ganaxolone at high doses (e.g., daily doses) to achieve therapeutic efficacy. For example, ganaxolone is administered twice a day at higher doses.

[0014] Administration of ganaxolone in an amount that achieves a ganaxolone plasma concentration of at least about 100 ng / ml or greater over a 24 hour period by about 70% or more can reduce the frequency of seizures and / or the severity of seizures in a subject relative to baseline. Typically, a reduction in seizure frequency of at least about 20% or greater relative to baseline seizure frequency can be achieved. During treatment, the plasma concentration of ganaxolone in the subject can be monitored and / or the subject can be monitored for seizure activity using EEG. If the subject develops signs of a seizure (e.g., a recurrence of seizure), the amount of ganaxolone administered can be adjusted accordingly.

[0015] The methods disclosed herein are suitable for treating any form of epileptic seizure associated with TSC or TSC-related epilepsy, such as, but not limited to, infantile spasms, focal impaired awareness seizures, focal seizures, or generalized seizures.

[0016] It is an object of the present disclosure to provide treatments for tuberous sclerosis complex. It is an object of the present disclosure to provide treatments for epilepsy associated with tuberous sclerosis complex (TSC). It is another object of the present disclosure to provide treatments for seizures associated with TSC-associated epilepsy. It is another object of the present disclosure to provide therapeutic benefits to humans with TSC and TSC-associated epilepsy using the gamma-aminobutyric acid (GABA)ergic mechanism of action of ganaxolone.

[0017] In furtherance of the above objectives and other objectives, the present disclosure relates in part to a method of treating a human suffering from tuberous sclerosis, the method comprising administering to the human a therapeutically effective amount of a pharmaceutically acceptable pregnenolone neurosteroid in an amount effective to alleviate or reduce one or more symptoms of tuberous sclerosis in the human. The pharmaceutically acceptable pregnenolone neurosteroid can be administered parenterally and / or orally in an amount of about 1 mg per day to about 5000 mg per day. Humans particularly likely to benefit from the administration of a pharmaceutically acceptable pregnenolone neurosteroid are those with low levels of allopregnanolone-sulfate.

[0018] Plasma levels of allopregnanolone-sulfate appear to be positively correlated with plasma levels of allopregnanolone and may be qualitatively representative of allopregnanolone levels in the brain. Therefore, low levels of allopregnanolone-sulfate may indicate a lack of allopregnanolone in the brain. Plasma allopregnanolone-sulfate levels of about 2500 pg / ml or less are considered low and may indicate a lack of endogenous neurosteroids in the body. Low levels of allopregnanolone-sulfate may be 2400 pg / ml or less, 2300 pg / ml or less, 2200 pg / ml or less, 2100 pg / ml or less, 2000 pg / ml or less, 1900 pg / ml or less, 1800 pg / ml or less, 1700 pg / ml or less, 1600 pg / ml or less, 1500 pg / ml or less, 1400 pg / ml or less, or 1500 pg / ml or less. pg / ml or less, 1300 pg / ml or less, 1200 pg / ml or less, 1100 pg / ml or less, 1000 pg / ml or less, 900 pg / ml or less, 850 pg / ml or less, 800 pg / ml or less, 750 pg / ml or less, 700 pg / ml or less, 650 pg / ml or less, 600 pg / ml or less, 550 pg / ml or less , 500pg / ml or less, 450pg / ml or less, 400pg / ml or less, 350pg / ml or less, 300pg / ml or less, 250pg / ml or less, 200pg / ml or less, 150pg / ml or less, 100pg / ml or less, 90pg / ml or less, 80pg / ml or less, 70pg / ml or less, 60pg / ml or less, 50pg / ml or lower, 40pg / ml or lower, 30pg / ml or lower, 20pg / ml or lower, 15pg / ml, 10pg / ml or lower, 9pg / ml or lower, 8pg / ml or lower, 7pg / ml or lower, 6pg / ml or lower, 5pg / ml or lower, 4pg / ml or lower, 3pg / ml or lower, 2pg / ml or lower, 1pg / ml or lower.

[0019] Because the lack of allopregnanolone in the brain may cause one or more symptoms of tuberous sclerosis, in the methods disclosed herein, the administration of a pharmaceutically acceptable pregnenolone neurosteroid according to the methods disclosed herein can correct this lack, and thus alleviate and / or reduce the severity of one or more symptoms of tuberous sclerosis and / or reduce the frequency of the symptoms. Symptoms of tuberous sclerosis that can be alleviated or alleviated by the administration of a pharmaceutically acceptable pregnenolone neurosteroid include, but are not limited to, for example, epileptic seizures, intellectual disability, developmental delay, behavioral problems, skin abnormalities, lung disease, and nephropathy. Epileptic seizures may include, for example, focal motor seizures with no consciousness or impaired perception, focal seizures with impairment of consciousness or perception, focal seizures that evolve into bilateral generalized seizures, tonic-clonic seizures, and generalized seizures with countable motor components, including, for example, tonic-clonic, bilateral tonic, bilateral clonus, or atonic / drop seizures.

[0020] The present disclosure also relates to a method of treating a human suffering from tuberous sclerosis, the method comprising administering to the human a therapeutically effective amount of ganaxolone in an amount effective to alleviate or reduce one or more symptoms of tuberous sclerosis in the human. Ganaxolone can be administered parenterally and / or orally. Humans particularly well-positioned to benefit from the administration of ganaxolone are those with low levels of allopregnanolone-sulfate.

[0021] When ganaxolone is administered orally, a therapeutically effective amount of ganaxolone can be, for example, about 600 mg per day to about 2000 mg per day. In certain embodiments, the dosage can be increased to about 2100 mg per day, 2200 mg per day, 2300 mg per day, or higher to provide a better response than lower doses, the limiting factor being the increased side effects at higher doses. Ganaxolone is typically administered up to about 1,500 mg per day or 1,800 mg per day. In certain embodiments, the dosage can be reduced to about 550 mg per day, 500 mg per day, 450 mg per day, 300 mg per day, or less than 300 mg per day to mitigate or reduce the severity of side effects experienced by a person at higher doses. Symptoms of tuberous sclerosis that can be alleviated and / or can have their frequency and / or severity reduced by administering ganaxolone according to the methods disclosed herein include, for example, seizures, intellectual disability, developmental delays, behavioral problems, skin abnormalities, lung disease, and kidney disease. Seizures can include, for example, focal motor seizures without impaired consciousness or perception, focal seizures with impaired consciousness or perception, focal seizures that evolve into bilateral generalized seizures, tonic-clonic seizures, and generalized seizures with a countable motor component, including, for example, tonic-clonic, bilateral tonic, bilateral clonic, or atonic / drop seizures.

[0022] The present disclosure also relates to a method of treating a human suffering from TSC or TSC-related epilepsy, the method comprising chronically administering to the human a pharmaceutically acceptable pregnenolone neurosteroid (e.g., ganaxolone) in an amount effective to reduce the frequency of seizures in the human, wherein the human has low plasma levels of endogenous neurosteroids (e.g., allopregnanolone-sulfate (Allo-S) as outlined above).

[0023] The present disclosure is also directed to a method of treating a human suffering from TSC or TSC-related epilepsy, the method comprising orally administering to the human twice a day (e.g., every 10-13 hours) a solid oral immediate release formulation comprising a pharmaceutically acceptable pregnenolone neurosteroid (e.g., ganaxolone), wherein the neurosteroid has a half-life of about 18 hours to about 24 hours, the formulation releases not less than about 70% or about 80% of the ganaxolone when placed in simulated gastrointestinal fluid (SGF and / or SIF) for 45 minutes, and the administration reduces the frequency of seizures per 28 days in the human by at least about 35%, about 40%, about 45%, or about 50% compared to the frequency of seizures during the 28-day period prior to the first administration.

[0024] The present disclosure is also directed to a method of treating a human suffering from TSC or TSC-related epilepsy, the method comprising orally administering to the human three times a day (e.g., every 6 to 8 hours) a liquid oral immediate release formulation comprising a pharmaceutically acceptable pregnenolone neurosteroid (e.g., ganaxolone), wherein the neurosteroid has a half-life of about 18 hours to about 24 hours, the formulation releases not less than about 70% or about 80% of the ganaxolone when the formulation is placed in simulated gastrointestinal fluid (SGF and / or SIF) for 45 minutes, and the administration reduces the frequency of seizures per 28 days in the human by at least about 35%, about 40%, about 45%, or about 50% compared to the frequency of seizures during the 28 day period prior to the first administration.

[0025] The present disclosure also relates to a method of treating a human being with a pregnenolone neurosteroid, wherein the human being suffers from TSC or TSC-related epilepsy, the method comprising the steps of: determining whether the human being has a low level of endogenous neurosteroids (e.g., Allo-S) by obtaining or having obtained a biological sample (e.g., a blood sample) from the human being; and conducting or having conducted an assay on the biological sample to determine the plasma level of the endogenous neurosteroid in the biological sample. A level of endogenous neurosteroids of 2500 pg mL-1 or less, 2000 pg mL-1 or less, 1500 pg mL-1 or less, 1000 pg mL-1 or less, 900 pg mL-1 or less, 800 pg mL-1 or less, 700 pg mL-1 or less, 600 pg mL-1 or less, 500 pg mL-1 or less, 400 pg mL-1 or less, 300 pg mL-1 or less, 200 pg mL-1 or less, 100 pg mL-1 or less, 75 pg mL-1 or less, 50 pg mL-1 or less, or 25 pg mL-1 or less indicates that the human has low levels of endogenous steroids. A subject (e.g., a human) with low levels of endogenous steroids (e.g., Allo-S) can be orally administered a pregnenolone neurosteroid (e.g., ganaxolone) at a dosage of 1 mg / kg to about 63 mg / kg per day, about 2 mg / kg to about 63 mg / kg per day, about 3 mg / kg to about 63 mg / kg per day, about 4 mg / kg to about 63 mg / kg per day, about 5 mg / kg to about 63 mg / kg per day, about 6 mg / kg to about 63 mg / kg per day, or about 7 mg / kg to about 63 mg / kg per day for at least one day in two or three divided doses.

[0026] In some embodiments of these embodiments, 2500 pg mL-1 or less, 2000 pg mL-1 or less, 1500 pg mL-1 or less, 1000 pg mL-1 or less, 900 pg mL-1 or less, 800 pg mL-1 or less, 700 pg mL-1 or less, 600 pg mL-1 or less, 500 pg mL-1 or less, 400 pg mL-1 or less, 300 pg mL-1 or less, 200 pg mL-1 or less, 100 pg mL-1 or less, 75 pg mL-1 or less, 50 pg mL-1 or less, A level of endogenous neurosteroids of 25 pg mL-1 or less, or 25 pg mL-1 or less, indicates that administration of ganaxolone may reduce the patient's seizure frequency after 28 days of administration, e.g., by 35% or more; about 40% or more; about 45% or more; or about 50% or more, compared to the seizure frequency during the 28 days prior to the first administration. Endogenous neurosteroids may be selected from the group comprising or consisting of pregnanolone, pregnanolone-sulfate, 5-αDHP, allopregnanolone, allopregnanolone-S, pregnanolone, pregnanolone-S, DHEA, and combinations thereof; and the pregnenolone neurosteroid may, for example, be selected from the group comprising or consisting of allopregnanolone, ganaxolone, alphaxalone, alphadolone, hydroxydione, minaxolone, pregnanolone, acebrochol, or tetrahydrocorticosterone, and pharmaceutically acceptable salts thereof. In some of these embodiments, the method further comprises communicating the results of the assay to the patient or medical provider before or after administration of the pregnenolone neurosteroid. Ganaxolone is a preferred pregnenolone neurosteroid.

[0027] The present disclosure is also directed to a method of treating a human with ganaxolone, wherein the human suffers from TSC or TSC-related epilepsy, the method comprising the steps of determining whether the human has an allopregnanolone-sulfate (Allo-S) level of 2500 pg mL-1 or less, and if the human has an allopregnanolone-sulfate level of 2500 pg mL-1 or less, orally administering ganaxolone to the human at a dose of 1 mg / kg per day to about 63 mg / kg per day, about 2 mg / kg per day to about 63 mg / kg per day, about 3 mg / kg per day to about 63 mg / kg per day, about 4 mg / kg per day to about 63 mg / kg per day, about 5 mg / kg per day to about 63 mg / kg per day, about 6 mg / kg per day to about 63 mg / kg per day, or about 7 mg / kg per day to about 63 mg / kg per day, for at least one day, divided into two or three doses. In some of these embodiments, a level of allopregnanolone sulfate of 2500 pg mL-1 or less indicates that administration of ganaxolone may reduce seizure frequency in a human, e.g., by at least about 35%, about 40%, about 45%, or about 50%, after 28 days of administration, compared to the frequency of seizures during the 28 days prior to the first administration.

[0028] The present disclosure also relates to a method for treating a human suffering from TSC or TSC-related epilepsy, the method comprising the steps of determining whether the human has a level of allopregnanolone-sulfate of 2500 pg mL-1 or less, and if the human has a level of allopregnanolone-sulfate of 2500 pg mL-1 or less, orally administering to the human an endogenous neurosteroid (e.g., allopregnanolone, pregnanolone, etc.) or a synthetic neurosteroid (e.g., Co26749 / WAY-141839, Co134444, Co177843, Sage-217 (3α-hydroxy-3β-methyl-21-(4-cyano-1H-pyrazol-1'-yl)-19-nor-5β-pregnane-20-one), ganaxolone, etc.) at a dose of 1 mg / day. kg to about 200 mg / kg per day, about 2 mg / kg per day to about 150 mg / kg per day, about 3 mg / kg per day to about 100 mg / kg per day, about 4 mg / kg per day to about 90 mg / kg per day, about 5 mg / kg per day to about 80 mg / kg per day, about 6 mg / kg per day to about 70 mg / kg per day, or about 7 mg / kg per day to about 65 mg / kg per day for at least one day, divided into two or three doses, and if the human has a level of allopregnanolone-sulfate exceeding 2500 pg mL-1, avoid administering endogenous or synthetic neurosteroids to the human and / or administering a different anticonvulsant. The different anticonvulsant can, for example, be selected from the group consisting of: benzodiazepines. Benzodiazepines (e.g., clobazam, diazepam, clonazepam, midazolam, etc.), chlordiazepoxide Acid (clorazepic acid), levetiracetam, felbamate, lamotrigine, fatty acid derivatives (e.g., valproic acid), carboxamide derivatives (rufinamide, carbamazepine, oxcarbazepine, etc.), amino acid derivatives (e.g., levocarnitine), barbiturates (e.g., phenobarbital), or a combination of two or more of the foregoing agents. Any number of other anticonvulsants may be administered. Those skilled in the art will be familiar with anticonvulsants.

[0029] The present disclosure is also directed to a method for treating a human suffering from TSC or TSC-related epilepsy, the method comprising the steps of determining whether the human has an allopregnanolone-sulfate level of 2500 pg mL-1 or less, and if the human has an allopregnanolone-sulfate level of 2500 pg mL-1 or less, orally administering ganaxolone to the human at a dose of 1 mg / kg per day to about 63 mg / kg per day, about 2 mg / kg per day to about 63 mg / kg per day, about 3 mg / kg per day to about 63 mg / kg per day, about 4 mg / kg per day to about 63 mg / kg per day, about 5 mg / kg per day to about 63 mg / kg per day, about 6 mg / kg per day to about 63 mg / kg per day, or about 7 mg / kg per day to about 63 mg / kg per day, for at least one day, divided into two or three doses. In some of these embodiments, a level of allopregnanolone sulfate of 2500 pg mL-1 or less indicates that administration of ganaxolone may reduce seizure frequency in a human, e.g., by at least about 35%, about 40%, about 45%, or about 50%, after 28 days of administration, compared to the frequency of seizures during the 28 days prior to the first administration.

[0030] The present disclosure is also directed to a method for treating a human with TSC or TSC-related epilepsy, the method comprising the steps of determining whether the human has an allopregnanolone level of 200 pg mL-1 or less, and if the human has an allopregnanolone level of 200 pg mL-1 or less, orally administering ganaxolone to the human at a dose of 1 mg / kg per day to about 80 mg / kg per day, about 2 mg / kg per day to about 75 mg / kg per day, about 3 mg / kg per day to about 70 mg / kg per day, about 4 mg / kg per day to about 65 mg / kg per day, about 5 mg / kg per day to about 63 mg / kg per day, about 6 mg / kg per day to about 63 mg / kg per day, or about 7 mg / kg per day to about 63 mg / kg per day, for at least one day, divided into two or three doses, and avoiding administration of ganaxolone to the human if the human has an allopregnanolone level of more than 200 pg mL-1. In some of these embodiments, a level of allopregnanolone of 200 pg mL-1 or less after 28 days of administration, compared to the frequency of seizures during the 28-day period prior to the first administration, indicates that administration of ganaxolone may reduce seizure frequency in a human, e.g., by at least about 35%, about 40%, about 45%, or about 50%.

[0031] The methods disclosed herein may also include the step of measuring the plasma level of allopregnanolone in a human suffering from TSC or TSC-related epilepsy. An allopregnanolone plasma level of about 200 pg / ml or less is a low level and may indicate that the human may be deficient in endogenous neurosteroids. Thus, in some embodiments, low endogenous neurosteroid levels in a human can be, for example, 200 pg / ml or less, 199 pg / ml or less, 198 pg / ml or less, 197 pg / ml or less, 196 pg / ml or less, 195 pg / ml or less, 194 pg / ml or less, 193 pg / ml or less, 192 pg / ml or less, 191 pg / ml or less, 190 pg / ml or less, 189 pg / ml or less, 188 pg / ml or less, 187 pg / ml or less, 186 pg / ml or less, 185 pg / ml or less, 184 pg / ml or less, 183pg / ml or less, 182pg / ml or less, 181pg / ml or less, 180pg / ml or less, 179pg / ml or less, 178pg / ml or less, 177pg / ml or less, 176pg / ml or less, 175pg / ml or less, 174pg / ml or less, 173pg / ml or less, 172pg / ml or less, 171pg / ml or less, 170pg / ml or less, 169pg / ml or less, 168pg / ml or less, 167pg / ml or less, 166pg / ml or less, 168pg / ml or less, pg / ml or less, 154pg / ml or less, 153pg / ml or less, 152pg / ml or less, 151pg / ml or less, 150pg / ml or less, 149pg / ml or less, 148pg / ml or less, 149pg / ml or less, 150pg / ml or less, 151pg / ml or less, 152pg / ml or less, 153pg / ml or less, 154pg / ml or less, 155pg / ml or less, 156pg / ml or less, 157pg / ml or less, 158pg / ml or less, 159pg / ml or less, 160pg / ml or less, 161pg / ml or less, 162pg / ml or less, 163pg / ml or less, 164pg / ml or less, 165pg / ml or less, 166pg / ml or less, 167pg / ml or less, 168pg / ml or less, 169pg / ml or less, 170pg / ml or less, 171pg / ml or less, 46pg / ml or less, 145pg / ml or less, 144pg / ml or less, 143pg / ml or less, 142pg / ml or less, 141pg / ml or less, 140pg / ml or less, 139pg / ml or less, 138pg / ml or less, 137pg / ml or less, 136pg / ml or less, 135pg / ml or less, 134pg / ml or less, 133pg / ml or less, 132pg / ml or less, 131pg / ml or less, 130pg / ml or less, 129pg / ml or less, 128pg / ml or less,127pg / ml or less, 126pg / ml or less, 125pg / ml or less, 124pg / ml or less, 123pg / ml or less, 122pg / ml or less, 121pg / ml or less, 120pg / ml or less, 119pg / ml or less, 118pg / ml or less, 117pg / ml or less, 116pg / ml or less, 115pg / ml or less, 114pg / ml or less, 113pg / ml or less, 112pg / ml or less, 111pg / ml or less, 110pg / ml or less, 109pg / ml or less, 108pg / ml or less, 107pg / ml or less pg / ml or less, 94pg / ml or less, 93pg / ml or less, 92pg / ml or less, 91pg / ml or less, 90pg / ml or less, 89pg / ml or less, 88pg / ml or less, 87pg / ml or less, 86pg / ml or less, 85pg / ml or less, 86pg / ml or less, 87pg / ml or less, 88pg / ml or less, 89pg / ml or less, 89pg / ml or less, 81pg / ml or less, 81pg / ml or less, 82pg / ml or less, 83pg / ml or less, 84pg / ml or less, 85pg / ml or less, 86pg / ml or less, 87pg / ml or less, 88pg / ml or less, 89pg / ml or less, 81pg / ml or less, 82pg / ml or less, 83pg / ml or less, 84pg / ml or less, 85pg / ml or less pg / ml or less, 84pg / ml or less, 83pg / ml or less, 82pg / ml or less, 81pg / ml or less, 80pg / ml or less, 79pg / ml or less, 78pg / ml or less, 77pg / ml or less, 76pg / ml or less, 75pg / ml or less, 74pg / ml or less, 73pg / ml or less, 72pg / ml or less, 71pg / ml or less, 70pg / ml or less, 69pg / ml or less, 68pg / ml or less, 67pg / ml or less, 66pg / ml or less, 65pg / ml or less, 64pg / ml or less, 63pg / ml or less, 62pg / ml or less / ml or less, 61pg / ml or less, 60pg / ml or less, 59pg / ml or less, 58pg / ml or less, 57pg / ml or less, 56pg / ml or less, 55pg / ml or less, 54pg / ml or less, 53pg / ml or less, 52pg / ml or less, 51pg / ml or less, 50pg / ml or less, 49pg / ml or less, 48pg / ml or less, 47pg / ml or less, 46pg / ml or less, 45pg / ml or less, 44pg / ml or less, 43pg / ml or less, 42pg / ml or less, 41pg / ml or less, 40pg / ml or less,39pg / ml or less, 38pg / ml or less, 37pg / ml or less, 36pg / ml or less, 35pg / ml or less, 34pg / ml or less, 33pg / ml or less, 32pg / ml or less, 31pg / ml or less, 30pg / ml or less, 29pg / ml or less, 28pg / ml or less, 27pg / ml or less, 26pg / ml or less, 25pg / ml or less, 24pg / ml or less, 23pg / ml or less, 22pg / ml or less, 21pg / ml or less, 20pg / pg / ml or less, 19pg / ml or less, 18pg / ml or less, 17pg / ml or less, 16pg / ml or less, 15pg / ml or less, 14pg / ml or less, 13pg / ml or less, 12pg / ml or less, 11pg / ml or less, 10pg / ml or less, 9pg / ml or less, 8pg / ml or less, 7pg / ml or less, 6pg / ml or less, 5pg / ml or less, 4pg / ml or less, 3pg / ml or less, 2pg / ml or less, 1pg / ml or less, or 0pg / ml.

[0032] The present disclosure is also directed to a method for treating TSC or TSC-related epilepsy, the method comprising the steps of determining whether the human has an allopregnanolone level of 200 pg mL-1 or less, and if the human has an allopregnanolone level of 200 pg mL-1 or less, orally administering ganaxolone to the human at a dose of 1 mg / kg per day to about 100 mg / kg per day, about 2 mg / kg per day to about 80 mg / kg per day, about 3 mg / kg per day to about 70 mg / kg per day, about 4 mg / kg per day to about 65 mg / kg per day, about 5 mg / kg per day to about 65 mg / kg per day, about 6 mg / kg per day to about 65 mg / kg per day, or about 7 mg / kg per day to about 65 mg / kg per day for at least one day.

[0033] The present disclosure also relates to a method of treating endogenous neurosteroid deficiency in a human in need thereof, the method comprising administering a pharmaceutically acceptable pregnenolone neurosteroid (e.g., ganaxolone) to the human at a dose of about 1800 mg or less per day for at least 1 day, wherein the human has a genetic mutation in the TSC1 gene located on chromosome 9q34 and / or the TSC2 gene located on chromosome 16p13.3, and has one or more symptoms selected from the group consisting of: depigmented macules (≥3, straight diameter of at least 5 mm), angiofibroma (≥3) or fibrous plaques, onychofibroma (≥2), shark spots, multiple retinal hamartomas, cortical dysplasia, subependymal nodules, subependymal giant cell astrocytoma, cardiac rhabdomyoma, lymphangioleiomyomatosis (LAM), angiomyolipoma (≥2), "confetti" skin lesions, enamel pits (≥3), oral fibroma (≥2), retinal achromatic plaques, multiple renal cysts, and non-renal hamartomas.

[0034] In some of these embodiments, the pharmaceutically acceptable pregnenolone neurosteroid is ganaxolone and is administered orally in an amount of about 200 mg per day to about 2500 mg per day, about 200 mg per day to about 2250 mg per day, about 200 mg per day to about 2000 mg per day, about 300 mg per day to about 1800 mg per day, about 400 mg per day to about 1800 mg per day, about 450 mg per day to about 1800 mg per day, about 675 mg per day to about 1800 mg per day, about 900 mg per day to about 1800 mg per day, about 1125 mg per day to about 1800 mg per day, about 1350 mg per day to about 1800 mg per day, about 1575 mg per day to about 1800 mg per day, or about 1800 mg per day, in two or three divided doses. In some embodiments, the human experiences seizures and administration of a pharmaceutically acceptable pregnenolone neurosteroid reduces the average seizure frequency per 28 days by 35% or better (e.g., about 40%, about 45%, about 50%, about 55%) compared to the frequency of seizures during a 28-day period prior to the first administration. In some embodiments, the improvement is 50% or more.

[0035] The methods disclosed herein may also include periodic measurement of the plasma levels of the administered pharmaceutically acceptable pregnenolone neurosteroid (e.g., ganaxolone) and / or concomitant AED medication (if any) and / or allopregnanolone (3α-hydroxy-5α-pregnane-20-one) and / or related endogenous CNS active steroids. In some embodiments, the plasma levels of liver enzymes (AST, ALT, and ALK Phos) are also measured before, during, or after treatment with a pharmaceutically acceptable pregnenolone neurosteroid. Plasma levels may be measured, for example, once a week, every 2 weeks, every 3 weeks, every 4 weeks, every 5 weeks, every 6 weeks, every 7 weeks, every 8 weeks, every 9 weeks, every 10 weeks, every 11 weeks, or every 12 weeks.

[0036] In the methods described herein, the pregnenolone neurosteroid (e.g., ganaxolone) can be administered orally or parenterally for a long term. In certain preferred embodiments, the pregnenolone neurosteroid is ganaxolone and is administered as an oral suspension or oral solid dosage form (e.g., oral capsule) at a dose of up to a total of 63 mg / kg per day, and preferably a maximum amount of up to 1800 mg of ganaxolone is administered per day. Preferably, ganaxolone is administered for a long term, e.g., as long as the patient is obtaining a therapeutic benefit from treatment without adverse side effects requiring cessation of treatment. In certain embodiments, ganaxolone is administered for at least one day, at least 2 days, at least 3 days, 2 weeks, at least 3 weeks, at least 4 weeks, at least 5 weeks, at least 6 weeks, at least 7 weeks, at least 8 weeks, at least 9 weeks, at least 10 weeks, at least 11 weeks, or at least 12 weeks. In some embodiments, ganaxolone can be administered for a period of 2 weeks to 100 years, or for a person's lifetime.

[0037] When the pregnenolone neurosteroid is administered as an oral suspension, it can be administered, for example, any number of times between once and three times per day. In certain preferred embodiments, when the pregnenolone neurosteroid (e.g., ganaxolone) is administered orally, it can be administered with food (for better absorption) or without food. When the pregnenolone neurosteroid is administered as an oral tablet or capsule, it can be administered, for example, any number of times between once and four times per day. When the pregnenolone neurosteroid is administered parenterally, it can be administered, for example, any number of times between once and about three times per day or as needed.

[0038] The present disclosure also relates in part to an immediate release formulation for TSC and / or TSC-related epilepsy comprising particles comprising (i) a pregnenolone neurosteroid (e.g., ganaxolone) and (ii) one or more pharmaceutically acceptable excipients (e.g., an oral suspension, tablet, or capsule), wherein the particles have a particle size that ensures that they do not agglomerate after dispersion in simulated gastrointestinal fluid (SGF and / or SIF) and do not change after the formulation is stored at 25°C / 60% RH for 1 month. In preferred embodiments, the formulation releases not less than about 70% or about 80% of the pregnenolone neurosteroid when the formulation is placed in 500 ml of dissolution medium (e.g., 5% SLS in SGF (simulated gastric fluid) and / or 5% SLS in SIF (simulated intestinal fluid)) at 37°C + 0.5°C at 100 rpm in USP apparatus 1 (basket), and provides a plasma level of pregnenolone neurosteroid of about 55 ng / mL, about 60 ng / ml, or about 65 ng / ml to a plasma level of pregnenolone neurosteroid of about 240 ng / ml to 400 ng / ml (e.g., 262 ng / mL) for a period of at least about 6 hours, about 7 hours, about 8 hours, about 9 hours, about 10 hours, or about 12 hours after single and / or multiple dose administration. In some of these embodiments, the volume weighted median diameter of the particles is about 250 nm to about 450 nm (e.g., about 332 nm). In some embodiments, the particles have a D(10) particle size of about 200 nm to about 220 nm, a D(50) particle size of about 250 nm to about 450 nm, and a D(90) particle size of about 480 nm to about 700 nm, and the formulation is free of cyclodextrins, including sulfoalkyl ether cyclodextrins and modified forms thereof, and is used to treat TSC-related epilepsy.

[0039] The present disclosure is also directed, in part, to an oral immediate release formulation for TSC and / or TSC-related epilepsy comprising particles comprising (i) ganaxolone and (ii) one or more pharmaceutically acceptable excipients (e.g., an oral suspension, tablet, or capsule), wherein the particles have an average particle size of about 0.3 microns (i.e., a volume weighted median diameter (D50) of about 0.3 microns); the particle size does not change after the formulation is stored at 25°C / 60% RH for 1 month; and the particle size does not change after the formulation is placed in 500 ml of dissolution medium (e.g., 5% SLS in SGF (simulated gastric fluid) and / or 5% SIF (simulated intestinal fluid) at 37°C+0.5°C in USP Apparatus 1 (basket) at 100 rpm. The formulation releases no less than about 70% or about 80% of the ganaxolone at 45 minutes in a 5% slurry (SLS); the formulation provides a plasma level of ganaxolone of about 55 ng / mL, about 60 ng / ml, or about 65 ng / ml to a plasma level of about 240 ng / ml to about 400 ng / ml (e.g., 262 ng / mL) after a single dose and / or multiple doses, which persists for at least 6 hours to 12 hours after administration, and is useful for treating TSC or TSC-related epilepsy. The formulation can provide a plasma level of ganaxolone of about 55 ng / mL, about 60 ng / ml, or about 65 ng / ml to a plasma level of about 240 ng / ml to 400 ng / ml (e.g., 262 ng / mL) after administration on an empty stomach and / or after feeding.

[0040] In some of these embodiments, an average particle size of about 0.3 microns is critical to providing not less than about 70% or about 80% dissolution of pregnenolone neurosteroid when the formulation is placed in simulated gastrointestinal fluid (SGF and / or SIF) for 45 minutes, and plasma levels of pregnenolone neurosteroid of about 55 ng / mL, about 60 ng / ml, or about 65 ng / ml to plasma levels of pregnenolone neurosteroid of about 240 ng / ml to 400 ng / ml (e.g., 262 ng / mL) for a period of at least about 6 hours, about 7 hours, about 8 hours, about 9 hours, about 10 hours, or about 12 hours.

[0041] The present disclosure also relates in part to an immediate release formulation for TSC and / or TSC-related epilepsy comprising particles comprising (i) ganaxolone and (ii) one or more pharmaceutically acceptable excipients (e.g., an oral suspension, tablet, or capsule), wherein the particles have an average particle size of about 0.3 microns; the particle size does not change after the formulation is stored at 25°C / 60% RH for 2 months and / or 3 months and / or 4 months; and the particle size does not change after the formulation is placed in 500 ml of dissolution medium (e.g., 5% SLS in SGF (simulated gastric fluid) and / or 5% SIF (simulated intestinal fluid) at 37°C+0.5°C in USP Apparatus 1 (basket) at 100 rpm. The formulation releases not less than 80% of the ganaxolone at 45 minutes in SLS; the formulation provides a plasma level of ganaxolone of about 55 ng / mL, about 60 ng / ml, or about 65 ng / ml to a plasma level of about 240 ng / ml to about 400 ng / ml (e.g., 262 ng / mL) for at least 6 hours to 12 hours after administration. BRIEF DESCRIPTION OF THE DRAWINGS

[0042] Figure 1 Baseline endogenous sex-specific allopregnenolone-sulfate ("Allo-S") levels in female patients with PCDH19-related epilepsy, stratified by responders versus non-responders, are depicted. When stratified for patients with Allo-S levels <2.5 ng mL-1, a mean reduction of 50% in seizure frequency (n=7) compared with baseline was observed. This analysis was performed retrospectively in a small open-label cohort. However, the 1.5-2 order of magnitude difference in allopregnenolone-sulfate between responders and non-responders suggests that plasma levels of allopregnenolone-sulfate may be used to predict the efficacy of pharmaceutically acceptable pregnenolone neurosteroids. These data provide preliminary evidence that plasma levels of allopregnenolone-sulfate may be used as a predictive biomarker to prospectively identify patients who may experience enhanced treatment effects with ganaxolone.

[0043] Figure 2Based on plasma samples from the Biosample Repository, allopregnanolone-sulfate (Allo-S) levels in TSC patients versus control patients were profiled. The goal was to quantify endogenous neurosteroid levels using our proprietary validated analytical method (LC / MS / MS) and compare levels to healthy (unaffected) age-matched control samples. Plasma samples from TSC epilepsy patients showed a trend toward reduced Allo-S levels (n=47, median 1.8 ng mL-1) compared to controls (n=60, median 4.1 ng mL-1). This finding was strengthened when only patients / subjects aged 1-14 years were sampled (TSC n=28, control n=28). Patients / subjects included 29 females and 18 males. The median age of females was 15 years (range 2-27 years). The median age of males was 10.5 years (range 2-33 years). Fluctuations in neurosteroid levels after puberty may confound the analysis in all patients.

[0044] Figure 3 The distribution of Allo-S in TSC patients and unaffected subjects (all individuals) is depicted.

[0045] Figure 4 A comparison of PCDH19 and allopregnanolone-sulfate (Allo-S) in TSC and potential expansion opportunities to explore Allo-S biomarkers in TSC are depicted.

[0046] Figure 5 A positive correlation between allopregnanolone (Allo) and allopregnanolone-sulfate (Allo-S) is depicted. DETAILED DESCRIPTION

[0047] Effective treatment of TSC and / or TSC-related epilepsy has been challenging, with conventional treatment options ineffective for many patients. Indeed, approximately 70% of patients are resistant to treatment, and epilepsy is intractable. Most seizures begin within the first 12 months of life, and the prevalence of intellectual disability is high as a result. Seizure control could reduce this developmental consequence. Therefore, improved methods for the treatment of TSC and / or TSC-related epilepsy are urgently needed.

[0048] The present disclosure relates to new methods for treating SE. As illustrated and described herein, treatment according to the method provides a reduction in the frequency of seizures and / or inhibition of seizures in TSC-related epilepsy. The method can be used to treat any form of TSC-related epilepsy. For example, but not limited to, infantile spasms, focal motor seizures without impaired consciousness or perception, focal seizures with impaired consciousness or perception, focal seizures that evolve to bilateral, tonic-clonic seizures, and generalized seizures. Motor seizures include countable tonic-clonic, bilateral tonic, bilateral clonus, atonic / drop seizures, myoclonic seizures, or epileptic seizures.

[0049] The methods described herein comprise administering to a subject a therapeutically effective amount of a neurosteroid. Ganaxolone is a preferred neurosteroid.

[0050] The methods described herein may also include administering ganaxolone in a therapeutically effective amount to achieve a ganaxolone plasma concentration of 100 ng / ml or greater by about 70% or more within a 24-hour day. This may be achieved by administering ganaxolone at least three times a day. Three times a day is preferred, although in some cases, ganaxolone may be suitably administered more than three times a day to achieve a desired trough concentration of ganaxolone. A plasma concentration of at least about 100 ng / ml or greater within a 24-hour day of about 70% or more provides for a reduction in seizures and / or improved seizure suppression. For example, a reduction in seizure frequency of at least 20% or greater relative to baseline seizure frequency may be achieved. A lower maximum dose of ganaxolone administered more than three times daily may achieve a desired trough concentration of ganaxolone. Typically, a maximum daily dose of ganaxolone of about 1,800 mg, preferably 1,500 mg, is administered. The maximum daily dose of ganaxolone is administered at the same or different doses over at least three time intervals within a 24-hour day.

[0051] The methods disclosed herein may also include determining whether a patient with TSC or TSC-related epilepsy will benefit from neurosteroid (e.g., ganaxolone) treatment. A subpopulation of patients with TSC-related epilepsy have low plasma concentrations of Allo-S (e.g., less than 2,500 pg / ml) and may respond better to ganaxolone treatment. The methods described herein may include measuring the level of endogenous neurosteroids in a subject before starting treatment with a neurosteroid (e.g., ganaxolone). Low levels of endogenous neurosteroids may indicate that the subject will respond to treatment with neurosteroids. Once it is determined that a subject has low endogenous neurosteroid levels, a therapeutically effective amount of a neurosteroid may be administered to the subject.

[0052] Additional descriptions of the methods and guidance for practicing the methods are provided herein. For ease of illustration, further details and guidance are provided regarding preferred aspects of using ganaxolone. Further details and guidance are also intended to relate to treatment with other neurosteroids.

[0053] I. Definitions

[0054] Unless otherwise indicated herein, the enumeration of value ranges is intended only to be used as a shorthand method for individually referring to each individual value belonging to the scope, and each individual value is incorporated into this specification as if individually enumerated herein. The endpoints of all scopes are included in the scope and can be independently combined. Unless otherwise indicated herein or clearly contradictory to the context, all methods described herein can be carried out in a suitable order. The use of any and all examples or exemplary language (e.g., "such as") is intended only to be illustrated, and unless otherwise required, will not impose limitations on the scope of the present invention. The language in this specification should not be interpreted as indicating any unrequired element as necessary for practicing the present invention.

[0055] The terms "a" or "an" do not denote a limitation of quantity, but rather denote the presence of at least one of the referenced item.

[0056] The term "about" is used synonymously with the term "approximately". As will be understood by one of ordinary skill in the art, the exact boundaries of "about" will depend on the components of the composition. As an example, the term "about" is used to indicate a value slightly beyond the quoted value, i.e., ±0.1% to 10%, which is also safe and effective. Therefore, compositions slightly beyond the quoted range are also included within the scope of the claims of the present invention.

[0057] "Active agent" is any compound, element or mixture that directly or indirectly produces a physiological effect on the patient when it is applied to the patient alone or in combination with another agent. When the active agent is a compound, it includes a salt, a solvate (including a hydrate) of a free compound or a salt, a crystalline and non-crystalline form, and various polymorphs of the compound. The compound may contain one or more asymmetric elements, such as a stereogenic center, a stereoaxis, etc., such as an asymmetric carbon atom, so that the compound can exist in different stereoisomeric forms. These compounds can be, for example, racemates or optically active forms. For compounds with two or more asymmetric elements, these compounds can also be mixtures of diastereomers. For compounds with an asymmetric center, it should be understood that all pure forms of optical isomers and their mixtures are covered. In addition, compounds with carbon-carbon double bonds can exist in Z-form and E-form, and all isomeric forms of the compound are included in the present invention. In these cases, a single enantiomer, i.e., an optically active form, can be obtained by asymmetric synthesis, synthesis from an optically pure precursor, or by resolution of a racemate. Resolution of racemates may also be achieved, for example, by conventional methods such as crystallization in the presence of a resolving agent or chromatography using, for example, a chiral HPLC column.

[0058] The term "endogenous neurosteroids" means steroids that are produced in the brain and are capable of regulating neuronal excitability by interacting with neuronal membrane receptors and ion channels (mainly GABA-A receptors), and include, for example, pregnane neurosteroids (e.g., allopregnanolone, allotetrahydrodeoxycorticosterone, etc.), androstane neurosteroids (e.g., androstanediol, bencholanolone, etc.), and sulfated neurosteroids (e.g., pregnanolone sulfate, dehydroepiandrosterone sulfate (DHEAS)).

[0059] The term "pregnenolone neurosteroid" means an endogenous or exogenous steroid capable of modulating neuronal excitability by interacting with neuronal membrane receptors and ion channels, primarily GABA-A receptors, and encompasses endogenous neurosteroids and synthetic neurosteroids synthesized or derived from pregnenolone, e.g., in vitro and in vivo.

[0060] The term "biomarker" refers to serum or plasma levels of neurosteroids that distinguish drug responders from non-responders.

[0061] As disclosed herein, the terms "serum" and "plasma" may be used interchangeably.

[0062] The terms "comprising," "including," and "containing" are non-limiting. Other elements not recited may be present in the embodiments claimed by these transitional phrases. Where "comprising," "including," or "comprising" are used as transitional phrases, other elements may be included and still form an embodiment within the scope of the claims. The open transitional phrase "comprising" encompasses the intermediate transitional phrase "consisting essentially of" and the closed transitional phrase "consisting of."

[0063] A "bolus dose" is a relatively large dose of a drug administered over a short period of time, such as over 1 to 30 minutes.

[0064] “C max ” is the concentration of the active agent in plasma at the point of maximum concentration.

[0065] "Ganexolone" is also known as 3α-hydroxy-5α-pregnane-20-one, or "GNX" herein.

[0066] "Infusion" administration is non-oral administration, typically intravenous administration, but in some embodiments includes other non-oral routes, such as epidural administration. Infusion administration is performed over a longer period of time than bolus administration, for example, for a period of at least 15 minutes, at least 30 minutes, at least 1 hour, at least 2 hours, at least 3 hours, or at least 4 hours.

[0067] A "patient" is a human or non-human animal in need of medical treatment. Medical treatment includes treatment of an existing condition, such as a disorder or injury. In certain embodiments, treatment also includes prophylactic or preventive treatment, or diagnostic treatment.

[0068] "Child" means a human between 1 day and 18 years of age (eg, 1 day to 15 years of age), inclusive.

[0069] "Adult" means a person over the age of 18.

[0070] A "pharmaceutical composition" is a composition comprising at least one active agent, such as a compound of formula (I) or a salt, solvate or hydrate, and at least one other substance, such as a carrier. A pharmaceutical composition optionally contains one or more additional active agents. When specified, a pharmaceutical composition complies with the GMP (Good Manufacturing Practice) standards of the U.S. FDA for human or non-human drugs. A "pharmaceutical combination" is a combination of at least two active agents, which may be combined in a single dosage form or provided together in separate dosage forms with instructions for using the active agents together to treat a condition, such as an epileptic seizure condition.

[0071] "Polyvidone", also known as povidone and polyvinylpyrrolidone (PVP), is a water-soluble polymer made from the monomer N-vinylpyrrolidone. Plasdone C-12 and C-17 are pharmaceutical grade homopolymers of N-vinylpyrrolidone. Plasdone C-12 has a K value of 10-2-13.8 and a nominal molecular weight of 4000d. Plasdone C-17 has a K value of 15.5-17.5 and a nominal molecular weight of 10,000d.

[0072] "Sterilization" means the inactivation of substantially all biological contaminants in a sample, preparation or product. For most pharmaceutical applications, a 1 million-fold reduction in bioburden is also considered "sterile".

[0073] The term "reduction" of seizures or seizure activity refers to a detectable decrease in the frequency, severity, and / or duration of seizures. The reduction in the frequency, severity, and / or duration of seizures can be measured by self-assessment (e.g., by patient report) or by a trained clinical observer. The reduction in the frequency, severity, and / or duration of seizures can be determined by comparing the patient's status before and after treatment.

[0074] A "therapeutically effective amount" or "effective amount" is the amount of an agent that achieves a pharmacological effect. The term "therapeutically effective amount" includes, for example, a prophylactically effective amount. An "effective amount" of a neurosteroid is the amount required to achieve the desired pharmacological effect or therapeutic improvement without undue side effects. One skilled in the art will select an effective amount of a neurosteroid based on the specific patient and disease. It should be understood that an "effective amount" or "therapeutically effective amount" may vary between subjects due to variations in the metabolism of the neurosteroid, age, weight, general condition of the subject, the disorder being treated, the severity of the disorder being treated, and the judgment of the prescribing physician.

[0075] "Treat" or "treatment" refers to any treatment of a disorder or disease, such as inhibiting the disorder or disease, such as arresting the development of the disorder or disease, relieving the disorder or disease, causing regression of the disorder or disease, alleviating the affliction caused by the disease or disease, or alleviating the symptoms of the disease or disease.

[0076] "Alkyl" is a branched or straight chain saturated aliphatic hydrocarbon radical having a specific number of carbon atoms, typically 1 to about 8 carbon atoms. As used herein, the term C1-C6 alkyl refers to an alkyl having 1, 2, 3, 4, 5, or 6 carbon atoms. Other embodiments include alkyls having 1 to 6 carbon atoms, 1 to 4 carbon atoms, or 1 or 2 carbon atoms, such as C1-C8 alkyl, C1-C4 alkyl, and C1-C2 alkyl. Examples of alkyls include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, 3-methylbutyl, tert-butyl, n-pentyl, and sec-pentyl.

[0077] "Aryl" means an aromatic group containing only carbon in one or more aromatic rings. Typical aryl groups contain 1 to 3 separate, fused or side chain rings and 6 to about 18 ring atoms, with no heteroatoms as ring members. When indicated, such aryl groups may be further substituted with carbon or non-carbon atoms or groups. Aryl groups include, for example, phenyl, naphthyl, including 1-naphthyl, 2-naphthyl and biphenyl. An "arylalkyl" substituent is an aryl as defined herein, which is connected to the group it replaces via an alkylene linker. Alkylene is an alkyl as described herein, except that it is divalent.

[0078] "Cycloalkyl" is a saturated hydrocarbon ring radical having a specified number of carbon atoms. Monocyclic cycloalkyls typically have 3 to about 8 carbon ring atoms or 3 to 6 (3, 4, 5, or 6) carbon ring atoms. The cycloalkyl substituents may be pendant from the substituted nitrogen, oxygen, or carbon atom, or the substituted carbon atom may have two substituents with the cycloalkyl attached in a spiro group. Examples of cycloalkyls include cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl.

[0079] "Heteroalkyl" is an alkyl group as described wherein at least one carbon is replaced with a heteroatom (eg, N, O, or S).

[0080] As used herein, the term "substituted" means that any one or more hydrogens on a designated atom or group are selected from the indicated group, provided that the normal valence of the designated atom is not exceeded. When the substituent is an oxo group (i.e., =O), 2 hydrogens on the atom are replaced. When an oxo group replaces a heteroaromatic moiety, the resulting molecule can sometimes adopt a tautomeric form. For example, a pyridyl group substituted by an oxo group at the 2nd or 4th position can sometimes be written as pyridine or hydroxypyridine. Such combinations are only allowed when the combination of substituents and / or variables produces a stable compound or a suitable synthetic intermediate. A stable compound or a stable structure means a compound that is strong enough to withstand separation from a reaction mixture and is subsequently formulated into an effective therapeutic agent. Unless otherwise indicated, the substituent is named into the core structure. For example, it should be understood that aminoalkyl means that the point of attachment of the substituent to the core structure is in the alkyl portion, while alkylamino means that the point of attachment is a bond to the nitrogen of the amino group.

[0081] Suitable groups that may be present in a "substituted" or "optionally substituted" position include, but are not limited to, for example, halogen; cyano; -OH; oxo; -NH2; nitro; azido; alkanoyl (e.g., C2-C6 alkanoyl); C(O)NH2; alkyl (including cycloalkyl and (cycloalkyl)alkyl) groups having 1 to about 8 carbon atoms or 1 to about 6 carbon atoms; alkenyl and alkynyl groups, including groups having one or more unsaturated bonds and 2 to about 8 or 2 to about 6 carbon atoms; alkoxy groups having one or more oxygen bonds and 1 to about 8 or 1 to about 6 carbon atoms; aryloxy groups, such as phenoxy; alkylthio groups, including groups having one or more thioether bonds and 1 to about 8 carbon atoms or 1 to about 6 carbon atoms; alkylsulfinyl, including groups having one or more sulfinyl bonds and 1 to about 8 carbon atoms or 1 to about 6 carbon atoms; alkylsulfonyl, including groups having one or more sulfonyl bonds and 1 to about 8 carbon atoms or 1 to about 6 carbon atoms; aminoalkyl, including groups having one or more N atoms and 1 to about 8 or 1 to about 6 carbon atoms; monoalkylamino or dialkylamino, including groups having alkyl groups having 1 to about 6 carbon atoms; monoalkylaminocarbonyl or dialkylaminocarbonyl (i.e., alkylNHCO- or (alkyl1)(alkyl2)NCO-) having alkyl groups having about 1 to about 6 carbon atoms; aryl groups having 6 or more carbons.

[0082] II. Tuberous sclerosis

[0083] Tuberous sclerosis complex (TSC) is a multisystem disorder of embryonic cortical development that can affect many organs through the overgrowth of benign tumors called hamartomas. Combinations of symptoms may include seizures, intellectual disability, developmental delays, behavioral problems, skin abnormalities, lung disease, and kidney disease. While the disease phenotype of TSC varies greatly, up to 90% of TSC patients experience neurological manifestations such as epilepsy (Krueger et al., 2013). This condition is caused by inherited mutations in the TSC1 gene located on chromosome 9q34 or the TSC2 gene located on chromosome 16p13.3. The frequency of TSC is 1:6,000, and mutations have been found in 85% of patients (Jülich and Sahin, 2014). The gene products, hamartomatin (TSC1) and tuberin (TSC2), form a regulatory complex responsible for limiting the activity of mammalian target of rapamycin complex 1 (mTORC1), an important regulator of intracellular growth and metabolism, which it regulates by inhibiting the small GTPase brain-enriched Ras homolog (Rheb) (Krueger et al., 2013). Everolimus is an mTOR inhibitor that has been shown to reduce epileptic seizures (French et al., 2016; Mizuguchi et al., 2019).

[0084] TSC is one of the most common genetic causes of epilepsy, and symptoms of seizures vary depending on the age of onset (Jülich and Sahin, 2014). Effective treatment options include the use of adrenocorticotropic hormone (ACTH), which includes possible mechanisms consistent with the use of ganaxolone. ACTH has been shown to have a stimulatory effect on deoxycorticosterone (DOC), which undergoes further synthesis to form various neurosteroids. Specifically, ACTH has been shown to rapidly increase endogenous plasma and brain levels of allopregnanolone, which may further explain the beneficial effects on IS.

[0085] Infantile spasms (“IS”) are the most common type of seizure presenting in infancy and represent the first manifestation of epilepsy in 50% of patients. In older children and adults, focal impaired perception seizures (formerly known as complex partial seizures) are the most common. Other focal and generalized seizures may also occur, and more than 30% of patients develop refractory epilepsy (Jülich and Sahin, 2014). Although seizures are often attributed to the tubercular and perirhinal cortex, epilepsy in TSC can be considered multifactorial in origin, as seizures may originate from other brain regions or may occur in patients with TSC who do not have tubercular involvement (Jülich and Sahin, 2014).

[0086] The clinical diagnosis of TSC is clear, including two major features or one major feature plus ≥ 2 minor features listed in Table 7 below.

[0087] Table 7.

[0088]

[0089] *Includes nodules and radial migration lines of white matter

[0090] **Combination of two major clinical features (LAM and angiomyolipoma) without other features that do not meet the definite diagnostic criteria.

[0091] Gamma-aminobutyric acid (GABA) appears to play a central role in the development of TSC-related epilepsy, possibly due to endogenous GABA A Altered expression of receptor modulators (di Michele et al., 2003).

[0092] There is evidence to support that a deficiency of the neuroactive steroid 3α,5α-allopregnanolone (THP) or allopregnanolone is a factor in epileptogenesis in TSC. Allopregnanolone is a positive modulator of GABAA receptors and has been shown to have antiepileptic effects in experimental animals and humans. Allopregnanolone is reduced relative to its functional GABAA antagonist 3β-THP in patients with TSC-associated epilepsy, but not in TSC patients without epilepsy or in controls (di Michele et al., 2003). This reduced ratio may alter neuronal excitability mediated by GABAA receptors, leading to the development of epilepsy. The greater efficacy of vigabatrin, a specific and irreversible GABA aminotransferase inhibitor, in TSC-induced seizures relative to other epilepsy also supports a GABAA receptor-mediated role (di Michele et al., 2003).

[0093] TSC patients with epilepsy have reduced levels of endogenous neurosteroids, especially Allo, similar to previous reports in PCDH19. Approximately 25% of all TSC patients have plasma allopregnanolone-sulfate levels below 6 ng / ml.

[0094] Neuroactive steroids with anticonvulsant properties may be used to treat TSC and TSC-related epilepsy. Neuroactive steroids with anticonvulsant properties may increase GABAA-mediated signaling and may improve not only seizure control but also behavioral abnormalities in individuals with TSC and TSC-related epilepsy.

[0095] Upregulation of the TSC-mTOR pathway results in an increased inflammatory response, with evidence suggesting increased pro-inflammatory toll-like receptor 4 (TLR4) signaling. Neurosteroids, including Allo, have been shown to act as inhibitors of various neuroinflammatory pathways, including TLR4, which expands the mechanism of action of these compounds beyond positive regulation of GABAA receptors.

[0096] III. Neurosteroids

[0097] Endogenous neurosteroids play a key role in maintaining homeostasis of brain activity. Neurosteroids are able to rapidly produce brain changes in response to changes in the brain environment. Neurosteroids do not interact with classical steroid hormone receptors that regulate gene transcription; they primarily regulate brain excitability through interactions with neuronal membrane receptors and ion channels.

[0098] Depending on the chemical structure of the steroid molecule, neurosteroids can be GABA A Positive or negative regulator of receptor function (Pinna and Rasmussen, 2014, Reddy, 2003). GABA AReceptors mediate most synaptic inhibition in the CNS. Structurally, GABA A The receptor is a heteropentamer of five protein subunits that form a chloride channel. There are seven different classes of subunits, some of which have multiple homologous variants (α1-6, β1-3, γ1-3, σ1-3, δ, ε, θ); most GABA A The receptors are composed of alpha, beta, and either gamma or delta subunits. The neurotransmitter GABA activates the opening of chloride channels, allowing chloride ion influx and subsequent hyperpolarization. GABA A The receptors prevent the generation of action potentials by altering the depolarization produced by excitatory neurotransmission. A There are 2 types of receptor-mediated inhibitory neurotransmission: synaptic (phasic) and extrasynaptic (tonic) inhibition. Neurosteroids regulate synaptic and extrasynaptic GABA A receptors, thereby enhancing phasic and tonic currents. Phasic inhibition is due to the activation of γ2-containing receptors in synapses by intermittent release of millimolar concentrations of GABA from the axon terminals of presynaptic GABAergic interneurons. In contrast, tonic inhibition is mediated by sustained activation of δ-containing extrasynaptic receptors outside the synaptic cleft by low levels of ambient GABA that escape reuptake by GABA transporters. Tonic inhibition plays a unique role in controlling hippocampal excitability by setting a baseline for excitability (Reddy 2010).

[0099] Neurosteroids such as ganaxolone are GABA A A potent positive allosteric modulator of the GABA receptor (Akk et al., 2009). Neurosteroids enhance the A The first observation of a receptor-mediated GABA-evoked response was reported in 1984 with alfaxalone (Harrison and Simmonds, 1984). This modulatory effect of neurosteroids occurs through interaction with GABA A Binding occurs via discrete sites on the receptors, which are located within the transmembrane domains of the α- and β-subunits (Hosier et al., 2007; Hosier et al., 2009). The binding sites for benzodiazepines and barbiturates are different. Although the exact location of the neurosteroid binding site is still unknown, it has been shown that the highly conserved glutamine at position 241 in the α-subunit M1 domain plays a key role in neurosteroid regulation (Hosie et al., 2009). Class in their respective GABA A There are also differences in how they interact with GABA receptors. A receptor subtype, but benzodiazepine The class acts only on GABA containing γ2 subunits and no α4-subunits or α6-subunits A receptors (Lambert et al., 2003; Reddy, 2010). The specific α-subunit may affect the efficacy of neurosteroids, while the γ-subunit type may affect GABA A Efficacy and potency of neurosteroid modulation of receptors (Lambert et al., 2003).

[0100] Recent studies have shown that GABA A There are at least three neurosteroid binding sites on the receptor: one for allosteric enhancement of GABA-evoked currents by allopregnanolone at low (nM) concentrations, one for direct activation by allopregnanolone, and one for antagonism by sulfated neurosteroids such as pregnanolone sulfate (Lambert et al., 2003; Hosie et al., 2007). A The enhancement of chloride current in the receptor occurs through an increase in both the frequency and duration of channel opening (Reddy, 2010). Thus, neurosteroids greatly increase GABA A The possibility of the receptor chloride channel opening, thereby allowing a large influx of chloride ions, thus promoting the enhancement of inhibitory GABAergic transmission. These effects occur at physiological concentrations of neurosteroids. Therefore, endogenous neurosteroid levels continuously regulate GABA A Function of receptors (Reddy, 2010).

[0101] Extrasynaptic GABA containing delta-subunit A The receptors show increased sensitivity to neurosteroids, suggesting a key regulatory role in tonic inhibition (Wohlfarth et al., 2002). GABA containing delta subunits A The receptor is more sensitive to neurosteroid-induced enhancement of the GABA response (Stell et al., 2003). Mice lacking the δ subunit have a significantly reduced sensitivity to neurosteroids (Mihalek et al., 1999). The δ-subunit has no effect on the neurosteroid binding site but appears to enhance the conversion of neurosteroid action after neurosteroid binding to the receptor. GABA containing the δ subunit A The receptors have a low degree of desensitization, which helps mediate tonic GABA activation by ambient concentrations of GABA in the extracellular space A Receptor current. Tonic GABA A The receptor current leads to a stable inhibition of the neuron and reduces its excitability. GABA is a relatively inefficient δGABA Areceptor agonists, even though it binds with high affinity (Glykys and Mody, 2007). Therefore, even in the presence of saturating GABA concentrations, neurosteroids can significantly enhance the activity of neurons induced by δGABA A During neuronal activity, GABA is expected to be released in large quantities from active GABAergic interneurons, which can interact with perisynaptic and extrasynaptic GABA containing delta subunits. A Overall, the powerful effects of neurosteroids may be due to their effects on synaptic and perisynaptic / extrasynaptic GABA A The role of receptors (Reddy, 2010).

[0102] Pregnane and pregnenolone neurosteroids are a class of compounds that are useful as anesthetics, sedatives, hypnotics, anxiolytics, antidepressants, antitremors, treatments for autistic behaviors, and anticonvulsants. These compounds are characterized by very low water solubility, which limits their formulation options. Orally and parenterally bioavailable nanoparticle formulations of pregnane and pregnenolone neurosteroids are available.

[0103] Injectable formulations of pregnane neurosteroids and pregnenolone neurosteroids are particularly desirable because these compounds are used in clinical indications that preclude oral administration, such as anesthesia, and particularly for the emergency treatment of active epileptic seizures.

[0104] The present disclosure includes injectable nanoparticle neurosteroid formulations.

[0105] The pregnane neurosteroid and pregnenolone neurosteroid of the present invention may each be a compound of formula IA:

[0106]

[0107] or a pharmaceutically acceptable salt thereof, wherein:

[0108] X is O, S or NR 10 ;

[0109] R 1 is hydrogen, hydroxy, -CH2A, optionally substituted alkyl, optionally substituted heteroalkyl, optionally substituted aryl, or optionally substituted arylalkyl;

[0110] A is hydroxyl, O, S, NR 11 or an optionally substituted nitrogen-containing five-membered heteroaryl or an optionally substituted nitrogen-containing bicyclic heteroaryl or bicyclic heterocyclyl,

[0111] R 4 is hydrogen, hydroxy, oxo, optionally substituted alkyl or optionally substituted heteroalkyl,

[0112] R2 , R 3 , R 5 , R 6 and R 7 each independently absent, hydrogen, hydroxy, halogen, optionally substituted C1-C6 alkyl, optionally substituted C1-C6 alkoxy (e.g., methoxy), or optionally substituted heteroalkyl;

[0113] R 8 and R 9 are each independently selected from the group consisting of hydrogen, C1-C6 alkyl (e.g., methyl), halogenated C1-C6 alkyl (e.g., trifluoromethyl) or C1-C6 alkoxy (e.g., methoxy), or R 8 and R 9 forming an oxo group;

[0114] R 10 is hydrogen, hydroxy, optionally substituted alkyl, optionally substituted heteroalkyl, optionally substituted aryl, or optionally substituted arylalkyl, wherein each alkyl is C1-C 10 Alkyl, C3-C6 cycloalkyl, (C3-C6 cycloalkyl)C1-C4 alkyl, and optionally containing a single bond replaced by a double bond or a triple bond;

[0115] Each heteroalkyl group is an independently selected -O-, -S-, -N(R 10 )-, -S(=O)- or -S(=O)2-substituted alkyl group, wherein R 10 is hydrogen, alkyl, or alkyl in which one or more methylene groups are replaced by -O-, -S-, -NH, or -N-alkyl;

[0116] R 11 Is -H2 or -HR 12 ;

[0117] R 12 It is a C1-C6 alkyl group or a C1-C6 alkoxy group.

[0118] The pregnane neurosteroid and pregnenolone neurosteroid of the present invention can each be a compound of formula IA, wherein

[0119] X is O;

[0120] R 1 It is hydrogen, -CH3, -CH2OH, 1H-imidazol-1-yl, 1-oxoquinolin-6-yloxy and 4-cyano-1H-pyrazol-1'-yl.

[0121] R 4 is hydrogen, hydroxy, oxo, optionally substituted alkyl or optionally substituted heteroalkyl,

[0122] R 2 , R 3 , R 5 , R 6 and R 7 each independently absent, hydrogen, hydroxy, halogen, optionally substituted C1-C6 alkyl, optionally substituted C1-C6 alkoxy (e.g., methoxy), or optionally substituted heteroalkyl;

[0123] R 8 and R 9 are each independently selected from the group consisting of hydrogen, C1-C6 alkyl (e.g., methyl), halogenated C1-C6 alkyl (e.g., trifluoromethyl) or C1-C6 alkoxy (e.g., methoxy), or R 8 and R 9 forming an oxo group;

[0124] R 10 is hydrogen, hydroxy, optionally substituted alkyl, optionally substituted heteroalkyl, optionally substituted aryl, or optionally substituted arylalkyl, wherein each alkyl is C1-C 10 Alkyl, C3-C6 cycloalkyl, (C3-C6 cycloalkyl)C1-C4 alkyl, and optionally containing a single bond replaced by a double bond or a triple bond;

[0125] Each heteroalkyl group is an independently selected -O-, -S-, -N(R 10 )-, -S(=O)- or -S(=O)2-substituted alkyl group, wherein R 10 is hydrogen, alkyl, or alkyl in which one or more methylene groups are replaced by -O-, -S-, -NH, or -N-alkyl.

[0126] The pregnane neurosteroid and pregnenolone neurosteroid of the present invention can each be a compound of formula IB

[0127]

[0128] or a pharmaceutically acceptable salt thereof, wherein:

[0129] X is O, S or NR 10 ;

[0130] R 1 is hydrogen, hydroxy, optionally substituted alkyl, optionally substituted heteroalkyl, optionally substituted aryl, or optionally substituted arylalkyl;

[0131] R 4 is hydrogen, hydroxy, oxo, optionally substituted alkyl or optionally substituted heteroalkyl,

[0132] R 2 , R3 , R 5 , R 6 and R 7 each independently hydrogen, hydroxy, halogen, optionally substituted alkyl or optionally substituted heteroalkyl;

[0133] R 8 is hydrogen or alkyl and R 9 is hydroxyl; or

[0134] R 8 and R 9 Together they form an oxo group;

[0135] R 10 is hydrogen, hydroxy, optionally substituted alkyl, optionally substituted heteroalkyl, optionally substituted aryl, or optionally substituted arylalkyl, wherein each alkyl is C1-C 10 Alkyl, C3-C6 cycloalkyl, (C3-C6 cycloalkyl)C1-C4 alkyl, and optionally containing a single bond replaced by a double bond or a triple bond;

[0136] Each heteroalkyl group is an independently selected -O-, -S-, -N(R 10 )-, -S(=O)- or -S(=O)2-substituted alkyl group, wherein R 10 is hydrogen, alkyl, or alkyl in which one or more methylene groups are replaced by -O-, -S-, -NH, or -N-alkyl.

[0137] Compounds of Formulae IA and IB include, for example, allopregnanolone, ganaxolone, alfaxolone, alfadolone, hydroxypregnantone, minaxolone, pregnanolone, acebucloral or tetrahydrocorticosterone and pharmaceutically acceptable salts thereof.

[0138] The pregnane neurosteroid and pregnenolone neurosteroid of the present invention may also each be a compound of formula II:

[0139]

[0140] or a pharmaceutically acceptable salt thereof, wherein:

[0141] X is O, S or NR 10 ;

[0142] R 1 is hydrogen, hydroxy, -CH2A, optionally substituted alkyl, optionally substituted heteroalkyl, optionally substituted aryl, or optionally substituted arylalkyl;

[0143] A is hydroxyl, O, S, NR 11 or an optionally substituted nitrogen-containing bicyclic heteroaryl or bicyclic heterocyclyl,

[0144] R 4 is hydrogen, hydroxy, oxo, optionally substituted alkyl or optionally substituted heteroalkyl,

[0145] R 2 , R 3 , R 5 , R 6 and R 7 each independently absent, hydrogen, hydroxy, halogen, optionally substituted C1-C6 alkyl, optionally substituted C1-C6 alkoxy (e.g., methoxy), or optionally substituted heteroalkyl;

[0146] R 8 and R 9 are each independently selected from the group consisting of hydrogen, C1-C6 alkyl (e.g., methyl), halogenated C1-C6 alkyl (e.g., trifluoromethyl) or C1-C6 alkoxy (e.g., methoxy), or R 8 and R 9 forming an oxo group;

[0147] R 10 is hydrogen, hydroxy, optionally substituted alkyl, optionally substituted heteroalkyl, optionally substituted aryl, or optionally substituted arylalkyl, wherein each alkyl is C1-C 10 Alkyl, C3-C6 cycloalkyl, (C3-C6 cycloalkyl)C1-C4 alkyl, and optionally containing a single bond replaced by a double bond or a triple bond;

[0148] Each heteroalkyl group is an independently selected -O-, -S-, -N(R 10 )-, -S(=O)- or -S(=O)2-substituted alkyl group, wherein R 10 is hydrogen, alkyl, or alkyl in which one or more methylene groups are replaced by -O-, -S-, -NH, or -N-alkyl;

[0149] R 11 Is -H2 or -HR 12 ;

[0150] R 12 It is a C1-C6 alkyl group or a C1-C6 alkoxy group.

[0151] The pregnane neurosteroid and pregnenolone neurosteroid of the present invention may also each be a compound of formula III:

[0152]

[0153] or a pharmaceutically acceptable salt thereof, wherein:

[0154] X is O, S or NR 10 ;

[0155] R 1 is hydrogen, hydroxy, -CH2A, optionally substituted alkyl, optionally substituted heteroalkyl, optionally substituted aryl, or optionally substituted arylalkyl;

[0156] A is hydroxyl, O, S, NR 11 or an optionally substituted nitrogen-containing bicyclic heteroaryl or bicyclic heterocyclyl,

[0157] R 4 is hydrogen, hydroxy, oxo, optionally substituted alkyl or optionally substituted heteroalkyl,

[0158] R 2 , R 3 , R 5 , R 6 and R 7 each independently absent, hydrogen, hydroxy, halogen, optionally substituted C1-C6 alkyl, optionally substituted C1-C6 alkoxy (e.g., methoxy), or optionally substituted heteroalkyl;

[0159] R 8 and R 9 are each independently selected from the group consisting of hydrogen, C1-C6 alkyl (e.g., methyl), halogenated C1-C6 alkyl (e.g., trifluoromethyl) or C1-C6 alkoxy (e.g., methoxy), or R 8 and R 9 forming an oxo group;

[0160] R 10 is hydrogen, hydroxy, optionally substituted alkyl, optionally substituted heteroalkyl, optionally substituted aryl, or optionally substituted arylalkyl, wherein each alkyl is C1-C 10 Alkyl, C3-C6 cycloalkyl, (C3-C6 cycloalkyl)C1-C4 alkyl, and optionally containing a single bond replaced by a double bond or a triple bond;

[0161] Each heteroalkyl group is an independently selected -O-, -S-, -N(R 10 )-, -S(=O)- or -S(=O)2-substituted alkyl group, wherein R 10 is hydrogen, alkyl, or alkyl in which one or more methylene groups are replaced by -O-, -S-, -NH, or -N-alkyl;

[0162] R 11 Is -H2 or -HR 12 ;

[0163] R 12 It is a C1-C6 alkyl group or a C1-C6 alkoxy group.

[0164] a) Ganaxolone

[0165] Ganaxolone (CAS Reg. No. 38398-32-2, 3α-hydroxy-3β-methyl-5α-pregnane-20-one) is a 3β-methylated synthetic analog of allopregnanolone, an endogenous allosteric modulator of CNS GABAA receptors. The structural formula of ganaxolone is:

[0166]

[0167] 3α-Hydroxy, 3β-methyl-5α-pregnane-20-one ganaxolone

[0168] Benzodiazepines Ganaxolone has comparable potency and efficacy to allopregnanolone in activating synaptic and extrasynaptic GABAA receptors at sites distinct from those of barbiturates. (Carter 1997) Ganaxolone has protective activity in multiple rodent seizure models (Reddy 2012; Bialer 2010). Clinical studies have shown that ganaxolone has anticonvulsant activity with an acceptable safety and tolerability profile in adults and children at doses ranging from 900 to 1800 mg (Sperling 2017; Laxer 2000; Kerrigan 2000; Pieribone 2007). In addition, ganaxolone reduces seizures in children with IS and refractory pediatric epilepsy. In an open-label (OL) study, pediatric patients aged 2 to 60 months with a history of refractory seizures and IS were treated with doses of up to 36 mg / kg of ganaxolone for up to 3 months (Kerrigan 2000). Sixteen of 20 patients completed treatment, 15 of whom had a history of IS. Five of 15 patients had a ≥50% decrease in spasticity from baseline, five had a 25% to 50% decrease, and five had a <25% decrease. From Week 2 to Week 7, one patient was spasticity-free, and one non-responder (<25% decrease) was spasticity-free.

[0169] In addition to its anticonvulsant activity, ganaxolone has been shown to reduce anxiety, hyperactivity, and attention in children with fragile X syndrome (Ligsay 2016). Individuals with TSC also experience similar behavioral problems, with rates of ADHD and autism being approximately 50% (Jülich and Sahin, 2014). Therefore, ganaxolone treatment may increase GABAA-mediated signaling and improve not only seizure control but also behavioral abnormalities in individuals with TSC and TSC-related epilepsy.

[0170] Ganaxolone has the same core chemical structure as allopregnanolone, but with the addition of a 3β-methyl group designed to prevent conversion back into an entity active at nuclear hormone receptors, thereby eliminating the chance of unwanted hormonal effects while increasing the bioavailability of the neurosteroid and retaining its desired CNS activity.

[0171] Like allopregnanolone, ganaxolone (a neuroactive steroid) exhibits potent antiepileptic, anxiolytic, sedative, and hypnotic activities in animals through allosteric modulation of gamma-aminobutyric acid type A (GABAA) receptors in the central nervous system (CNS). Ganaxolone has comparable potency and efficacy to allopregnanolone in activating synaptic and extrasynaptic GABAA receptors at sites distinct from the site class.

[0172] Ganaxolone binds to synaptic and extrasynaptic GABA at a binding site unique to this class A Outside the synapse, ganaxolone can be absorbed into the cell membrane and diffuse to activate extrasynaptic GABA A receptors, thereby providing constant or tonic regulation of GABA inhibitory signals, calming overexcited neurons.

[0173] Ganaxolone is insoluble in water. Its solubility in 95% alcohol, propylene glycol, and polyethylene glycol is 13 mg / mL, 3.5 mg / mL, and 3.1 mg / mL, respectively.

[0174] Ganaxolone is primarily metabolized by the CYP3A family of hepatic enzymes, but interactions based on hepatic metabolism are limited to those caused by other drugs that induce or inhibit CYP3A4 / 5, such as ketoconazole.

[0175] In vitro, the clearance of ganaxolone appears to be driven primarily by CYP3A4. In clinical studies in adults, administration of grapefruit increased ganaxolone exposure in healthy volunteers. Ganaxolone levels were reduced in patients treated concomitantly with enzyme-inducing AEDs. These data further support the hypothesis that CYP3A4 is the primary factor in the clearance of ganaxolone in humans.

[0176] In adults, plasma concentrations of ganaxolone following oral administration are characterized by a high degree of variability. Single-dose PK parameters are strongly influenced by the rate and extent of ganaxolone absorption and whether the subject is in a fed or fasted state.

[0177] In the pediatric population, CYP3A4 expression levels approach those of adults at approximately 2 years of age (de Wildt et al., 2003), albeit with high inter-individual variability. Therefore, patients older than 2 years of age are expected to have ganaxolone clearance similar to that of adults.

[0178] Ganaxolone has a relatively long half-life - approximately 20 hours in human plasma after oral administration (Nohria, V. and Giller, E., Neurotherapeutics, (2007) 4(1): 102-105). In addition, the T of ganaxolone is max Ganaxolone is used to treat seizures in adults and pediatric patients with epilepsy.

[0179] Ganaxolone binds to GABAA receptors differently from other allosteric GABAA receptor modulators (e.g., benzodiazepines). Ganaxolone affects GABAA receptors by interacting with the recognition sites of benzodiazepines. Ganaxolone binds to both intrasynaptic and extrasynaptic receptors, mediating phasic and tonic modulation, respectively. Ganaxolone’s unique binding to these two receptors does not result in the same Compared to allopregnanolone, ganaxolone has oral bioavailability and cannot be converted back into intermediates with classical steroid hormone activity such as progesterone in the body, thus not activating the progesterone receptor directly or indirectly through metabolic conversion.

[0180] Ganaxolone administered intravenously has also been evaluated and shown to induce a burst suppression-like electroencephalographic (EEG) pattern in otherwise normal rats and block seizure responses in a model representing clinical status epilepticus (SE). Ganaxolone induced a sedative response but not a full anesthetic response.

[0181] In addition to anticonvulsant activity, ganaxolone has been shown to have anxiolytic properties as well as improve behaviors associated with autism. In a mouse model of post-traumatic stress disorder (PTSD), ganaxolone treatment reduced aggression and anxiety-like behaviors induced by social isolation (Pinna and Rasmussen, 2014). In another study, ganaxolone treatment improved social skills in the BTBR mouse model of autism (Kazdoba et al., 2016). Clinical studies of ganaxolone treatment of children and adolescents with fragile X syndrome (FXS) have shown that ganaxolone reduces anxiety and hyperactivity and improves attention in children and adolescents with higher baseline anxiety levels (Ligsay et al., 2017).

[0182] Ganaxolone did not interact with the human ether-a-go-go related gene (hERG) receptor at a measured concentration of 70 nM (n=2). Ganaxolone had no effects on cardiovascular parameters in dogs at single doses up to 15 mg / kg (maximum concentration [Cmax] of 1000 ng / mL, area under the concentration-time curve (AUC) (0-24) of 10000 ng·h / mL). In a 1-year canine toxicity study (Cmax>1500 ng / mL), transient sinus tachycardia (>190 beats per minute [bpm]) with shortening of the PR and QT intervals was observed in 4 animals after 3 months of dosing, but there was no therapeutic effect on QRS duration or corrected QT interval (QTc). No pulmonary effects were observed in female rats at doses up to 40 mg / kg.

[0183] In response to higher heart rates, the PR and QT intervals are physiologically shortened normally. There was no effect on QRS duration or QTc interval. No pulmonary effects were observed in female rats at doses up to 40 mg / kg.

[0184] Ganaxolone induces the major cytochrome P450 (CYP) isozymes 1A1 / 2 and 2B1 / 2 in female rats but not in male rats. Autoinduction has also been observed in mice and rats but not in dogs.

[0185] Tissue distribution studies in mice and rats have shown that [ 14 C]-Ganexolone is rapidly distributed throughout the body to highly perfused organs, intestine, and adipose tissue, with brain concentrations of ganaxolone approximately 5-fold higher than those in plasma.

[0186] The majority of radioactivity excreted in all species was in the feces (>70%), with the remainder excreted in the urine.

[0187] In toxicology studies, the most common effect of ganaxolone treatment was dose-related sedation, which is a result of GABA A Expected pharmacology of a receptor positive modulator. There was little evidence of target organ or systemic toxicity associated with single or repeated doses of ganaxolone in both oral and intravenous regimens. No functional or anatomical changes within hematopoietic tissues or any specific organ, such as the liver, kidney, or gastrointestinal (GI) system, were observed in repeated dose studies. In a 6-month study, ganaxolone induced hepatic enzymes in rats, with more pronounced effects in females, which were associated with increased liver weights and dose-related hepatocyte hypertrophy.

[0188] In a chronic oral toxicity study in dogs, mean C values ​​greater than 1500 ng / mL (10 and 15 mg / kg per day) weremax levels caused an increase in body weight and total plasma cholesterol levels.

[0189] When administered intravenously to rats and dogs, the major dose-limiting toxicity was found to be sedation. The no observed adverse effect level (NOAEL) in rats after 14 days of intravenous administration was determined to be 42 mg / kg per day for males and 30 mg / kg per day for females. After 28 days of administration of ganaxolone by intravenous bolus followed by continuous intravenous infusion, the NOAEL in dogs was 7.20 mg / kg per day, corresponding to steady-state concentrations of approximately 330 ng / mL and 333 ng / mL. No local tolerance was found in rabbit studies. Finally, ganaxolone did not cause hemolysis in vitro and was compatible with human plasma.

[0190] Ganaxolone was not teratogenic in rats or mice and had no significant developmental effects on offspring. Ganaxolone had no effects on fertility or early embryonic development in rats. No mutagenic potential was detected. Treatment of neonatal rats with ganaxolone produced the expected signs of sedation but did not affect development or show any postmortem changes.

[0191] b) Allopregnanolone

[0192] Allopregnanolone (CAS Reg. No. 516-54-1, 3α,5α-altropregnanolone) is an endogenous progesterone derivative with anticonvulsant activity.

[0193]

[0194] 3α,5α-Allopregnanolone

[0195] Allopregnanolone has a relatively short half-life of approximately 45 minutes in human plasma.

[0196] Allopregnanolone exhibits potent antiepileptic, anxiolytic, sedative, and hypnotic activities in animals by virtue of its GABAA receptor modulating activity.

[0197] In addition to its efficacy in treating epileptic seizures, allopregnanolone is being evaluated for use in treating neurodegenerative diseases, including Alzheimer's disease, Parkinson's disease, Huntington's disease, and amyotrophic lateral sclerosis, as well as for the treatment of lysosomal storage diseases characterized by abnormal cholesterol synthesis, such as Niemann Pick A, B, and C disease, Gaucher disease, and Tay Sachs disease. (See US 8,604,011, which is incorporated herein by reference for its teachings on the use of allopregnanolone to treat neurological disorders.)

[0198] The relationship between progesterone and its metabolite allopregnanolone and seizures has been extensively studied in women with catamenial epilepsy, a condition in which the frequency of seizures varies with the phases of the menstrual cycle. During the menstrual cycle, when progesterone is low (e.g., around the time of menstruation), the likelihood of seizures tends to increase (French 2005). Circulating allopregnanolone levels parallel those of progesterone. While the reproductive effects of progesterone are related to its interaction with intracellular progesterone receptors, the anticonvulsant effects of progesterone are independent of this (Reddy and Rogawski 2009). The anticonvulsant activity of progesterone results from its conversion to the neurosteroid allopregnanolone (Kokate et al. 1999). Due to its effects on GABAA receptors, allopregnanolone has been shown to protect against seizures in a number of animal models (Reddy and Rogawski 2009). Ganaxolone is a synthetic analog of allopregnanolone without the progesterone-related effects that may be used to treat TSC-related epilepsy.

[0199] c) Alfaxalone

[0200] Alfaxalone, also known as alfaxalone (CAS Reg. No. 23930-19-0, 3α-hydroxy-5α-pregnane-11,20-dione), is a neurosteroid with anesthetic activity. It is used as a general anesthetic in veterinary practice. Anesthetics are often used in combination with anticonvulsants to treat refractory epileptic seizures. Injectable nanoparticle neurosteroid dosage forms containing alfaxalone alone or in combination with ganaxolone or allopregnanolone are within the scope of the present disclosure.

[0201]

[0202] 3α-Hydroxy-5α-pregnane-11,20-dione alfaxalone

[0203] d) Aphadolon

[0204] Alfadolone, also known as alfadolone (CAS Reg. No. 14107-37-0, 3α,21-dihydroxy-5α-pregnane-11,20-dione) is a neurosteroid with anesthetic properties. Its hydroacetate is used in combination with alfaxalone as a veterinary anesthetic.

[0205]

[0206] 3α,21-dihydroxy-5α-pregnane-11,20-dione afadolone

[0207] e) Additional neurosteroids

[0208] Other neurosteroids that can be used in the nanoparticle neurosteroid formulations of the present disclosure and the methods disclosed herein include, but are not limited to, pregnenolone, hydroxypregnanedione (CAS Reg. No. 303-01-5, (5β)-21-hydroxypregnane-3,20-dione), minaxolone (CAS Reg. No. 62571-87-3, 2β, 3α, 5α, 11α)-11-(dimethylamino)-2-ethoxy-3-hydroxypregnane-20-one), pregnanolone (CAS Reg. No. 128-20-1, (3α, 5β)-d-hydroxypregnane-20-one), renanolone (CAS Reg. No. 565-99-1, 3α-hydroxy-5β-pregnane-11,20-dione) or tetrahydrocorticosterone (CAS Reg. No. 565-99-1, 3α-hydroxy-5β-pregnane-11,20-dione). Reg. No. 68-42-8, 3α,5α-pregnane-20-dione).

[0209] Additional neurosteroids useful in the nanoparticle neurosteroid formulations of the present disclosure and methods disclosed herein include Co26749 / WAY-141839, Co134444, Co177843, and Sage-217, Sage-324, and Sage-718. Co26749 / WAY-141839, Co134444, Co177843, and Sage-217 have the following structures:

[0210]

[0211] Additional neurosteroids useful in the nanoparticle neurosteroid formulations of the present disclosure and methods disclosed herein include compounds disclosed in U.S. Patent Publication No. 2016-0229887 (U.S. Serial No. 14 / 913,920, filed February 23, 2016), which is incorporated herein by reference in its entirety.

[0212] IV. Dosage

[0213] The pregnenolone neurosteroid used in the methods disclosed herein can be administered in one, two, three or four doses in an amount of about 1 mg per day to about 5000 mg per day. In certain embodiments, doses of 1600 mg per day and 2000 mg per day may cause drowsiness, and a dose of 1800 mg per day defines the best combination of drug exposure, ease of administration and tolerability.

[0214] When the pregnenolone neurosteroid is ganaxolone, the target and maximum dose of ganaxolone is about 1800 mg per day. In these embodiments, this dose provides the highest possible exposure based on the nonlinear kinetics of ganaxolone. Thus, when the pregnenolone neurosteroid is ganaxolone, the amount of ganaxolone administered in the methods disclosed herein is generally about 200 mg per day to about 1800 mg per day, about 300 mg per day to about 1800 mg per day, about 400 mg per day to about 1800 mg per day, about 450 mg per day to about 1800 mg per day, about 675 mg per day to about 1800 mg per day, about 900 mg per day to about 1800 mg per day, about 1125 mg per day to about 1800 mg per day, about 1350 mg per day to about 1800 mg per day, about 1575 mg per day to about 1800 mg per day, or about 1800 mg per day, at a dose of 1 mg / kg per day to about 80 mg / kg per day, divided into one, two, three, or four doses. In certain embodiments, the target and maximum doses of ganaxolone may be higher, if desired, to achieve improved therapeutic benefit, but limited by side effects (e.g., somnolence).

[0215] In certain embodiments, about 300 mg to about 2000 mg, about 900 mg to about 1800 mg, about 950 mg to about 1800 mg, about 1000 mg to about 1800 mg, about 1100 mg to about 1800 mg, or about 1200 mg of ganaxolone is administered orally daily for two or more consecutive days (e.g., for a period of 1 week to 50 years, or for the life of the patient). Ganaxolone may be administered orally or parenterally in one, two, three, or four doses per day.

[0216] Whether a person receives ganaxolone two or three times a day may depend on the formulation. For patients taking oral immediate-release capsules, ganaxolone is usually administered twice a day, with each dose separated by 8 to 12 hours from the next and / or previous dose. For patients taking the oral suspension, ganaxolone is usually administered three times a day, with each dose separated by 4 to 8 hours from the next and / or previous dose.

[0217] When the pregnenolone neurosteroid is ganaxolone, the methods disclosed herein comprise administering ganaxolone at a dosage of about 1 mg / kg per day to about 80 mg / kg per day, provided that the total amount of ganaxolone administered does not exceed 2000 mg per day.

[0218] The methods described herein may also include administering ganaxolone in a therapeutically effective amount to achieve a ganaxolone plasma concentration of 100 ng / ml or greater for about 70% or more over a 24-hour period in a day. In some cases, the plasma concentration of ganaxolone may be above 100 ng / ml for about at least about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or more over a 24-hour period.

[0219] In order to achieve about 70% or more of a ganaxolone plasma concentration of 100 ng / ml or more within a 24 hour day, ganaxolone may be administered at least three times a day. Three times a day is preferred, and if necessary or desired, ganaxolone may be administered more than three times a day to achieve the desired ganaxolone trough concentration. For example, ganaxolone may be administered three times a day, four times a day, five times a day, six times a day, seven times a day, eight times a day, or more.

[0220] Ganaxolone administered according to the methods disclosed herein can be administered at the same or lower daily doses used in clinical trials and can increase drug exposure by, for example, maintaining a serum concentration of ganaxolone of at least about 100 ng / ml for at least about 70% or more over a 24-hour day. A total daily dose of ganaxolone of about 1800 mg, about 1700 mg, about 1600 mg, about 1500 mg, or 63 mg / kg per day can be administered, provided that the total daily dose is administered in three or more separate administrations (preferably each containing the same amount of ganaxolone) to produce a serum ganaxolone concentration of at least about 100 ng / ml for at least 70% or more over a 24-hour day. Typically, a total daily dose of ganaxolone of 1500 mg per day can produce a plasma concentration of at least about 100 ng / ml or greater for about 70% or more over a 24-hour day when administered three times a day.

[0221] For example, when administering a total daily dose of 1500 mg of ganaxolone, a dose of about 500 mg may be administered three times a day. For example, when administering a total daily dose of 1800 mg of ganaxolone, a dose of about 600 mg may be administered three times a day. The maximum daily dose of ganaxolone is administered at least three time intervals over a 24 hour period in the same or different doses. A skilled clinician will appreciate that the amount of ganaxolone administered three times a day may be adjusted to achieve a desired trough level of ganaxolone as long as the total amount does not exceed the maximum daily dose of ganaxolone.

[0222] Although a plasma concentration of at least about 100 ng / ml is preferred, some variability may exist based on, for example, differences in subject weight, metabolism, age, seizure duration, and seizure severity.

[0223] Ganaxolone can be administered orally (e.g., as an oral suspension or oral capsule) or as an intravenous formulation. Preferably, ganaxolone is administered orally. Oral administration may include, but is not limited to, oral suspension formulations and oral capsules.

[0224] Plasma concentrations of at least about 100 ng / ml or more over a 24-hour day of about 70% or more result in a reduction in seizures and / or improved seizure inhibition. For example, a reduction in seizure frequency of at least 20% or greater relative to baseline seizure frequency can be achieved. For example, a reduction in seizure frequency of at least 35% or greater relative to baseline can be achieved. EEG can be used to monitor seizure load and / or frequency.

[0225] The pharmacokinetics of ganaxolone in a formulation comprising immediate release 0.3 micron particles (e.g., the formulation of Example 2) are linear up to approximately 1200 mg per day (twice a day ("BID")), with a modest increase in exposure at 1600 mg per day, and little further increase in exposure at 2000 mg per day. Thus, in order to maintain the highest possible trough levels in all subjects, a dose of 1800 mg per day is generally targeted, but may be adjusted higher or lower in some individuals to provide optimal therapeutic effect.

[0226] In certain embodiments, ganaxolone is administered at a dose exceeding 5 mg / kg per day, such as from about 6 mg / kg per day to about 80 mg / kg per day, provided that the total amount of ganaxolone administered does not exceed 1800 mg per day.

[0227] In certain embodiments, the dose of ganaxolone is adjusted during the treatment period from 15 mg / kg per day to 100 mg / kg per day up to a maximum dose of 1800 mg per day.

[0228] In certain embodiments, the method of treatment comprises administering at least 33 mg / kg of ganaxolone per day in one, two, three, or four doses, with a maximum daily dose of about 1800 mg.

[0229] In certain embodiments, the human is about 0.6 to about 7 years old, and the dose of ganaxolone administered is about 1.5 mg / kg twice a day ("BID") (3 mg / kg per day) to 12 mg / kg three times a day ("TID") (36 mg / kg per day). In embodiments where the human receives a 12 mg / kg TID dosing regimen, a trough concentration of at least about 38.5 ± 37.4 ng / mL is achieved.

[0230] In certain embodiments, ganaxolone is orally administered in a beta-cyclodextrin formulation at a dose of 6 mg / kg BID (12 mg / kg per day) to 12 mg / kg TID (36 mg / kg per day) with food and achieves plasma ganaxolone concentrations of up to 22.1 ng / mL and 5.7 to 43.7 ng / mL in the 4th and 8th weeks of administration, respectively.

[0231] In certain embodiments, ganaxolone is administered orally with food at a dose of 1 to 12 mg / kg TID (3 to 36 mg / kg per day) and achieves plasma concentrations of ganaxolone of up to 5.78 ng / mL (1 mg / kg TID) to 10.3 to 16.1 ng / mL (12 mg / kg TID).

[0232] In certain embodiments, ganaxolone is administered orally in an oral suspension formulation at a dose of 3 to 18 mg / kg TID (9 to 54 mg / kg per day) and achieves a ganaxolone C of about 123 ng / mL. max and a trough concentration of approximately 23 ng / mL.

[0233] In certain embodiments, the average ganaxolone C is based on 1000 mg ganaxolone administered orally three times a day. min (trough) is 55 ng / ml to about 100 ng / ml, and C max The level was about 240 ng / ml to 400 ng / ml (eg, 262 ng / mL).

[0234] In certain embodiments, the methods provide for an average C based on oral administration of 1000 mg ganaxolone twice a day. min (Valley) and C max The levels were about 56.9 ng / ml and about 262 ng / mL, respectively.

[0235] In certain embodiments, administration of ganaxolone provides a C of greater than 3, 3.5, 4, 4.5, 5, or 6. min / C max ratio. min / C max The ratio can be provided after a single dose administration and / or at steady state after administration. In certain embodiments, C min / C max The ratio remained the same regardless of the dose of ganaxolone administered.

[0236] In certain embodiments, the dose administered is determined from a pediatric pharmacokinetic model that allows determination of a dose across a wide pediatric age range that will produce a C similar to that achieved based on an effective dose determined in an adult epilepsy population. maxand AUC exposure of ganaxolone dose. This model can be constructed, for example, using standard methods taking into account the pharmacokinetic data in this application.

[0237] In certain embodiments, multiple titration steps may be used to administer the pregnenolone neurosteroid to a patient until a therapeutically effective dosage regimen is achieved. For example, depending on the patient's size, about six to eight titration steps may be used.

[0238] In certain embodiments, the methods disclosed herein include establishing a baseline seizure frequency for a patient, initially administering a dose of ganaxolone to the patient in an amount of about 0.5 mg / kg per day to about 15 mg / kg per day; and gradually increasing the dose of ganaxolone to an amount of about 18 mg / kg per day to about 60 mg / kg per day over the course of 4 weeks, wherein for patients weighing more than 30 kg, the total dose of ganaxolone is up to about 1800 mg per day. For patients weighing less than 30 kg, the total daily dose of ganaxolone may be less (e.g., about 63 mg per day). In certain preferred embodiments, the initial dose of ganaxolone is about 4.5 mg / kg per day. In certain preferred embodiments, the dose of ganaxolone is increased to about 36 mg / kg per day. In certain preferred embodiments, if the patient experiences a dose-limiting adverse event, the ganaxolone dose is reduced to the previous level.

[0239] In certain embodiments, for subjects weighing more than 30kg, treatment is started with a dose of 900mg per day in divided doses. The dose is then increased by about 20% to 50% (e.g., an increase of 33% from 900mg per day to 1200mg per day) at intervals of no less than 3 days and no more than 2 weeks, provided that the current dose is reasonably tolerated until the desired efficacy is achieved or the maximum tolerated dose (MTD) level is reached. Subsequent dose adjustments can be made in increments of about 20% to 50%, with at least 3 days between dose changes, unless for safety considerations. The maximum allowed dose in these embodiments is 1800mg per day.

[0240] In certain embodiments, for subjects weighing less than 30 kg, treatment is started at 18 mg / kg per day and increased by about 20% to 50% increments at intervals of not less than 3 days and not more than 2 weeks, provided that the current dose is reasonably tolerated until the desired efficacy is achieved or the maximum tolerated dose (MTD) level is reached. Subsequent dose adjustments can be made in increments of about 20% to 50%, with at least 3 days between dose changes, unless for safety considerations. The maximum allowed dose in these embodiments is 63 mg / kg per day.

[0241] For people weighing ≥ 28 kg (62 lbs), ganaxolone may be started at a dose of about 300 mg per day to about 600 mg per day (e.g., 400 mg per day) in divided doses. The dose will be increased by 450 mg per day every 7 days until 1800 mg per day or the maximum tolerated dose is reached.

[0242] For humans weighing <28 kg (62 lbs), ganaxolone can be started at a dose of about 10 mg / kg per day to about 30 mg / kg per day (e.g., 18 mg / kg per day) and increased by about 15 mg / kg per day each week until 63 mg / kg per day is reached.

[0243] In certain embodiments, ganaxolone is administered as an oral suspension in increments of 10 mg per day to 20 mg per day (e.g., 15 mg / kg per day), up to 63 mg / kg per day (maximum 1800 mg per day), or as an oral capsule in increments of 225 mg per day to 900 mg per day (e.g., 450 mg per day). In some of these embodiments, ganaxolone may be administered, for example, as follows: 6 mg / kg three times daily (TID) (18 mg / kg per day) suspension / 225 twice daily (BID) (450 mg per day) capsules - Days 1-7; 11 mg / kg TID (33 mg / kg per day) suspension / 450 BID (900 mg per day) capsules - Days 8-14; 16 mg / kg TID (48 mg / kg per day) suspension / 675 BID (1350 mg per day) capsules - Days 15-21; 21 mg / kg TID (63 mg / kg per day, not to exceed 1800 mg per day) suspension / 900 BID (1800 mg per day) capsules - Days 22-28.

[0244] In certain embodiments, ganaxolone is administered as an oral suspension using the following titration regimen:

[0245] 15kg(33lbs)

[0246]

[0247] 20kg(44lbs)

[0248]

[0249] 25kg(55lbs)

[0250]

[0251] 30kg(66lbs)

[0252]

[0253] In certain embodiments, ganaxolone is administered in capsules and the following titration regimen is used:

[0254]

[0255] In certain embodiments, trough concentrations associated with maximum efficacy are in the range of about 55 ng / mL, about 60 ng / ml, or about 65 ng / ml (0.3 micron suspension; TID dosing), and a dose of 1800 mg per day (0.3 micron capsule, BID administration) provides trough plasma concentrations within this range.

[0256] The treatment methods disclosed herein encompass administration of neurosteroids (e.g., ganaxolone) with or without food. In certain embodiments, ganaxolone is administered with food.

[0257] V. Duration of treatment

[0258] The duration of treatment according to the methods disclosed herein can range from 1 day to more than 2 years. For example, the duration of treatment can be from about 1 day to about 80 years, from about 1 day to about 70 years, from about 1 day to about 60 years, from about 1 day to about 50 years, from about 1 day to about 45 years, from about 2 days to about 45 years, from about 2 days to about 40 years, from about 5 days to about 35 years, from about 10 days to about 30 years, from about 10 days to about 30 years, from about 15 days to about 30 years. In some embodiments, the duration of treatment continues as long as the subject continues to derive therapeutic benefit from the administration of the neurosteroid (e.g., ganaxolone). In some embodiments, the duration of treatment is 14 days, 28 days, 30 days, 6 weeks, 8 weeks, 10 weeks, 12 weeks, 6 months, 1 year, 2 years, 2.5 years, 3 years, 3.5 years, 4 years, 4.5 years, 5 years, 5.5 years, 6 years, 6.5 years, 7 years, 7.5 years, 8 years, 8.5 years, 9 years, 9.5 years, or 10 years.

[0259] In certain embodiments, at the end of the treatment period, or upon cessation of treatment, the dosage is gradually reduced over a period of 1 to 4 weeks based on the subject's age, weight, dosage, and duration of treatment.

[0260] VI. Preparation

[0261] Any desired formulation comprising a pregnenolone neurosteroid (e.g., ganaxolone) and one or more pharmaceutically acceptable excipients can be administered according to the methods disclosed herein. A therapeutically effective amount of a pregnenolone neurosteroid is included to treat one or more symptoms of TSC or TSC-related epilepsy. In certain embodiments, the formulation is free of cyclodextrins, including sulfoalkyl ether cyclodextrins and modified forms thereof.

[0262] In preferred embodiments, the amount of pregnenolone neurosteroid in the formulation is therapeutically effective for treating symptoms of TSC-related epilepsy, e.g., after oral administration of the formulation for 1 week and / or 2 weeks and / or 3 weeks and / or 4 weeks and / or 6 weeks and / or 7 weeks and / or 8 weeks and / or 9 weeks and / or 10 weeks and / or 11 weeks and / or 12 weeks or more.

[0263] In a preferred embodiment, the pregnenolone neurosteroid (eg, ganaxolone) is incorporated into a pharmaceutically acceptable composition for oral administration. In certain preferred embodiments, such formulations may be liquids (e.g., aqueous liquids (encompassing suspensions, solutions, etc.). In other preferred embodiments, the oral formulation may be an oral solid dosage form (e.g., an oral capsule or tablet). In the most preferred embodiment, the oral formulation is an oral suspension comprising a pregnenolone neurosteroid or an oral capsule comprising a pregnenolone neurosteroid. Preferably, a unit dose of the oral formulation contains a therapeutically effective amount of a pregnenolone neurosteroid, which can be orally administered to a (e.g., human) patient (e.g., an infant, child, adolescent, or adult). In certain embodiments, the oral suspension is administered to the patient via the use of an oral syringe. For example, oral suspensions are contemplated for use in humans weighing less than about 30 kg (e.g., about 28 kg). On the other hand, the oral suspension may be administered to humans who have difficulty swallowing solid oral dosage forms. Children greater than 30 kg may take solid dosage forms, such as ganaxolone capsules. Ganaxolone oral suspensions may be administered by an oral administration syringe, for example, three times a day. Ganaxolone capsules may be administered, for example, twice a day. Ganaxolone may be better absorbed by the patient with a meal (milk).

[0264] As described in U.S. Pat. No. 8,022,054, the liquid formulation can be an aqueous dispersion of stabilized pregnenolone neurosteroid (e.g., ganaxolone) particles, the particles comprising ganaxolone, a hydrophilic polymer, a wetting agent, and an effective amount of a complexing agent to stabilize particle growth after initial particle growth and endpoint is reached, the complexing agent being selected from the group of small organic molecules having a molecular weight of less than 550 and containing a moiety selected from the group consisting of a phenol moiety, an aromatic ester moiety, and an aromatic acid moiety, wherein the stabilized particles have a volume weighted median diameter (D50) of the particles of about 50 nm to about 500 nm, the complexing agent is present in an amount of about 0.05% to about 5% w / w based on the weight of the particles, and the particles are dispersed in an aqueous solution further containing at least two preservatives, the preservatives being present in an amount sufficient to inhibit microbial growth. The amount of the hydrophilic polymer can be about 3% to about 50% w / w based on the weight of the solid particles. The amount of the wetting agent can be about 0.01% to about 10% w / w based on the weight of the solid particles. The amount of pregnenolone neurosteroid (e.g., ganaxolone) may be from about 10% to about 80% (and in certain embodiments from about 50% to about 80%) based on the weight of the stabilized particles. When the particles are dispersed in simulated gastric fluid (SGF) or simulated intestinal fluid (SIF) at a concentration of 0.5 to 1 mg ganaxolone / mL and placed in a heating bath at 36° C. to 38° C. for 1 hour, the D50 of the stabilized particles increases by no more than about 150% compared to the volume weighted median diameter (D50) of the stabilized particles when the particles are dispersed in distilled water under the same conditions, wherein the volume weighted median diameter (D50) of the stabilized particles dispersed in SGF or SIF is less than about 750 nm. When the formulation is dispersed in 15 mL of SGF or SIF at a concentration of 0.5 to 1 mg ganaxolone / mL, the volume weighted median diameter (D50) of the stabilized particles increases by no more than about 150% compared to the D50 of the stabilized particles when the particles are dispersed in distilled water under the same conditions, wherein the volume weighted median diameter (D50) of the stabilized particles dispersed in SGF or SIF is less than about 750 nm. The complexing agent can be a paraben, benzoic acid, phenol, sodium benzoate, methyl anthranilate, etc. The hydrophilic polymer can be a cellulosic polymer, a vinyl polymer, and mixtures thereof. The cellulosic polymer can be a cellulose ether, such as hydroxypropyl methylcellulose. The vinyl polymer can be a polyvinyl alcohol, such as vinyl pyrrolidone / vinyl acetate copolymer (S630). The wetting agent can be sodium lauryl sulfate, a pharmaceutically acceptable salt of docusate, and mixtures thereof. The aqueous dispersion can also contain a sweetener, such as sucralose. The preservative is selected from the group consisting of potassium sorbate, methylparaben, propylparaben, benzoic acid, butylparaben, ethanol, benzyl alcohol, phenol, benzalkonium chloride, and mixtures of any of the foregoing.

[0265] In some embodiments, a liquid pregnenolone neurosteroid (e.g., ganaxolone) formulation for oral administration to a subject is provided, comprising ganaxolone particles as described herein and at least one dispersing agent or suspending agent. The ganaxolone formulation can be a powder and / or granules for suspension, and upon mixing with water, a substantially uniform suspension is obtained. As described herein, the aqueous dispersion can contain amorphous and non-amorphous ganaxolone particles composed of a variety of effective particle sizes, such that ganaxolone particles with smaller effective particle sizes are absorbed faster and ganaxolone particles with larger effective particle sizes are absorbed slower. In certain embodiments, the aqueous dispersion or suspension is an immediate release formulation. In another embodiment, the aqueous dispersion containing amorphous ganaxolone particles is formulated such that about 50% of the ganaxolone particles are absorbed within about 3 hours after administration and about 90% of the ganaxolone particles are absorbed within about 10 hours after administration. In other embodiments, the addition of a complexing agent to the aqueous dispersion results in a greater span of ganaxolone-containing particles extending the drug absorption phase such that 50-80% of the particles are absorbed within the first 3 hours and about 90% to about 10 hours.

[0266] A suspension is "substantially homogeneous" when the suspension is mostly homogeneous, i.e., the suspension is composed of approximately the same concentration of pregnenolone neurosteroid (e.g., ganaxolone) at any point throughout the suspension. Preferred embodiments are those that provide substantially the same concentration (within 15%) when measured at different points in the aqueous oral formulation of ganaxolone after shaking. Particularly preferred are aqueous suspensions and dispersions that maintain homogeneity (up to 15% variation) when measured 2 hours after shaking. Homogeneity should be determined by sampling methods consistent with determining homogeneity of the entire composition. In one embodiment, an aqueous suspension can be resuspended into a homogeneous suspension by physical agitation lasting less than 1 minute. In another embodiment, an aqueous suspension can be resuspended into a homogeneous suspension by physical agitation lasting less than 45 seconds. In another embodiment, an aqueous suspension can be resuspended into a homogeneous suspension by physical agitation lasting less than 30 seconds. In yet another embodiment, agitation is not required to maintain a homogeneous aqueous dispersion.

[0267] In some embodiments, the pregnenolone neurosteroid (e.g., ganaxolone) powder for use in the aqueous dispersion described herein comprises stable ganaxolone particles having an effective particle size of less than 500 nm by weight formulated with ganaxolone particles having an effective particle size of greater than 500 nm by weight. In such embodiments, the formulation has a particle size distribution wherein about 10% to about 100% by weight of the ganaxolone particles are between about 75 nm and about 500 nm, about 0% to about 90% by weight of the ganaxolone particles are between about 150 nm and about 400 nm, and about 0% to about 30% by weight of the ganaxolone particles are greater than about 600 nm. The ganaxolone particles described herein can be amorphous, semi-amorphous, crystalline, semi-crystalline, or mixtures thereof.

[0268] In one embodiment, the aqueous suspensions or dispersions described herein comprise ganaxolone particles or ganaxolone complexes at a concentration of about 20 mg / ml to about 150 mg / ml of suspension. In another embodiment, the oral aqueous dispersions described herein comprise ganaxolone particles or ganaxolone complex particles at a concentration of about 25 mg / ml to about 75 mg / ml of solution. In yet another embodiment, the oral aqueous dispersions described herein comprise ganaxolone particles or ganaxolone complexes at a concentration of about 50 mg / ml of suspension. The aqueous dispersions described herein are particularly useful for administering ganaxolone to infants (less than 2 years old), children under 10 years old, and any patient group that is unable to swallow or ingest solid oral dosage forms.

[0269] The dosage form of a liquid pregnenolone neurosteroid (e.g., ganaxolone) preparation for oral administration can be an aqueous suspension selected from the group including, but not limited to, pharmaceutically acceptable oral aqueous dispersions, emulsions, solutions, and syrups. See, e.g., Singh et al., Encyclopedia of Pharmaceutical Technology, 2nd Edition, pp. 754-757 (2002). In addition to the ganaxolone particles, the liquid dosage form can also include additives such as: (a) a disintegrant; (b) a dispersant; (c) a wetting agent; (d) at least one preservative, (e) a viscosity increasing agent, (f) at least one sweetener, (g) at least one flavoring agent, (h) a complexing agent, and (i) an ionic dispersion modifier. In some embodiments, the aqueous dispersion can also include a crystallization inhibitor.

[0270] Examples of disintegrants for aqueous suspensions and dispersions include, but are not limited to, starches, for example, natural starches such as corn starch or potato starch, pregelatinized starches such as National 1551 or or sodium starch glycolate or Cellulose, such as wood products, microcrystalline cellulose (e.g. and ), methylcellulose, cross-linked carboxymethylcellulose or a cross-linked cellulose, such as cross-linked carboxymethylcellulose sodium Cross-linked carboxymethyl cellulose or cross-linked cross-linked carboxymethyl cellulose; cross-linked starch, such as sodium starch glycolate; cross-linked polymers, such as crospovidone; cross-linked polyvinyl pyrrolidone; alginates, such as alginic acid or alginates, such as sodium alginate; clays, such as HV (magnesium aluminum silicate); gums such as agar, guar, locust bean, karaya, pectin or tragacanth; sodium starch glycolate; bentonite; natural sponge; surfactants; resins such as cation exchange resins; citrus pulp; sodium lauryl sulfate; sodium lauryl sulfate in combination with starch, etc.

[0271] In some embodiments, dispersants suitable for use in the aqueous suspensions and dispersions described herein are known in the art and include, for example, hydrophilic polymers, electrolytes, or 80, PEG, polyvinylpyrrolidone (PVP; commercially known as ) and carbohydrate-based dispersants such as hydroxypropyl cellulose and hydroxypropyl cellulose ethers (e.g., HPC, HPC-SL, and HPC-L), hydroxypropyl methylcellulose and hydroxypropyl methylcellulose ethers (e.g., HPMC K100, HPMC K4M, HPMC K15M, and HPMC K100M), sodium carboxymethyl cellulose, methylcellulose, hydroxyethyl cellulose, hydroxypropyl methylcellulose phthalate, hydroxypropyl methylcellulose acetate stearate, amorphous cellulose, magnesium aluminum silicate, triethanolamine, polyvinyl alcohol (PVA), polyvinyl pyrrolidone / vinyl acetate copolymer ( S-630), 4-(1,1,3,3-tetramethylbutyl)-phenol polymer with ethylene oxide and formaldehyde (also known as tyloxapol), poloxamers (e.g., Pluronics and They are block copolymers of ethylene oxide and propylene oxide); and poloxamines (e.g., Tetronic 9080, also known as poloxamine 9080, which is a tetrafunctional block copolymer derived from the sequential addition of propylene oxide and ethylene oxide to ethylenediamine (BASF Corporation, Parsippany, NJ)). In other embodiments, the dispersant is selected from the group that does not include one of the following agents: a hydrophilic polymer; an electrolyte; or 80; PEG; polyvinylpyrrolidone (PVP); hydroxypropyl cellulose and hydroxypropyl cellulose ethers (e.g., HPC, HPC-SL and HPC-L); hydroxypropyl methylcellulose and hydroxypropyl methylcellulose ethers (e.g., HPMC K100, HPMC K4M, HPMC K15M, HPMC K100M and USP 2910 (Shin-Etsu); sodium carboxymethylcellulose; methylcellulose; hydroxyethylcellulose; hydroxypropylmethylcellulose phthalate, hydroxypropylmethylcellulose acetate stearate; non-crystalline cellulose; magnesium aluminum silicate; triethanolamine; polyvinyl alcohol (PVA); 4-(1,1,3,3-tetramethylbutyl)-phenol polymer with ethylene oxide and formaldehyde; poloxamers (e.g., and which are block copolymers of ethylene oxide and propylene oxide); or poloxamines (e.g., Also known as Poloxamine 908%).

[0272] Wetting agents (including surfactants) suitable for use in the aqueous suspensions and dispersions described herein are known in the art and include, but are not limited to, acetyl alcohol, glyceryl monostearate, polyoxyethylene sorbitan fatty acid esters (e.g., commercially available For example and (ICI Specialty Chemicals)), and polyethylene glycol (e.g. and and (Union Carbide), oleic acid, glyceryl monostearate, sorbitan monooleate, sorbitan monolaurate, triethanolamine oleate, polyoxyethylene sorbitan monolaurate, polyoxyethylene sorbitan monolaurate, sodium oleate, sodium lauryl sulfate, docusate sodium, triacetin, vitamin E TPGS, sodium taurocholate, simethicone, phosphatidylcholine, etc.

[0273] Suitable preservatives for aqueous suspensions or dispersions described herein include, for example, potassium sorbate, parabens (e.g., methylparaben and propylparaben) and salts thereof, benzoic acid and salts thereof, other esters of parabens such as butylparaben, alcohols such as ethanol or benzyl alcohol, phenolic compounds such as phenol, or quaternary compounds such as benzalkonium chloride. As used herein, preservatives are incorporated into the dosage form at a concentration sufficient to inhibit microbial growth. In one embodiment, the aqueous liquid dispersion may contain methylparaben and propylparaben in a concentration range of about 0.01% to about 0.3% by weight of methylparaben and 0.005% to 0.03% by weight of propylparaben, based on the total weight of the aqueous dispersion. In yet another embodiment, the aqueous liquid dispersion may include 0.05 to about 0.1% methylparaben and 0.01-0.02% propylparaben by weight of the aqueous dispersion.

[0274] Suitable viscosity enhancers for aqueous suspensions or dispersions described herein include, but are not limited to, methylcellulose, xanthan gum, carboxymethylcellulose, hydroxypropylcellulose, hydroxypropylmethylcellulose, Plasdone.RTM.S-630, carbomer, polyvinyl alcohol, alginate, gum arabic, chitosan, and combinations thereof. The concentration of the viscosity enhancer will depend on the selected agent and the desired viscosity.

[0275] Examples of natural and artificial sweeteners suitable for use in the aqueous suspensions or dispersions described herein include, for example, gum arabic syrup, acesulfame potassium, alitame, anise, apple, aspartame, banana, Bavarian cream, berries, blackcurrant, butterscotch, calcium citrate, camphor, caramel, cherry, cherry cream, chocolate, cinnamon, bubble gum, citrus, citrus punch, citrus cream, marshmallow, cocoa, cola, cool cherry, cool citrus, sodium cyclamate, cylamate, dextrose, eucalyptus, eugenol, fructose, fruit punch, ginger, glycyrrhetinate, glycyrrhiza (licorice) syrup, grape, grapefruit, honey, isomalt, lemon, lime, lemon cream, monoammonium glycyrrhizinate Maltol, mannitol, maple, marshmallow, menthol, mint cream, mixed berries, neohesperidin DC, neotame, orange, pear, peach, mint, mint cream, Powder, raspberry, root beer, rum, saccharin, safrole, sorbitol, spearmint, spearmint cream, strawberry, strawberry cream, stevia, sucralose, sucrose, saccharin sodium, saccharin, aspartame, acesulfame potassium, mannitol, talin, sucralose, sorbitol, Swiss cream, tagatose, orange, thaumatin, tutti fruitti, vanilla, walnut, watermelon, wild cherry, wintergreen, xylitol or any combination of these flavoring ingredients, such as anise-menthol, cherry-anise, cinnamon-orange, cherry-cinnamon, chocolate-mint, honey-lemon, lemon-lime, lemon-mint, menthol-eucalyptus, orange-cream, vanilla-mint and mixtures thereof. In one embodiment, the aqueous liquid dispersion may contain a sweetener or flavoring agent in a concentration range of about 0.0001% to about 10.0% by weight of the aqueous dispersion. In another embodiment, the aqueous liquid dispersion may include a sweetener or flavoring agent in a concentration range of about 0.0005% to about 5.0% wt % of the aqueous dispersion. In yet another embodiment, the aqueous liquid dispersion may include a sweetener or flavoring agent in a concentration range of about 0.0001% to 0.1 wt %, about 0.001% to about 0.01 wt %, or 0.0005% to 0.004% of the aqueous dispersion.

[0276] In addition to the additives listed above, liquid pregnenolone neurosteroid (eg, ganaxolone) formulations may also contain inert diluents commonly used in the art, such as water or other solvents, solubilizers, and emulsifiers.

[0277] In some embodiments, the pharmaceutical pregnenolone neurosteroid (e.g., ganaxolone) formulations described herein can be a self-emulsifying drug delivery system (SEDDS). An emulsion is a dispersion of an immiscible phase in another phase, usually in the form of droplets. Typically, an emulsion is produced by vigorous mechanical dispersion. In contrast to emulsions or microemulsions, SEDDS spontaneously forms an emulsion when added to excess water without any external mechanical dispersion or stirring. An advantage of SEDDS is that the droplets can be distributed throughout the solution with just gentle mixing. In addition, water or an aqueous phase can be added just before administration, which ensures the stability of unstable or hydrophobic active ingredients. Therefore, SEDDS provides an effective delivery system for oral and parenteral delivery of hydrophobic active ingredients. SEDDS can improve the bioavailability of hydrophobic active ingredients. Methods for producing self-emulsifying dosage forms are known in the art, including, but not limited to, for example, U.S. Patent Nos. 5,858,401, 6,667,048, and 6,960,563, each of which is specifically incorporated by reference.

[0278] Exemplary emulsifiers are ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol, dimethylformamide, sodium lauryl sulfate, sodium docusate, cholesterol, cholesterol esters, taurocholic acid, phosphatidylcholine, oils (e.g., cottonseed oil, peanut oil, corn germ oil, olive oil, castor oil, and sesame oil), glycerol, tetrahydrofurfuryl alcohol, polyethylene glycol, fatty acid esters of sorbitan, or mixtures of these substances, and the like.

[0279] In certain preferred embodiments, the liquid pharmaceutical formulation comprises ganaxolone, hydroxypropyl methylcellulose, polyvinyl alcohol, sodium lauryl sulfate, simethicone, methylparaben, propylparaben, sodium benzoate, citric acid and sodium citrate, pH 3.8-4.2. The suspension may comprise ganaxolone at a concentration of 50 mg / ml. The formulation may also comprise a pharmaceutically acceptable sweetener (e.g., sucralose) and / or a pharmaceutically acceptable flavoring (e.g., cherry). The formulation may be packaged in, for example, 120 mL, 180 mL, 240 mL or 480 mL bottles.

[0280] In certain preferred embodiments, an oral solid formulation as described and prepared in applicant's previous U.S. Patent No. 7,858,609, entitled "Solid Ganaxolone Formulations and Methods for the Making and Use Thereof" (incorporated herein in its entirety by reference) is used. Oral solid formulations (e.g., oral capsules or tablets) of pregnenolone neurosteroids can be prepared according to any suitable method.

[0281] For example, as disclosed in U.S. Pat. No. 7,858,609, an oral solid dosage form comprises stabilized particles comprising a pregnenolone neurosteroid (e.g., ganaxolone), a hydrophilic polymer, a wetting agent, and an effective amount of a complexing agent to stabilize particle growth after initial particle growth and endpoint is reached, the complexing agent being an organic small molecule having a molecular weight of less than 550 and containing a moiety selected from the group consisting of a phenol moiety, an aromatic ester moiety, and an aromatic acid moiety, wherein the stabilized particles have a particle volume weighted median diameter (D50) of about 50 nm to about 500 nm, and the complexing agent is present in an amount of about 0.05% to about 5% w / w based on the weight of the solid particles. The amount of the hydrophilic polymer may be about 3% to about 50% w / w based on the weight of the solid particles. The amount of the wetting agent may be about 0.01% to about 10% w / w based on the weight of the solid particles. The amount of pregnenolone neurosteroid (e.g., ganaxolone) may be from about 10% to about 80% (and in certain embodiments from about 50% to about 80%) based on the weight of the stabilized particles. When the particles are dispersed in simulated gastric fluid (SGF) or simulated intestinal fluid (SIF) at a concentration of 0.5 to 1 mg ganaxolone / mL and placed in a heating bath at 36° C. to 38° C. for 1 hour, the D50 of the stabilized particles increases by no more than about 150% compared to the volume weighted median diameter (D50) of the stabilized particles when the particles are dispersed in distilled water under the same conditions, wherein the volume weighted median diameter (D50) of the stabilized particles dispersed in SGF or SIF is less than about 750 nm. When the formulation is dispersed in 15 mL of SGF or SIF at a concentration of 0.5 to 1 mg ganaxolone / mL, the volume weighted median diameter (D50) of the stabilized particles increases by no more than about 150% compared to the D50 of the stabilized particles when the particles are dispersed in distilled water under the same conditions, wherein the volume weighted median diameter (D50) of the stabilized particles dispersed in SGF or SIF is less than about 750 nm. The solid stabilized particles can be combined with optional excipients and prepared in powder form for administration, or they can be incorporated into a dosage form selected from the group consisting of tablets or capsules. The complexing agent can be a paraben, benzoic acid, phenol, sodium benzoate, methyl anthranilate, etc. The hydrophilic polymer can be a cellulosic polymer, a vinyl polymer, and mixtures thereof. The cellulosic polymer can be a cellulose ether, such as hydroxypropyl methylcellulose. The vinyl polymer can be a polyvinyl alcohol, such as a vinyl pyrrolidone / vinyl acetate copolymer (S630). The wetting agent can be sodium lauryl sulfate, a pharmaceutically acceptable salt of docusate, and a mixture thereof. When the particles are incorporated into a solid dosage form, the solid dosage form may further comprise at least one pharmaceutically acceptable excipient, such as an ion dispersion regulator, a water-soluble spacer, a disintegrant, a binder, a surfactant, a plasticizer, a lubricant, a diluent, and any combination or mixture thereof.The water-soluble spacer can be a sugar or an ammonium salt, such as fructose, sucrose, glucose, lactose, mannitol. The surfactant can be, for example, polysorbate. The plasticizer can be, for example, polyethylene glycol. The disintegrant can be cross-linked sodium carboxymethylcellulose, cross-linked polyvinylpyrrolidone, a mixture thereof, and the like.

[0282] Capsules can be prepared, for example, by placing a pregnenolone-admixed neurosteroid (e.g., ganaxolone) formulation described above inside a capsule. In some embodiments, the ganaxolone formulation (non-aqueous suspensions and solutions) is placed in a soft gelatin capsule. In other embodiments, the ganaxolone formulation is placed in a standard gelatin capsule or a non-gelatin capsule, such as a capsule containing HPMC. In other embodiments, the ganaxolone formulation is placed in a spray capsule, where the capsule can be swallowed whole or the capsule can be opened and the contents can be sprayed on food before eating. The therapeutic dose can be divided into multiple (e.g., two, three, or four) capsules. In some embodiments, the entire dose of the ganaxolone formulation is delivered in capsule form.

[0283] Preferably, each capsule contains about 200 to about 600 mg ganaxolone, about 300 to about 600 mg ganaxolone, about 400 to about 600 mg ganaxolone, about 500 to about 600 mg ganaxolone, about 200 mg ganaxolone, about 250 mg ganaxolone, about 300 mg ganaxolone, about 500 mg ganaxolone, or about 600 mg ganaxolone.

[0284] In certain embodiments, each capsule contains 200 mg or 225 mg of ganaxolone, as well as hydroxypropyl methylcellulose, sucrose, polyethylene glycol 3350, polyethylene glycol 400, sodium lauryl sulfate, sodium benzoate, anhydrous citric acid, sodium methylparaben, microcrystalline cellulose, 30% simethicone emulsion, gelatin capsule, polysorbate 80, and sodium chloride. In some embodiments, the size of the capsule is 00.

[0285] Alternatively, the oral dosage form can be in the form of a controlled release dosage form, as described in US Pat. No. 7,858,609.

[0286] Suitable pregnenolone neurosteroid (e.g., ganaxolone) formulations may also be administered parenterally. In such embodiments, formulations suitable for parenteral, subcutaneous or intravenous injection may include physiologically acceptable sterile aqueous or non-aqueous solutions, dispersions, suspensions or emulsions, and sterile powders for reconstitution into sterile injectable solutions or dispersions. Examples of suitable aqueous and non-aqueous carriers, diluents, solvents or vehicles include water, ethanol, polyols (propylene glycol, polyethylene glycol, glycerol, cremophor, etc.), suitable mixtures thereof, vegetable oils (e.g., olive oil), and injectable organic esters such as ethyl oleate. In addition, ganaxolone may be dissolved using water-soluble β-cyclodextrins (e.g., β-sulfobutyl-cyclodextrin and 2-hydroxypropyl β-cyclodextrin) at concentrations >1 mg / ml. A particularly suitable cyclodextrin is a substituted β-cyclodextrin, For example, proper fluidity can be maintained by the use of coatings such as lecithin, by maintaining the desired particle size (in the case of dispersions), and by the use of surfactants. Ganaxolone formulations suitable for subcutaneous injection may also contain additives such as preservatives, wetting agents, emulsifiers, and dispensing agents. Prevention of microbial growth may be ensured by various antibacterial and antifungal agents such as parabens, benzoic acid, benzyl alcohol, chlorobutanol, phenol, sorbic acid, etc. It may also be desirable to include isotonic agents such as sugars, sodium chloride, etc. Drug absorption from injectable drug forms may be prolonged by the use of agents that delay absorption such as aluminum monostearate and gelatin. Ganaxolone suspensions designed for extended release via subcutaneous or intramuscular injection may avoid first-pass metabolism, and lower doses of ganaxolone may be required to maintain plasma levels of about 50 ng / ml. In such formulations, the particle size of the ganaxolone particles and the particle size range of the ganaxolone particles may be used to control the release of the drug by controlling the dissolution rate in fat or muscle.

[0287] Particularly useful injectable formulations are disclosed in Applicant's U.S. Patent Publication No. 2017 / 0258812 (U.S. Serial No. 15 / 294,135 filed October 14, 2016), which is incorporated herein by reference in its entirety. Other useful pregnenolone neurosteroid injectable formulations known to those skilled in the art may also be used.

[0288] VII. Combination Therapy

[0289] The present disclosure includes embodiments in which the neurosteroid is the sole active agent and embodiments in which the neurosteroid is administered in combination with one or more additional active agents. When used in combination with additional active agents, the neurosteroid and the additional active agent may be combined in the same formulation or may be administered separately. The neurosteroid may be administered while the additional active agent is administered (simultaneous administration) or may be administered before or after the additional active agent is administered (sequential administration).

[0290] The present disclosure includes embodiments in which the additional active agent is an anticonvulsant. Anticonvulsants include GABAA receptor modulators, sodium channel blockers, GAT-1 GABA transporter modulators, GABA transaminase modulators, voltage-gated calcium channel blockers, and peroxisome proliferator-activated alpha modulators.

[0291] The present disclosure includes embodiments in which an anesthetic or sedative is administered to a patient in combination with a neurosteroid. The anesthetic or sedative may be administered at a concentration sufficient to render the patient unconscious, such as a concentration sufficient to medically induce coma or a concentration effective to induce general anesthesia. Alternatively, the anesthetic or sedative may be administered at a lower dose effective for sedation but insufficient to cause loss of consciousness.

[0292] Benzodiazepines Benzodiazepines can be used as both anticonvulsants and anesthetics. The class includes diazepam, flunitrazepam, lorazepam, and midazolam.

[0293] In certain embodiments, neurosteroids and benzodiazepines Class (e.g., clobazam, diazepam, clonazepam, midazolam, chlordiazepoxide Acid, levetiracetam, felbamate, lamotrigine, fatty acid derivatives (e.g., valproic acid), carboxamide derivatives (rufinamide, carbamazepine, oxcarbazepine, etc.), amino acid derivatives (e.g., levocarnitine), barbiturates (e.g., phenobarbital), or a combination of two or more of the foregoing agents are administered simultaneously.

[0294] The neurosteroid nanoparticle injectable formulations of the present disclosure may be administered with another anticonvulsant. Anticonvulsants include many drug classes and overlap to some extent with coma-inducing, anesthetic, and sedative drugs that can be used in combination with neurosteroids. Anticonvulsants that can be used in combination with the neurosteroid nanoparticle injectable formulations of the present disclosure include aldehydes, such as paraldehyde; aromatic allyl alcohols, such as stiripentol; barbiturates, including those listed above, as well as methylphenobarbital and barbexaclone; benzodiazepines. The drug class includes alprazolam, bretazenil, bromazepam, brotizolam, chloridazepoxide, cinolazepam, clonazepam, chorazepate, clopazam, clotiazepam, cloxazolam, delorazepam, diazepam, estazolam, etizolam, ethylloflazepate, flunitrazepam, flurazepam, flutoprazepam, halazepam, ketazolam ketazolam, loprazolam, lorazepam, lormetazepam, medazepam, midazolam, nimetazepam, nitrazepam, nordazepam, oxazepam, phenenazepam, pinazepam, prazepam, premazepam, pyrazolam, quazepam, temazepam, and tatrazepam; bromides, such as potassium bromide; carboxamides, such as carbamazepine, oxcarbazepine, and eslicarbazepine acetate fatty acids, such as valproic acid, sodium valproate and divalproexsodium; fructose derivatives, such as topiramate; GABA analogs, such as gabapentin and pregabalin; hydantoins, such as ethotoin, phenytoin, mephenytoin and fosphenytoin; other neurosteroids, such as allopregnanolone, oxasolidinedione (e.g. paramethadione, trimethadione and ethadione), propionates (e.g. beclamide);Pyrimidinediones (e.g., primidone), pyrrolidines (e.g., brivaracetam, levetiracetam, and seletracetam), succinimides (e.g., ethosuximide), pensuximide, and mesuximide); sulfonamides, such as acetazoloamide, sultiame, methazolamide, and zonisamide; triazines, such as lamotrigine; ureas, such as pheneturide and phenacemide; NMDA antagonists, such as felbamate, and valproylamides, such as valpromide and valnoctamide; and perampanel. ;

[0295] VIII. Biomarkers

[0296] Predictive biomarkers are used to identify patient populations that are more homogeneous and have a higher propensity to respond to therapy.

[0297] Allopregnanolone, a metabolite of progesterone, is a positive allosteric modulator (PAM) of GABAA receptors. Individuals with TSC-related epilepsy may exhibit this allopregnanolone deficiency, supporting the hypothesis that treatment with a pharmaceutically acceptable pregnenolone (e.g., ganaxolone) may reduce seizure frequency and potentially improve additional symptoms of TSC-related epilepsy.

[0298] Thus, in certain embodiments, allopregnanolone sulfate (Allo-S) is used as a predictive biomarker for response to ganaxolone (an analog of allopregnanolone). In these embodiments, 2,500 pg mL -1 Allo-S plasma levels of 2,500 pg mL-1 or lower indicate that the subject may respond to and benefit from ganaxolone therapy; and Allo-S plasma levels above 2,500 pg mL-1 -1 Plasma levels of 0.1% or less indicate that the subject is unlikely to respond to ganaxolone therapy and a different therapeutic agent should be used.

[0299] 5. Examples

[0300] The following examples of formulations according to the present invention should not be construed as limiting the invention in any way and are merely samples of the various formulations described herein.

[0301] During the development of ganaxolone formulations, various formulations have been evaluated to establish a formulation that exhibits adequate pharmacokinetic ("PK") parameters and is suitable for development and commercialization. Other ganaxolone formulations used include ganaxolone mixed with sodium lauryl sulfate for administration in the form of various suspensions, ganaxolone mixed with hydroxypropyl-β-cyclodextrin (HP-β-CD) in solution, and ganaxolone mixed with β-cyclodextrin (β-CD), as well as 0.5 micron particles of ganaxolone in suspension and tablet formulations, as well as controlled release capsule formulations, and ganaxolone mixed with sulfobutyl ether cyclodextrin (HP-β-CD). Intravenous solution of dissolved ganaxolone. Development work resulted in an oral suspension comprising 0.3 micron ganaxolone immediate release particles as described in Example 1, and an oral capsule formulation comprising 0.3 micron ganaxolone immediate release particles as described in Example 2.

[0302] Example 1.

[0303] A 50 mg / ml ganaxolone suspension was prepared having the ingredients set forth in Table 1 below:

[0304] Table 1. Composition of 50 mg / ml ganaxolone suspension

[0305]

[0306] Table 2 shows the function of the excipients used in the 50 mg / ml ganaxolone suspension.

[0307] Table 2. Summary of the functions of the ingredients of 50 mg / ml ganaxolone suspension

[0308]

[0309] The oral bioavailability of a 50 mg / ml ganaxolone suspension is dependent on the rate and extent of dissolution of the nanoparticle drug in a relevant physiological environment. The particle size determination method and specifications are designed to ensure that the ganaxolone drug product does not agglomerate after dispersion in simulated gastrointestinal fluid.

[0310] A dispersive nanomilling process was used to reduce the particle size of ganaxolone and obtain stable ganaxolone nanoparticles. The nanomilling process involved the use of yttria-stabilized zirconium oxide (YTZ) grinding media under high energy agitation within a nanomill. To ensure consistent slurry particle size prior to dispersive nanomilling, Marinus developed a high energy rotor / stator pre-milling process using a VakuMix DHO-1. After nanomilling, the dispersion was diluted from 25% w / w ganaxolone to 20% w / w ganaxolone and filtered through a 20 micron filter, and stabilizers (methyl paraben, sodium benzoate, and anhydrous citric acid) were added to promote controlled growth to approximately 300 nm over a 5-10 day curing period at room temperature. The stabilized 300 nm nanoparticles showed good stability to particle growth in pediatric suspension drug products and encapsulated drug product forms. The stabilization process was controlled by the accurate addition and dissolution of parabens, which are water-soluble stabilizers. The solidification process was controlled by adjusting the holding time and temperature of the stable dispersion prior to suspension dilution (in the case of the 50 mg / ml ganaxolone suspension) or fluid bed bead coating (in the case of the 225 mg ganaxolone capsules described in Example 2).

[0311] The three dispersion batches prepared in the dispersion nanomilling scale-up study were diluted and stabilized by adding sodium methylparaben, sodium benzoate and anhydrous citric acid and cured for 7 days. After curing, the particle size was measured and is shown in Table 3.

[0312] Table 3. Stabilized dispersion particle size after 7 days of curing

[0313] batch D(10)(nm) D(50)(nm) D(90)(nm) Dispersion Batch 1 212 298 689 Dispersion Batch 2 208 289 539 Dispersion Batch 3 209 291 498

[0314] D = diameter

[0315] As shown, the D(50) particle size is stable within the specification of 250-450 nm.

[0316] Example 2.

[0317] Ganaxolone capsules (225 mg) were prepared having the ingredients set forth in Tables 4 and 5 below:

[0318] Table 4. Composition of 225 mg Ganaxolone Capsules IR Beads

[0319]

[0320] Table 5 summarizes the functions of the excipients used in the formulation of 225 mg ganaxolone capsules.

[0321] Table 5. Summary of the functions of the ingredients in 225 mg ganaxolone capsules

[0322]

[0323] Table 5. Summary of the functions of the ingredients in 225 mg ganaxolone capsules IR = Immediate Release

[0324] The manufacturing process used to prepare these capsules utilizes the same drug product specifications and the same quantitative composition, as well as the same nano-milled dispersion dilution and dispersion stabilization processes. Thus, the product of Example 2 utilizes the same stabilized dispersion intermediate as the product of Example 1. Sodium methylparaben can be replaced with methylparaben.

[0325] Table 6 summarizes the thirty-six month formal stability data results for ganaxolone immediate release (IR) 225 mg capsules:

[0326] Table 6. Thirty-six Month Formal Stability Data for Ganaxolone Immediate Release (IR) 225 mg Capsules (25°C / 60% RH)

[0327]

[0328] Table 6. Thirty-six Month Formal Stability Data for Ganaxolone Immediate Release (IR) 225 mg Capsules (25°C / 60% RH)

[0329]

[0330] Example 3

[0331] Summary of past epilepsy clinical trials in TSC-related epilepsy

[0332]

[0333] Example 4

[0334] A Phase 2 clinical study of ganaxolone was conducted in TSC-related epilepsy. Approximately 30 male and / or female patients aged 2 to 65 years (inclusive) with TSC-related epilepsy were screened and enrolled. Patients had a clinical diagnosis of TSC and a mutation in the TSC1 or TSC2 gene. Patients completed a daily diary regarding the effects of ganaxolone on seizures.

[0335] The treatment phase consisted of 2 parts: Part A and Part B. In Part A, patients received ganaxolone for 12 weeks (4 weeks of titration, 8 weeks of maintenance) in addition to their standard anti-seizure therapy. Patients were titrated to 63 mg / kg per day (maximum 1800 mg per day) over 4 weeks and then maintained at that dose for an additional 8 weeks. Ganaxolone was administered in increments of 15 mg / kg per day, up to 63 mg / kg per day, as an oral suspension given with food. Patients ≤28 kg were dosed as mg / kg. Patients >28 kg were dosed on a fixed schedule in increments of 450 mg per day to a maximum of 1800 mg per day. Ganaxolone was administered during the 4-week titration period as follows:

[0336]

[0337] Any patient who cannot tolerate the next dose step will be maintained at the lower dose step for a few more days before moving on to the next dose. If the next dose is still not tolerated, the patient can be dropped back to the next lower dose step. After the escalation period, a minimum dose of 33 mg / kg per day or 900 mg per day is usually required during the maintenance period unless the sponsor agrees to a lower dose due to tolerability issues (e.g., somnolence).

[0338] Dose changes, including alternative dosing paradigms (e.g., lower dose during the day and higher dose at night), should be discussed with the Sponsor's Medical Supervisor prior to or within 48 hours of making a dose change. Patients who discontinue ganaxolone prior to completion of the maintenance phase of Part A will continue to be followed per protocol and will be encouraged to maintain daily seizure diaries at a minimum until Part A is completed. These patients will also return for a safety follow-up assessment after 2 weeks of tapering.

[0339] Patients who have a ≥35% reduction in seizure frequency during the 12-week treatment period of Part A compared to baseline (e.g., a 4-week baseline period) and do not have any other contraindications to continued treatment who continue to receive ganaxolone during the OLE period of the study (Part B) may continue into Part B ("OLE eligible"). Part B is an open-label extension and lasts approximately 24 weeks. Therefore, Part B is available for patients who are responsive to ganaxolone as defined by the protocol. The main differences between Parts A and B are the duration of treatment, less frequent assessments, and the ability to change medication doses (both ganaxolone and other AED treatments, including starting and stopping other medications) based on evaluation of the patient's clinical course during Part B.

[0340] In Part B, patients on ganaxolone continued ganaxolone treatment at the dose they completed Part A. During Part B, the ganaxolone dose could be adjusted to a maximum of 600 mg TID for patients weighing >28 kg and to a maximum of 21 mg / kg TID for patients ≤28 kg. During Part B, doses of other antiseizure medications could be adjusted (including tapering and initiation of therapy) at the investigator's discretion.

[0341] Unless medically contraindicated, any patient who completes Part A but does not continue Part B or who completes Part B or discontinues GNX treatment will undergo a 2-week drug taper (gradual reduction) period and return to the site for a follow-up safety assessment after 2 weeks.

[0342] Patients who completed Part A and were deemed eligible for Part B continued to take ganaxolone at the same dose as when they completed Part A for an additional 24 weeks of treatment in Part B.

[0343] Patients may receive a lower dose during the day and a higher dose at night.

[0344] Unless there are other medical contraindications, such as a drug-induced rash, anyone discontinuing ganaxolone will undergo a 2-week taper.

[0345] Patients who discontinue ganaxolone before completion of the treatment maintenance period will continue to be followed per protocol and will be encouraged to maintain a daily seizure diary at least until completion of the treatment period in Part A. These patients will return for a safety follow-up assessment after 2 weeks of tapering.

[0346] Patients may return for a 2-week follow-up safety visit (eg, due to early termination of Part A or Part B, not participating in Part B, or completing Part B).

[0347] The primary efficacy endpoint was to evaluate the potency of ganaxolone for TSC-related epilepsy over a 12-week period and / or the % change in primary seizure frequency by the end of the 12-week treatment (titration and maintenance) period in Part A. Primary seizure types included focal motor seizures without impaired consciousness or perception, focal seizures with impaired consciousness or perception, focal seizures that evolved into bilateral generalized convulsive seizures, and generalized seizures with countable motor components.

[0348] Secondary objectives were to evaluate the safety and tolerability of ganaxolone as adjunctive therapy at the end of the 12-week treatment (titration and maintenance) period of Part A; to evaluate the pharmacokinetic (PK) parameters in patients receiving ganaxolone doses up to 63 mg / kg per day (or a maximum of 1800 mg per day) throughout the study; and to evaluate the long-term safety and tolerability of GNX when administered as adjunctive therapy throughout Part B.

[0349] Exploratory objectives are to assess changes in quality of life; to assess behavioral / neuropsychiatric changes in patients receiving ganaxolone as adjunctive therapy at the end of the 12-week treatment (titration and maintenance) period of Part A; to assess the relationship between efficacy response to ganaxolone and biomarker levels (e.g., neurosteroids); to assess the potential effects of ganaxolone on EEG activity; to assess changes in other types of seizures (non-primary) in TSC; to assess the effect of ganaxolone on primary seizure-free days; to assess the effect of ganaxolone on infant / epilepsy seizure-free days; to assess the effect of ganaxolone on infant / epilepsy seizure-free days.

[0350] A. Pharmacokinetic Assessment:

[0351] The PK population will include all patients who have received at least 1 dose of GNX and have collected at least 1 sample and obtained valid bioanalytical results. Samples will be drawn between 1 hour and 5 hours or between 4 hours and 8 hours after the last dose of Part A and Part B. Pharmacokinetic analysis will be limited to the concentration list because there is not enough concentration-time data available for non-compartmental analysis, such as Cmax, AUC or tmax. The pharmacokinetic data of this study can be used for population PK analysis conducted separately from this study and reported separately.

[0352] B. Neurosteroid serum and concomitant AED levels:

[0353] Blood samples will be drawn at the Screening Visit, Week 12 in Part A, and the Final Visit in Part B to measure neurosteroid levels (including allopregnanolone and related endogenous CNS-active steroids and sulfate metabolites, including, for example, allopregnanolone-sulfate).

[0354] Example 5. Biomarkers

[0355] Between May and November 2015, individuals with confirmed PCDH19 mutations and minimal seizure burden were enrolled at six centers in the United States and Italy (n=11). Change in seizure frequency (%) was assessed as the primary endpoint, and responders were defined as a 25% or greater reduction in seizure rate. Plasma neurosteroid levels were quantified using a previously published GC / MS method (doi:10.1016 / S0028-3908(99)00149-5). In two cases, baseline neurosteroid levels were not measured. In these cases, 6-month values ​​were used because no significant changes in neurosteroid levels over time were observed.

[0356] The median change from baseline to 28-day seizure frequency (all seizure types) was reduced by 26% for all participants (n=11). In this group, the mean plasma allopregnanolone-sulfate (Allo-S) concentration was 4,741 pg mL-1 (median=433 pg mL-1). Responder analysis and correlation with Allo-S demonstrated two discrete populations. Plasma Allo-S concentrations in responders (n=6) (≥25% reduction in seizure rate) and non-responders (n=5) were 501±430 pg mL-1 and 9,829±6,638 pg mL-1, respectively (mean±SD, p=0.05, Mann-Whitney).

[0357] When comparing seizure frequency at 6 months to baseline, the biomarker-positive group improved significantly (p=0.02, Wilcoxon), while the biomarker-negative (high Allo-S) group did not improve, but also did not significantly worsen (p=0.25, Wilcoxon). Retrospective analysis of biomarker-positive (n=7, Allo-S<2,500pg mL-1) vs. biomarker-negative (n=4, Allo-S>2,500pg mL-1) subjects yielded median % changes in seizure rate of -53.9% and 247%, respectively (p=0.006, Mann-Whitney). In addition, when comparing seizure frequency at 6 months to baseline, the biomarker-positive group improved significantly (p=0.02, Wilcoxon Signed Rank), while the biomarker-negative group did not significantly worsen (p=0.25, Wilcoxon Signed Rank).

[0358] Example 6. Case Report of a TSC Subject Enrolled in Part A of an Open-Label Phase 2 Trial 1

[0359] A subject with tuberous sclerosis complex (Subject 001) was enrolled in Part A of an open-label Phase 2 trial of ganaxolone (GNX) as an adjunctive treatment for epilepsy associated with tuberous sclerosis complex according to the study protocol described in Example 4. The subject's baseline seizure load was 132.41 per 28 days. Ganaxolone was administered orally three times a day at a maximum daily dose of 1800 mg for 11 weeks (78 days). The subject has completed the protocol and experienced a 64% reduction in seizures relative to baseline. This is the first subject to complete the protocol. Additional subjects are currently enrolled but have not yet completed the study.

[0360] Table 8. Summary of changes in seizure activity in TSC subjects

[0361]

[0362] Example 7. Preliminary pharmacokinetic and pharmacodynamic (PK / PD) analysis

[0363] A preliminary PK / PD analysis was performed to explore the relationship, if any, between ganaxolone levels and the percent change in severe motor seizure frequency.

[0364] a) Study design

[0365] A global, randomized, double-blind, placebo-controlled, Phase 3 clinical trial to evaluate the safety and efficacy of adjunctive ganaxolone for the treatment of seizures associated with CDD. Patients aged 2 to 21 years with pathogenic or likely pathogenic mutations in the CDKL5 gene, with impaired neurodevelopment and seizures refractory to at least 2 prior anti-seizure medications, who experienced at least 16 seizures per 28 days in the 2 months prior to screening were eligible for enrollment. The study consisted of a 6-week baseline followed by a 17-week double-blind phase (ganaxolone or placebo, 1:1). The dose of 50 mg / mL ganaxolone suspension was titrated to 63 mg / kg per day (21 mg / kg TID) over 4 weeks, not to exceed 1800 mg / d (600 mg TID) or the maximum tolerated dose. Blood draws for PK analysis were scheduled at Visit 3 (Week 5), Visit 4 (Week 9), and Visit 5 (Week 17).

[0366] b) Methods

[0367] The mean ganaxolone concentration for each subject was calculated during the double-blind phase using available results from up to three laboratory assays. The percent reduction in severe motor seizures (log eLinear regression was performed using standard deviation (SD) of the mean ganaxolone concentration ([percent reduction + 100]) as the dependent variable and natural log-transformed mean ganaxolone concentration as the single explanatory variable. Regression diagnostics included inspection of residuals and normal probability plots and identification of outliers and effect sizes. Cases with standardized residuals >2 or <-2 were excluded from the model and the regression was repeated.

[0368] The resulting samples were used to determine the Pearson correlation coefficient (using log-transformed values). In addition, the percentage reduction in seizures was compared among three tertiles representing the mean of low (N=13), medium (N=13), and high (N=12) ganaxolone concentrations per subject using the Kruskal-Wallis test.

[0369] The number of CNS-related adverse events suggestive of potential dose-related toxicity (somnolence, sedation, lethargy, attention disturbances, drooling, and hypotonia) in the ganaxolone-treated participants was tabulated as well as the occurrence and duration of the events, and the number of participants who experienced a CNS adverse event was calculated for each week during the double-blind period.

[0370] c) Results

[0371] The 44 participants with data on seizure reduction had at least one plasma ganaxolone level measured (mean + standard deviation = 103.5 + 79.2 ng / mL). In the linear regression with percent seizure reduction as the dependent variable and mean plasma ganaxolone concentration as the independent variable, six cases were identified as outliers due to adjusted residuals >2 or <-2. Repeating the linear regression after excluding those cases (N = 38) yielded an adjusted R2 of 0.227 (F(1,36) = 11.89), p = 0.001). The correlation coefficient between mean plasma ganaxolone concentration and percent reduction in severe motor seizures using the same samples was -0.499 (p = 0.001). A robust regression was performed including all observations (N = 44) that replicated the results of this analysis.

[0372] Mean and median percent reductions in severe motor seizures were calculated for the low, medium, and high ganaxolone concentration tertiles (Table 9). There was a statistically significant difference between the groups in percent reduction in severe motor seizure frequency (H(2)=9.087, p=0.011)( Figure 2 ). Post hoc pairwise comparisons of the sample distributions of the three groups showed statistically significant differences between the low GNX level group and the high GNX level group, but no other inter-group tests showed statistically significant differences.

[0373] Table 9. Tertiles Based on Mean Ganaxolone Plasma Concentrations

[0374]

[0375] In conclusion, the logarithm of plasma ganaxolone levels was inversely correlated with the percent change in severe motor seizure frequency. In patients with CDKL5 deficiency (CDD), increases in plasma GNX levels were associated with greater reductions in seizure frequency in the range of 27 to 333 ng / mL. Backward transformation of the logarithmic values ​​indicated that plasma concentrations of approximately 100 ng / mL (mean for the CDD population) predicted an approximately 40% reduction in seizures in participants in this study.

[0376] Modeled PK profiles based on previous Phase 1 studies suggest that TID dosing could provide GNX trough levels compared with BID dosing. Results from preliminary PK / PD analyses suggest that increases in plasma ganaxolone concentrations are associated with improved seizure reduction, and that concentrations of approximately 100 ng / mL are associated with meaningful changes in seizure frequency. Based on modeled PK profiles, TID dosing may result in plasma ganaxolone levels >100 ng / mL approximately 78% of the time over a 24-hour day, compared with only 53% with BID dosing. Although these analyses could not maintain randomization and therefore may not represent a causal effect of GNX on changes in seizure frequency, they suggest that TID dosing may provide an incremental anti-seizure benefit.

Claims

1. A method for treating tuberous sclerosis complex or tuberous sclerosis complex-associated epilepsy, the method comprising administering to a subject in need thereof a therapeutically effective amount of a pharmaceutically acceptable pregnenolone neurosteroid or a pharmaceutically acceptable salt thereof.

2. The method of claim 1, wherein the pregnenolone neurosteroid is ganaxolone.

3. The method of any one of the preceding claims, wherein the pregnenolone neurosteroid is administered three times daily.

4. The method of any one of claims 1-2, wherein the pregnenolone neurosteroid is administered twice daily.

5. The method of any of the preceding claims, wherein about 200 mg per day to about 1,800 mg per day of ganaxolone is administered to the subject.

6. The method of any one of claims 1-4, wherein up to about 1,800 mg of ganaxolone per day is administered to the subject.

7. The method of any preceding claim, wherein the ganaxolone is administered at about 1,500 mg of ganaxolone per day.

8. The method of any one of the preceding claims, wherein ganaxolone is administered in an amount of up to 63 mg / kg per day.

9. The method of any one of the preceding claims, wherein the pregnenolone neurosteroid is administered orally.

10. The method of any one of the preceding claims, wherein the pregnenolone neurosteroid is administered as an oral suspension.

11. The method of any one of the preceding claims, wherein the pregnenolone neurosteroid is administered as an oral capsule.

12. The method of any one of the preceding claims, wherein the tuberous sclerosis complex-related epilepsy is infantile spasms.

13. The method of any of the preceding claims, wherein the tuberous sclerosis complex-associated epilepsy is a focal impaired perception seizure.

14. The method of any of the preceding claims, wherein the tuberous sclerosis complex-associated epilepsy is a focal epileptic seizure.

15. The method of any of the preceding claims, wherein the tuberous sclerosis complex-related epilepsy is a generalized epileptic seizure.

16. The method of any one of the preceding claims, wherein administration of the pregnenolone neurosteroid reduces seizure frequency and / or seizure severity in the subject relative to baseline.

17. The method of any of the preceding claims, wherein administration of the pregnenolone neurosteroid reduces seizure frequency by about 20% or greater relative to baseline seizure frequency.

18. The method of any of the preceding claims, wherein administration of the pregnenolone neurosteroid reduces seizure frequency by at least about 35% or greater relative to baseline seizure frequency.

19. The method of any one of the preceding claims, wherein the subject is monitored by electroencephalography (EEG).

20. The method of any one of the preceding claims, wherein seizure activity in the subject is monitored by electroencephalography (EEG).

21. The method of any of the preceding claims, wherein ganaxolone is administered in an amount sufficient to provide about 70% or more of a ganaxolone plasma concentration of about 100 ng / mL in the subject over a 24 hour period.

22. The method of claim 21, wherein ganaxolone is administered three times a day.

23. The method of any one of the preceding claims, further comprising: measuring the level of endogenous neurosteroids in the subject prior to administering the pregnanolone neurosteroid, wherein a subject having low endogenous neurosteroids indicates that the subject will be responsive to the pregnanolone neurosteroid; and A therapeutically effective amount of the pregnenolone neurosteroid is administered to the subject having low endogenous neurosteroid levels.

24. The method of claim 23, wherein the endogenous neurosteroid is allopregnanolone-sulfate.

25. The method of any one of claims 23 or 24, wherein the low endogenous neurosteroid level is 2500 pg mL -1 or lower amounts.

26. The method of claim 25, wherein the pregnenolone neurosteroid is ganaxolone.

27. A method for treating tuberous sclerosis complex or tuberous sclerosis complex-associated epilepsy, the method comprising administering to a subject in need thereof a therapeutically effective amount of ganaxolone that produces a plasma concentration of ganaxolone of at least about 100 ng / ml by at least about 70% or more over a 24 hour period in a day.

28. The method of claim 27, wherein ganaxolone is administered three times daily.

29. The method of any one of claims 27 or 28, wherein ganaxolone is administered orally.

30. The method of any one of claims 27-29, wherein ganaxolone is administered as an oral suspension.

31. The method of any one of claims 27-29, wherein ganaxolone is administered as an oral capsule.

32. The method of any one of claims 27-31, wherein ganaxolone is administered in an amount of up to 63 mg / kg per day.

33. The method of any one of claims 27-32, wherein ganaxolone is administered in an amount of up to 1,800 mg per day.

34. The method of any one of claims 27-32, wherein ganaxolone is administered in an amount of up to 1,500 mg per day.

35. The method of any one of claims 27-34, wherein the tuberous sclerosis complex-associated epilepsy is infantile spasms, focal impaired perception seizures, focal seizures, or generalized seizures.

36. The method of any one of claims 27-35, wherein administration of ganaxolone reduces seizure frequency and / or seizure severity in the subject relative to baseline.

37. The method of any one of claims 27-35, wherein administration of ganaxolone reduces the frequency of severe motor activity in the subject relative to baseline.

38. The method of any one of claims 27-35, wherein administration of ganaxolone reduces seizure frequency by about 20% or greater relative to baseline seizure frequency.

39. The method of any one of claims 27-35, wherein administration of ganaxolone reduces seizure frequency by at least about 35% or greater relative to baseline seizure frequency.

40. The method of any one of claims 27-39, wherein the subject is monitored by electroencephalography (EEG).

41. The method of any one of claims 27-39, wherein seizure activity in the subject is monitored by electroencephalography (EEG).

42. A method of treating a subject having or suspected of having tuberous sclerosis complex-related epilepsy, the method comprising determining whether the subject has low levels of endogenous neurosteroids; and If the subject has low levels of the endogenous neurosteroid, a therapeutically effective amount of a pharmaceutically acceptable pregnenolone neurosteroid or a pharmaceutically acceptable salt thereof is administered to the subject.

43. The method of claim 42, wherein the endogenous neurosteroid is allopregnanolone-sulfate and the pregnenolone neurosteroid is ganaxolone.

44. The method of any one of claims 42 or 43, wherein the endogenous neurosteroid is allopregnanolone sulfate and the low level of the endogenous steroid is 2500 pg mL -1 or lower level.

45. The method of any one of claims 42-44, wherein the pregnenolone neurosteroid is a compound of formula IA: or a pharmaceutically acceptable salt thereof, wherein: X is O, S or NR 10 ; R 1 is hydrogen, hydroxy, -CH2A, optionally substituted alkyl, optionally substituted heteroalkyl, optionally substituted aryl, or optionally substituted arylalkyl; A is hydroxyl, O, S, NR 11 , an optionally substituted nitrogen-containing five-membered heteroaryl, an optionally substituted nitrogen-containing five-membered heteroaryl, or an optionally substituted nitrogen-containing bicyclic heteroaryl or bicyclic heterocyclyl, R 4 is hydrogen, hydroxy, oxo, optionally substituted alkyl or optionally substituted heteroalkyl, R 2 , R 3 , R 5 , R 6 and R 7 each independently absent, hydrogen, hydroxy, halogen, optionally substituted C1-C6 alkyl, optionally substituted C1-C6 alkoxy (e.g., methoxy), or optionally substituted heteroalkyl; R 8 and R 9 are each independently selected from the group consisting of hydrogen, C1-C6 alkyl (e.g., methyl), halogenated C1-C6 alkyl (e.g., trifluoromethyl) or C1-C6 alkoxy (e.g., methoxy), or R 8 and R 9 forming an oxo group; R 10 is hydrogen, hydroxy, optionally substituted alkyl, optionally substituted heteroalkyl, optionally substituted aryl, or optionally substituted arylalkyl, wherein each alkyl is C1-C 10 Alkyl, C3-C6 cycloalkyl, (C3-C6 cycloalkyl)C1-C4 alkyl, and optionally containing a single bond replaced by a double bond or a triple bond; Each heteroalkyl group is an independently selected -O-, -S-, -N(R 10 )-, -S(=O)- or -S(=O)2-substituted alkyl group, wherein R 10 is hydrogen, alkyl, or alkyl in which one or more methylene groups are replaced by -O-, -S-, -NH, or -N-alkyl; R 11 Is -H2 or -HR 12 ; R 12 It is a C1-C6 alkyl group or a C1-C6 alkoxy group.

46. ​​The method of claim 45, wherein the pregnenolone neurosteroid is selected from the group consisting of allopregnanolone, pregnenolone, 5-αDHP (5-α dihydroprogesterone), pregnanolone, dehydroepiandrosterone (DHEA), ganaxolone, 3α-hydroxy-3β-methyl-21-(4-cyano-1H-pyrazol-1'-yl)-19-nor-5β-pregnane-20-one, pharmaceutically acceptable salts of any of the foregoing, and combinations of any of the foregoing.

47. The method of any one of claims 42-46, wherein the pregnenolone neurosteroid is administered orally.

48. The method of any one of claims 42-47, wherein up to about 63 mg / kg of ganaxolone per day is administered to the subject.

49. The method of claim 48, wherein up to about 33 mg / kg of ganaxolone per day is administered to the subject.

50. The method of any one of claims 42-47, wherein up to about 1,800 mg of ganaxolone per day is administered to the subject.

51. The method of any one of claims 42-47, wherein up to about 1,500 mg of ganaxolone per day is administered to the subject.

52. The method of claims 42-51, wherein the tuberous sclerosis complex-related epilepsy is selected from the group consisting of focal motor seizures, focal epileptic seizures, and generalized epileptic seizures.

53. The method of any one of claims 42-52, wherein administration of ganaxolone reduces seizure frequency and / or seizure severity in the subject relative to baseline.

54. The method of any one of claims 42-53, wherein administration of ganaxolone reduces the frequency of severe motor activity in said subject relative to baseline.

55. The method of any one of claims 42-53, wherein administration of ganaxolone reduces seizure frequency by about 20% or greater relative to baseline seizure frequency.

56. The method of any one of claims 42-53, wherein administration of ganaxolone reduces seizure frequency by at least about 35% or greater relative to baseline seizure frequency.

57. The method of any one of claims 42-56, wherein the subject is monitored by electroencephalography (EEG).

58. The method of any one of claims 42-56, wherein seizure activity in the subject is monitored by electroencephalography (EEG).

59. The method of claim 42, wherein the endogenous neurosteroid is allopregnanolone and the low level of allopregnanolone is 200 pg mL -1 or lower.

60. The method of claim 42, wherein the endogenous neurosteroid comprises allopregnanolone, allopregnanolone-sulfate, pregnenolone, pregnenolone-sulfate, and mixtures thereof.

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