RU2803464C2 - Neuroactive steroid solutions and methods for administration - Google Patents

Abstract

FIELD: allopregnanolone solution.

SUBSTANCE: pharmaceutically acceptable dilute aqueous solution of allopregnanolone contains (i) the first solution subjected to heat treatment and containing allopregnanolone, sulfo-butyl ether of beta-cyclodextrin and citrate buffer and (ii) diluent; wherein the diluted aqueous solution is made by mixing the first heat treated solution with a diluent, and wherein the first solution is autoclaved by subjecting it to cycles of heat sterilization for at least 10, 15, 20, 30 or 40 minutes at a temperature of 110 to 150 °C.

EFFECT: ensuring stable solution.

7 cl, 8 dwg, 16 tab, 8 ex

Description

RELATED APPLICATIONS

This application claims priority under 35 USC § 119(e) of US Provisional Application No. 62/181,550, filed June 18, 2015, the contents of which are incorporated herein by reference in their entirety.

BACKGROUND OF THE ART

Homogeneous solutions (eg, aqueous solutions) containing a therapeutic agent, such as a neuroactive steroid described herein, allow administration to a person in need through various routes of administration (eg, oral, parenteral (eg, intravenous, intramuscular, subcutaneous) delivery). Neuroactive steroids are typically highly lipophilic compounds with low intrinsic solubility in water. Particularly for intravenous administration, solutions are in most cases pH-stable or chemically stable, preferably over long periods of time.

SUMMARY OF THE INVENTION

Provided herein is a pharmaceutically acceptable aqueous solution containing (eg, consisting essentially of, consisting of) a neuroactive steroid (eg, allopregnanolone), beta-cyclodextrin sulfobutyl ether, and a buffer; wherein the solution is a stable solution at a pH of about 3 to about 9 (eg, about 5 to about 7, about 5.5 to about 6.5) for at least 1, 2, 3, 4 weeks; 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 months; 1, 2, 3 years or more.

In some embodiments, the solution is a stable solution at a pH of about 3 to about 9 (e.g., about 5 to about 7, about 5.5 to about 6.5) for at least 1, 2, 3, 4 weeks; 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 months; 1, 2, 3 years or more at temperatures between approximately 2°C and approximately 8°C.

In some embodiments, the solution is a stable solution at a pH of about 3 to about 9 (e.g., about 5 to about 7, about 5.5 to about 6.5) for at least 1, 2, 3, 4 weeks; 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 months; 1, 2, 3 years or more at a temperature of from about 0°C to about 45°C (for example, from about 0°C to about 30°C, from about 15°C to about 25°C).

Also provided herein is a pharmaceutically acceptable aqueous solution containing (eg, consisting essentially of, consisting of) a neuroactive steroid (eg, allopregnanolone), beta-cyclodextrin sulfobutyl ether, and a buffer; where: the buffer is present at a concentration of at least 0.1 mM (for example, at least 0.5 mM, 1 mM, 2 mM, 5 mM or 10 mM).

Also provided herein is a pharmaceutically acceptable aqueous solution containing (eg, consisting essentially of, consisting of) a neuroactive steroid (eg, allopregnanolone), beta-cyclodextrin sulfobutyl ether, and a buffer; where: the solution remains substantially free of (e.g. meets product regulatory requirements with less than 3, 2, 1, 0.5, 0.3, 0.2, 0.1% w/w) impurities (e.g. , a solution essentially free of (for example, meets the requirements of product regulatory documentation with less than 3, 2, 1, 0.5, 0.3, 0.2, 0.1% w/w) impurities at room temperature in for at least 1, 2, 3, 4 weeks, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 months, 1, 2, 3 years or more). In some embodiments, the solution is at least 97% pure for at least 1, 2, 3, 4 weeks; 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 months; 1, 2, 3 years or more). For example, the solution has a found base content value of 90-110 for at least 1, 2, 3, 4 weeks; 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 months; 1, 2, 3 years or more).

In some embodiments, the solution remains substantially free (e.g., less than 3, 2, 1, 0.5, 0.3, 0.2, 0.1% w/w) of impurities for at least 1. 2, 3, 4 weeks; 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 months; 1, 2, 3 years or more at temperatures between approximately 2°C and approximately 8°C.

In some embodiments, the solution remains substantially free (e.g., less than 3, 2, 1, 0.5, 0.3, 0.2, 0.1% w/w) of impurities for at least 1. 2, 3, 4 weeks; 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 months; 1, 2, 3 years or more at a temperature of from about 0°C to about 45°C (for example, from about 0°C to about 30°C, from about 15°C to about 25°C).

In some embodiments, the buffer in solution is present at a concentration of from about 5 to 10 mM. In some embodiments, the buffer in solution is present at a concentration of from about 0.1 to about 4 mM. In some embodiments, the buffer in solution is present at a concentration of about 0.1, about 0.5, about 1.67, or about 3.3 mM.

In some embodiments, the solution further contains a diluent.

In some embodiments, the solution is suitable for parenteral use.

In some embodiments, the solution is homogeneous.

In some embodiments, the neuroactive steroid is selected from pregnanolone, ganaxolone, alfadalone, alfaxalone, and allopregnanolone. In some embodiments, the neuroactive steroid is ganaxolone. In some embodiments, the neuroactive steroid is allopregnanolone.

In some embodiments, the neuroactive steroid is estrol.

In some embodiments, the neuroactive steroid assay content is reduced by less than 10% when stored for 1, 2, 3, 4, 5, 6, 7 days; 1, 2, 3, 4, 5, 6 months or more, or 1, 2, 3 years or more at room temperature (e.g. 23 +/- 2°C).

In some embodiments, the neuroactive steroid assay content is reduced by less than 10% when stored for 1, 2, 3, 4, 5, 6, 7 days; 1, 2, 3, 4, 5, 6 months or more, or 1, 2, 3 years or more at a temperature of about 2 to about 8°C.

In some embodiments, the assayed neuroactive steroid content is reduced by less than 10% when stored for at least 10, 15, 20, 25, 30, 40, 45 minutes or more at a temperature of from about 110 to about 130°C ( for example, from about 110 to about 125°C, for example, 122 +/- 2°C).

In some embodiments, the solution has an assayed base content of 100 +/- 10%.

In some embodiments, the solution is chemically stable. In some embodiments, the solution is physically stable. In some embodiments, the solution is pH stable.

In some embodiments, the solution contains less than 0.5, 0.4, 0.3, 0.2, or 0.1% w/w. a degradation product of a neuroactive steroid (for example, allopregnanolone). In some embodiments, the degradation product is an oxidation product of a neuroactive steroid (eg, an oxidation product of allopregnanolone, 136 ). In some embodiments, the degradation product is a racemate or epimer of a neuroactive steroid (eg, the epimeric product of allopregnanolone, 1269 ). In some embodiments, the amount of neuroactive steroid degradation product (e.g., racemate or epimer, or neuroactive steroid oxidation product) present in the solution is substantially the same (e.g., meets product regulatory requirements of +/- 0.1, 0.2 , 0.5, 1, 2% w/w) for 1, 2, 3, 4, 5, 6, 7 days or more; 1, 2, 3, 4 weeks or more; 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 months or more; 1, 2, 3 years or more. In some embodiments, the amount of neuroactive steroid degradation product present in the solution is less than 0.1% w/w. for 1, 2, 3, 4, 5, 6, 7 days or more; 1, 2, 3, 4 weeks or more; 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 months or more; 1, 2, 3 years or more.

In some embodiments, the pH of the solution is substantially the same (e.g., meets product regulatory requirements, pH is +/- 1.2, 1, 0.8, 0.5, 0.3 or less) for 1, 2, 3, 4, 5, 6, 7 days or more; 1, 2, 3, 4 weeks or more; 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 months or more; 1, 2, 3 years or more.

In some embodiments, the pH of the solution is from about 3 to about 9 (e.g., from about 5 to about 7, from about 5.5 to about 6.5) for 1, 2, 3, 4, 5, 6, 7 days or more; 1, 2, 3, 4 weeks or more; 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 months or more; 1, 2, 3 years or more.

In some embodiments, the solution is at a temperature between 3°C and 37°C. In some embodiments, the solution is at a temperature of 0°C to 45°C (eg, 0°C and 30°C, eg, 15°C to 25°C). In some embodiments, the solution is at room temperature (eg, 25°C).

In some embodiments, the buffer is selected from an acidic, basic, or neutral buffer. In some embodiments, the buffer is selected from an acidic or neutral buffer. In some embodiments, the buffer has a pKa of from about 2 to about 9. In some embodiments, the buffer contains a monoprotic acid. In some embodiments, the buffer contains a polybasic acid (eg, citrate). In some embodiments, the buffer is selected from the group consisting of citrate, phosphate, acetate, lactate, gluconate, malate, succinate, Tris, histidine and tartrate, and mixtures thereof.

In some embodiments, the buffer is a citrate buffer. In some embodiments, the citrate buffer has a pH of about 3 to about 8 (e.g., about 4.5 to about 7.0, about 5.5 to about 6.5, about 5.0 to about 6.0) .

In some embodiments, the buffer is a phosphate buffer. In some embodiments, the phosphate buffer has a pH of about 1 to about 9 (e.g., about 4.5 to about 7.0, about 5.5 to about 6.5, about 5.0 to about 6.0) .

In some embodiments, the buffer is a solution of one or more substances (eg, a salt of a weak acid and a weak base, a mixture of a weak acid and a salt of a weak acid and a strong base).

In some embodiments, the buffer is selected from 4-2-hydroxyethyl-1-piperazineethanesulfonic acid (HEPES), 2-{[tris(hydroxymethyl)methyl]amino}ethanesulfonic acid (TES), 3-(N-morpholino)propanesulfonic acid (MOPS ), piperazine-N,N'-bis(2-ethanesulfonic acid) (PIPES), dimethylarsinic acid (cacodylate), citrate (e.g. sodium citrate saline), 2-(N-morpholino)ethanesulfonic acid (MES), phosphate (e.g. PBS, D-PBS), succinate (i.e. 2(R)-2-(methylamino)succinic acid), acetate, dimethylglutarate, maleate, imidazole, N-(2-acetamido)-2-aminoethanesulfonic acid (ACES), N,N-bis(2-hydroxyethyl)-2-aminoethanesulfonic acid (BES), bicine, Bis-Tris, borate, N-cyclohexyl-3-aminopropanesulfonic acid (CAPS), glycine, 3-[4- (2-hydroxyethyl)-1-piperazinyl]propanesulfonic acid (HEPPS or EPPS), N-[tris(hydroxymethyl)methyl]-3-aminopropanesulfonic acid, [(2-hydroxy-1,1-bis(hydroxymethyl)ethyl)amino ]-1-propanesulfonic acid (TAPS), tricine, Tris, Tris base, Tris buffer, Tris-glycine, Tris-HCl, collidine, veronal acetate, N-(2-acetamido)iminodiacetic acid; N-(carbamoylmethyl)iminodiacetic acid (ADA), β-hydroxy-4-morpholinepropanesulfonic acid, 3-morpholino-2-hydroxypropanesulfonic acid (MOPSO), cholamine chloride, 3-(N,N-bis[2-hydroxyethyl]amino)- 2-hydroxypropanesulfonic acid (DIPSO), acetamidoglycine, 3-{[1,3-dihydroxy-2-(hydroxymethyl)-2-propanyl]amino}-2-hydroxy-1-propanesulfonic acid (TAPSO), piperazine-N,N -bis(2-hydroxypropanesulfonic acid) (POPSO), N-(2-hydroxyethyl)piperazine-N'-(2-hydroxypropanesulfonic acid) (HEPPSO), N-cyclohexyl-2-aminoethanesulfonic acid (CHES), 2-amino- methyl 1,3-propanediol (AMPd) and glycinamide. In some embodiments, the buffer contains piperazine (eg, PIPES, HEPES, POPSO, EPPS).

In some embodiments, the buffer contains a non-metal complexing compound (eg, MES, MOPS, PIPES).

In some embodiments, the buffer has a pH suitable for injection (eg, safe, tolerable, non-irritating).

In some embodiments, the buffer is within its effective buffer capacity range.

In some embodiments, the buffer is citrate. In some embodiments, the citrate buffer is present at a concentration of from about 1 to about 100 mM or more. In some embodiments, the citrate buffer is present at a concentration of 5, 10, 20, 50, 100 mM or more.

In some embodiments, the buffer is a phosphate. In some embodiments, the phosphate buffer is present at a concentration of from about 1 to about 100 mM or more. In some embodiments, the phosphate buffer is present at a concentration of 5, 10, 20, 50, 100 mM or more.

In some embodiments, the pH of the solution is from about 3 to about 9 (e.g., preferably from about 5 to about 9, from about 4.5 to about 7.0, from about 5.0 to about 6.5).

In some embodiments, the neuroactive steroid is present in an amount of 0.1, 0.5, 1, 1.25, 2.5, 3.75, 5, 6.25, 7.5, 8, 9, or 10 mg/ml, or more. In some embodiments, the neuroactive steroid is combined with 2.5, 5, 6, 7.5, 10, 15, 20, 30% w/v. or more sulfobutylether-β-cyclodextrin.

In some embodiments, the molar ratio of neuroactive steroid to sulfoalkyl ether-β-cyclodextrin is approximately 1:1, 1:2, 1:3, 1:4, 1:5, 1:6, 1:7, 1:8, 1:9, 1:10, 1:20: 1:30, 1:50, 1:75, 1:100, 1:120 or more. In some embodiments, the molar ratio of neuroactive steroid to sulfoalkyl ether-β-cyclodextrin is about 0.1, 0.05, 0.03, 0.02, 0.01, 0.008, 0.005 or less. In some embodiments, the neuroactive steroid is allopregnanolone. In some embodiments, the molar ratio of allopregnanolone to sulfoalkyl ether-β-cyclodextrin is about 1:1, 1:2, 1:3, 1:4, 1:5, 1:6, 1:7, 1:8, 1:9, 1:10, 1:20: 1:30, 1:50, 1:75. In some embodiments, the molar ratio of allopregnanolone to sulfoalkyl ether-β-cyclodextrin is approximately 1:1, 1:2, 1:3, 1:4, 1:5, 1:6, 1:7, 1:8, 1:9 , 1:10, 1:20. In some embodiments, the molar ratio of allopregnanolone to sulfoalkyl ether-β-cyclodextrin is from about 1:1 to about 1:60 (e.g., from about 1:1 to about 1:20, from about 1:1 to about 1:15). In some embodiments, the molar ratio of allopregnanolone to sulfoalkyl ether-β-cyclodextrin is from about 1:3 to about 1:20 (for example, from about 1:5 to about 1:10). In some embodiments, the solution further contains a surfactant.

In some embodiments, the solution further contains a chelating agent.

In some embodiments, the solution further contains a preservative.

In some embodiments, the solution further contains an isotonizing agent. In some embodiments, the isotonic agent is present in an amount to achieve isotonicity.

In some embodiments, the solution is sterilized by heat treatment.

In one aspect, provided herein is a pharmaceutically acceptable aqueous solution containing (eg, consisting essentially of, consisting of) a neuroactive steroid, beta-cyclodextrin sulfobutyl ether, and a buffer; wherein the composition contains less than 3, 2, 1, 0.5, 0.3, 0.2, 0.1% wt./mass. impurities (e.g., the solution is substantially free of (e.g., less than 3.2.1, 0.5, 0.3, 0.2, 0.1% w/w) impurities for at least 1, 2 , 3, 4 weeks, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 months, 1, 2, 3 years or more).

In one aspect, provided herein is a method of preparing a stable solution containing allopregnanolone, the method comprising contacting allopregnanolone with a pharmaceutically acceptable aqueous solution containing (eg, consisting essentially of, consisting of) beta-cyclodextrin sulfobutyl ether and a buffer.

In some embodiments, the solution is at a temperature of from about 0°C to about 60°C (eg, from about 20°C to about 50°C, from about 35°C to about 45°C). In some embodiments, the solution is at room temperature (eg, 35-45°C).

In some embodiments, the solution is chemically stable.

In some embodiments, the solution is autoclaved (e.g., subjected to heat sterilization cycles, e.g., subjected to at least 10 (e.g., at least 15, 20, 30, 40 minutes) of heat (e.g., 110 to 150°C (e.g. , 121 to 123°C)) In some embodiments, the solution is at a temperature of 110 to 150°C (eg, 121 to 123°C).

In some embodiments, the amount of neuroactive steroid degradation product present in the solution is less than 0.1% w/w. for 1, 2, 3, 4, 5, 6, 7 days or more; 1, 2, 3, 4 weeks or more; 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 months or more; 1, 2, 3 years or more.

In one aspect, provided herein is a pharmaceutically acceptable aqueous solution containing (eg, consisting essentially of, consisting of) a neuroactive steroid (eg, allopregnanolone), beta-cyclodextrin sulfobutyl ether, and a buffer; wherein the solution is a stable solution at a pH of about 3 to about 9 (e.g., about 5 to about 7, about 5.5 to about 6.5) for at least 5 minutes, such as at least 10, 15, 20, 25, 30, 35, 40, 45, 50, 55 or 60 minutes or more at a temperature of from about 120°C to about 124°C; or the buffer is present at a concentration of at least 0.1 mM; or the solution remains substantially free (e.g., meets product regulatory requirements with less than 3, 2, 1, 0.5, 0.3, 0.2, 0.1% w/w) impurities for up to at least 5 minutes, such as at least 10, 15, 20, 25, 30, 35, 40, 45, 50, 55 or 60 minutes or more at a temperature of from about 120°C to about 124°C.

In one aspect, provided herein is a method of parenteral administration, the method comprising mixing a primary solution containing allopragnanolone (e.g., the solution described herein) with a diluent (e.g., water or injection, or saline) to form a diluted solution ; and parenterally administering the diluted solution to the subject. In some embodiments, the primary solution is diluted with two parts diluent to one part primary solution. In some embodiments, the primary solution is diluted with nine parts diluent to one part primary solution.

In one aspect, provided herein is a method of preparing an aqueous solution containing a neuroactive steroid, a beta-cyclodextrin sulfoalkyl ether (e.g., beta-cyclodextrin sulfobutyl ether or sulfobutyl ether-beta-cyclodextrin) and a buffer in which the solution is mixed (e.g., by homogenization high shear) to produce a solution substantially free (e.g., less than about 1, 0.5, 0.2, 0.1% w/v) of solids (e.g., free of any solids substances with particle sizes of 0.22, 0.45, 1 micron or more in diameter).

In some embodiments, the solution is mixed with a suitable mixing device or method. In some embodiments, the mixing device is a high shear paddle mixer, a rotor-stator mixer, a homogenizer, an ultrasonic device, or a microfluidizer.

In some embodiments, the rotor-stator mixer rotates at a speed of 2,000 to 18,000 rpm. In some embodiments, the homogenizer operates at a pressure of 1000 to 5000 pounds per square inch.

In some embodiments, the solution is mixed using a suitable high shear mixing device, such as a rotor/stator device, homogenizer, microfluidizer, or sonicator. In some embodiments, the high shear mixing device (eg, a rotor/stator device, homogenizer, microfluidizer, or sonicator uses built-in high shear modules).

In some embodiments, the method is used for a suitable period of time to achieve solubilization (eg, at least 15, 30, 60 or more minutes).

In some embodiments, the solution is diluted with a diluent, for example, to form a mixture.

In one aspect, provided herein is a closed container containing a neuroactive steroid, a beta-cyclodextrin sulfoalkyl ether (eg, beta-cyclodextrin sulfobutyl ether or beta-cyclodextrin sulfobutyl ether) and a buffer; additionally containing a gaseous layer containing mainly (for example, containing more than 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 99.1, 99.5, 99.98, 99.99% inert gas (e.g. nitrogen, argon).

In some embodiments, the gaseous layer contains less than 21, 20, 17, 15, 12, 10, 8, 5, 3, 1, 0.5, 0.2, 0.1, 0.05% oxygen gas (e.g., not contains oxygen).

In some embodiments, the container includes a bottle, stopper, or cap.

In some embodiments, the container is a pre-filled syringe. In some embodiments, the container is a glass container. In some embodiments, the container is a plastic container. In some embodiments, the plastic container and low oxygen content are provided through a packaging wrapper (eg, a multi-layer aluminum foil pouch).

In one aspect, provided herein is a method of treating a subject (e.g., a subject suffering from a disease or disorder described herein (e.g., depression (e.g., postpartum depression), the method comprising administering an aqueous solution or mixture described herein, thereby thereby exposing the subject to treatment.

In one aspect, provided herein is a method of treating a subject (e.g., a subject suffering from a disease or disorder described herein (e.g., depression (e.g., postpartum depression), the method comprising administering one part of an aqueous solution described herein to two parts of the diluents described herein (eg, WFI), thereby exposing the subject to treatment.

In one aspect, provided herein is a method of treating a subject (e.g., a subject suffering from a disease or disorder described herein (e.g., depression (e.g., postpartum depression), the method comprising administering one part of an aqueous solution described herein to nine parts of the diluents described herein (eg, WFI), thereby exposing the subject to treatment.

DETAILED DESCRIPTION OF FIGURES

Figure 1 . Illustrative description of allopregnanolone degradation processes.

Figure 2 . Illustrative description of the solubility of allopregnanolone in SBECD.

Figure 3 . Stability of allopregnanolone in phosphate buffer at time points = 0, 4, 6 and 12 weeks ( A ) Area under the curve at 40°C; ( B ) Area under the curve at 60°C.

Figure 4 . Stability of allopregnanolone in citrate buffer at time points = 0, 4, 6 and 12 weeks ( A ) Area under the curve at 40°C; ( B ) Area under the curve at 60°C.

Figure 5 . Formation of 136 over time in different buffers ( A ) at 40°C; ( B ) at 60°C.

Figure 6 . Illustrative LC-MS characterization of 1269.

Figure 7 . Illustrative characteristics 136.

Figure 8 . Quantitative analysis of the unbuffered allopregnanolone formulation performed after 12 weeks ( A ) at 40°C and ( B ) at 60°C.

DETAILED DESCRIPTION OF SOME OPTIONS FOR IMPLEMENTING THE INVENTION

Disclosed herein are aqueous solutions or mixtures containing a neuroactive steroid, cyclodextrin and a buffer; methods of their use and administration, methods of their preparation and containers containing solutions or mixtures.

Definitions

As used herein, the terms “stabilized” and “stable” aqueous solution as described herein (eg, an aqueous solution containing a neuroactive steroid) refer to a solution that is “chemically stable” and “physically stable.” A solution containing a neuroactive steroid is chemically stable if the neuroactive steroid does not undergo chemical conversion or degradation (eg, racemization, epimerization, oxidation). For example, a chemically stable neuroactive steroid, e.g., in solution, will not racemize or epimerize (e.g., at receptive positions (e.g., racemize or epimerize at position C17 in a neuroactive steroid)) or oxidize (e.g., receptive positions (e.g., oxidize at position C3 of a neuroactive steroid)) or recover (e.g., at susceptible positions (e.g., recover at position C21 of a neuroactive steroid), e.g., over a period of time (e.g., within 1, 2, 3, 4, 5, 6, 8, 10, 12, 14, 16, 18, 20, 24 hours or more, 1, 2, 3, 4, 5, 6, 7 days or more, 1, 2, 3, 4 weeks or more, 1, 2, 3 , 4, 5, 6, 8, 10, 12 months or more, 1, 2, 3, 4, 5 years or more) or at temperatures (e.g., ambient or elevated). As used herein, the term "pH- "stable" refers to a solution in which the pH of the solution is substantially the same (for example, +/- 1.2, 1, 0.8, 0.5, 0.3 or less) for 1, 2, 3, 4, 5, 6, 7 days or more, 1, 2, 3, 4 weeks or more, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 months or more; 1, 2, 3, 4, 5 years or more, for example at least 1, 2, 3, 4, 5, 6, 7, 8 weeks; 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 months or more; 1, 2, 3, 4, 5 years or more. A solution containing a neuroactive steroid is "physically stable" if the solution does not undergo physical changes, such as changes in color or particle level, for example, over a period of time or at different temperatures. For example, a stable aqueous solution containing a neuroactive steroid is chemically stable and physically stable during manufacture (eg, preparation, mixing, filling, labeling and sterilization), transportation or storage.

"Analysis for the content of the main substance" in the context of the present description refers to a specific stability determination procedure that determines the content of the drug substance. For example, the analysis for the content of the main substance may be a chromatographic method (eg, HPLC) using a standard sample.

"Impurities" as used herein refers to organic and inorganic impurities and residual solvents. For example, impurities are racemized or epimerized (e.g., at receptive positions (e.g., racemized or epimerized at the C17 position of a neuroactive steroid)), or oxidized (e.g., at receptive positions (e.g., oxidized at the C3 position of a neuroactive steroid)), or reduced ( for example, at receptive positions (eg, reduced at the C21 position of a neuroactive steroid) a neuroactive steroid. A solution is free of impurities when it contains less than 3, 2, 1, 0.5, 0.3, 0.2 or 0.1% w/w impurities.

"Purity" as used herein refers to the absence of impurities, eg, in a solution or composition relative to its original state (eg, at time=0).

"Sterilization" as used herein refers to filling under aseptic conditions (eg, aseptic sterilization) or terminal sterilization.

Solutions

The aqueous solutions or mixtures described herein contain a neuroactive steroid. Neuroactive steroids are typically highly lipophilic compounds with low intrinsic solubility in water. Cyclodextrins, for example the cyclodextrins according to the present description, can help stabilize compounds, for example, neuroactive steroid compounds. Surprisingly, it has been discovered that some unbuffered solutions of neuroactive steroids containing sulfobutylether-β-cyclodextrin are not pH stable. For example, the pH of solutions (eg, unbuffered solutions) is from about 3 to about 9, such as from about 5 to about 8, such as from about 5.5 to about 7.5. In addition, it has been found that the pH of solutions (eg, unbuffered solutions) deviates (eg, the pH does not remain within the desired pH range). Some buffers have been found to be well suited for combined use with unbuffered neuroactive steroid solutions containing sulfobutylether-β-cyclodextrin, for example in a clinical setting, as the pH of the solution or mixture is not changed (for example, the pH remained in the range of 5.5 to 7 ,5). Surprisingly, it was found that some buffered solutions or mixtures were more stable than some unbuffered solutions when stored for 1, 2, 3, 4, 5, 6 or more months at temperatures ranging from 4 to 40°C. In addition, it has surprisingly been discovered that some of the buffered solutions or mixtures described herein are stable (e.g., physically and chemically stable), for example, at high temperatures (e.g., 121°C) for short periods of time, for processes sterilization (for example, the sterilization processes described in this document). For example, some buffered solutions or mixtures described herein are stable (e.g., physically and chemically stable) at high temperatures (e.g., 121°C) for 10, 20, 30, 40, 50, 60, 70, 80 , 90 minutes or more. In addition, it has surprisingly been discovered that certain buffered solutions or mixtures of neuroactive steroids described herein are less susceptible to the formation of impurities over a range of temperatures and time. For example, some buffered solutions or mixtures of neuroactive steroids may have lower impurity levels (eg, 2% w/v or lower) than some unbuffered solutions of neuroactive steroids over a range of storage temperatures or times.

Some buffered solutions or mixtures of neuroactive steroids described herein are also stable (e.g., chemically and physically stable) for 1, 2, 3, 4, 5, 6, 8, 10, 12, 14, 16, 18. 20, 24 hours or more; 1, 2, 3, 4, 5, 6, 7 days or more; 1, 2, 3, 4 weeks or more; 1, 2, 3, 4, 5, 6, 8, 10, 12 months or more; 1, 2, 3, 4, 5 years or more. Some buffered solutions or mixtures of neuroactive steroids are stable (eg, pH stable, chemically stable) at temperatures from about 3 to about 125°C. In some embodiments, buffered solutions or mixtures of neuroactive steroids are stable at a temperature of from about 3 to about 6°C. In some embodiments, buffered solutions or mixtures of neuroactive steroids are stable at a temperature of approximately 4°C. In some embodiments, the buffered solutions or mixtures of neuroactive steroids are stable at a temperature of from about 20 to about 40°C. In some embodiments, buffered solutions or mixtures of neuroactive steroids are stable at room (eg, ambient) temperature. In some embodiments, buffered solutions or mixtures of neuroactive steroids are stable at a temperature of approximately 25°C. In some embodiments, buffered solutions or mixtures of neuroactive steroids are stable at a temperature of approximately 37°C. In some embodiments, the buffered solutions or mixtures of neuroactive steroids are stable at a temperature of from about 115 to about 125°C, for example, for several minutes (e.g., 10, 20, 30, 40, 50, 60, 70, 80, 90 minutes or more, for several hours (e.g., 1, 2, 3 hours or more). In some embodiments, the buffered solutions or mixtures of neuroactive steroids are stable at the temperature of autoclaving. In some embodiments, the buffered solutions or mixtures of neuroactive steroids are stable at the temperature approximately 121°C.

In some embodiments, the buffered solutions or mixtures of neuroactive steroids described herein are stable at temperatures ranging from about 20 to 30°C for at least 1, 2, 3, 4, 5, 6, 7, 8 weeks ; 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 months or more; 1, 2, 3, 4, 5 years or more).

In some embodiments, the buffered solutions or mixtures of neuroactive steroids described herein are stable at temperatures ranging from about 2 to 8°C for at least 1, 2, 3, 4, 5, 6, 7, 8 weeks ; 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 months or more; 1, 2, 3, 4, 5 years or more).

In some embodiments, the buffered solutions or mixtures of neuroactive steroids described herein are prepared for administration by injection to a subject. As such, they will be prepared by processes designed to ensure they are sterile and free from pyrogens, particulate matter and other contaminants and, where appropriate, contain microbial growth inhibitors. Thus, buffered solutions or mixtures of neuroactive steroids will be substantially free of visible particulate matter. In some embodiments, the buffered solutions or mixtures of neuroactive steroids described herein may be filtered. In some embodiments, the buffered solutions or mixtures of neuroactive steroids described herein can be sterilized (e.g., filter sterilized (e.g., filtered through 0.45 and 0.22 μm filters), heat (e.g., steam sterilized at 121°C or by irradiation, such as gamma irradiation).In some embodiments, the sterilized buffered solutions or mixtures of neuroactive steroids do not contain elevated concentrations of impurities (e.g., an oxidized neuroactive steroid or a racemized or epimerized neuroactive steroid). For example, sterilized buffered solutions or mixtures of neuroactive steroids do not contain more than 0.001, 0.002, 0.005, 0.01, 0.02, 0.05, 0.1, 0.2, 0.5, 1% w/w impurities In some embodiments, sterilized buffered solutions or mixtures of neuroactive steroids have a pH of from about 3 to about 8 (eg, from about 5 to about 7, from about 5.5 to about 6.5).

In some embodiments, buffered solutions or mixtures of neuroactive steroids are safe, well tolerated, or non-irritating when administered to humans.

In some embodiments, the buffered neuroactive steroid described herein is formulated as an emulsion suitable for parenteral administration. Such emulsions will contain the neuroactive steroid described herein in a suitable oil or mixture of oils, suitable emulsifying ingredients, a suitable buffer and other ingredients necessary to modify tonicity and to provide chemical and physical stability of the composition. As such, they will be prepared by processes designed to ensure they are sterile and free from pyrogens, particulate matter and other contaminants and, where appropriate, contain microbial growth inhibitors. Thus, buffered solutions of neuroactive steroids will be substantially free of visible particulate matter. In some embodiments, the buffered solutions of neuroactive steroids described herein may be filtered. In some embodiments, the buffered solutions of neuroactive steroids described herein can be sterilized (e.g., sterilized by filtration (e.g., filtered through 0.45 and 0.22 μm filters), heat (e.g., steam sterilized at 121°C, or irradiated , such as gamma irradiation). In some embodiments, the sterilized buffered neuroactive steroid emulsions maintain the required globule or droplet size to allow safe and effective administration of the buffered neuroactive steroid emulsion. In some embodiments, the sterilized buffered neuroactive steroid emulsions do not contain elevated concentrations of impurities ( for example, an oxidized neuroactive steroid or a racemized or epimerized neuroactive steroid).For example, a sterilized buffered emulsion of a neuroactive steroid does not contain more than 0.001, 0.002, 0.005, 0.01, 0.02, 0.05, 0.1, 0.2, 0.5, 1% w/w impurities. In some embodiments, the sterilized buffered neuroactive steroid emulsion has a pH of about 3 to about 8 (eg, about 5 to about 7, about 5.5 to about 6.5).

In some embodiments, the buffered neuroactive steroid is provided as an oily solution suitable for injection. Such oil solutions will contain the neuroactive steroid in a suitable oil or mixture of oils and other ingredients as necessary to provide chemical and physical stability of the composition. In some embodiments, the selection of oils and excipients of the formulation provides the desired release and sustained activity of the neuroactive steroid. As such, they will be prepared by processes designed to ensure they are sterile and free from pyrogens, particulate matter and other contaminants and, where appropriate, contain microbial growth inhibitors. Thus, the buffered oil solution of the neuroactive steroid will be substantially free of visible particulate matter. In some embodiments, the buffered oil solutions of neuroactive steroids described herein may be filtered. In some embodiments, the buffered oil solution of a neuroactive steroid described herein can be sterilized (e.g., filter sterilized (e.g., filtered through 0.45 and 0.22 μm filters), heat (dry heat sterilization >150°C). In some embodiments, the sterilized buffered oil solution of the neuroactive steroid does not contain elevated concentrations of impurities (e.g., oxidized neuroactive steroid or racemized or epimerized neuroactive steroid).For example, the sterilized buffered oil solution of the neuroactive steroid does not contain more than 0.001, 0.002, 0.005, 0.01 , 0.02, 0.05, 0.1, 0.2, 0.5, 1% w/w impurities.

In some embodiments, the buffered solution or emulsion of the neuroactive steroid is lyophilized. Such lyophilized solution or emulsion may contain excipients similar to those used for the neuroactive steroid solution described herein. In some embodiments, the lyophilized buffered neuroactive steroid solution or emulsion may contain additional components known to those skilled in the art to enhance the lyophilization process, such as, but not limited to, sugars, modified carbohydrate compounds, and solvents such as tert-butyl alcohol. As such, they will be prepared by processes designed to ensure they are sterile and free from pyrogens, particulate matter and other contaminants and, where appropriate, contain microbial growth inhibitors. Thus, the lyophilized buffered solution or emulsion of the neuroactive steroid will be substantially free of visible particulate matter upon reconstitution. In some embodiments, the lyophilized buffered neuroactive steroid solution or emulsion described herein may be filtered before and after reconstitution. In some embodiments, the lyophilized buffered neuroactive steroid solution or emulsion described herein may be sterilized (e.g., filter sterilized (e.g., filtered through 0.45 and 0.22 μm filters) or irradiation (e.g., gamma irradiation). In some embodiments, the lyophilized sterilized buffered solution or emulsion of the neuroactive steroid does not contain more than 0.001, 0.002, 0.005, 0.01, 0.02, 0.05, 0.1, 0.2, 0.5, 1 wt%. by weight of impurities (e.g., oxidized neuroactive steroid or racemized or epimerized neuroactive steroid).In some embodiments, the sterilized lyophilized buffered solution or emulsion of the neuroactive steroid has a pH of from about 3 to about 8 (e.g., from about 5 to about 7, from about 5 .5 to approximately 6.5) after dilution.

Mixture

The aqueous solutions described herein may be mixed with the diluent described herein to form a "mixture". Suitable diluents include sterile water for injection ("WFI"), saline and dextrose. In some embodiments, an aqueous solution described herein is mixed with a diluent described herein in an aqueous solution:diluent ratio of 1:2. In some embodiments, an aqueous solution described herein is mixed with a diluent described herein in an aqueous solution:diluent ratio of 1:9.

In some embodiments, the mixture contains from about 1 to about 3 mg/ml of a neuroactive steroid. In some embodiments, the mixture contains from about 1.2 to about 2.5 mg/ml of neuroactive steroid. In some embodiments, the mixture contains from about 1.4 to about 2.0 mg/ml of neuroactive steroid. In some embodiments, the mixture contains from about 1.6 to about 1.7 mg/ml of neuroactive steroid. In some embodiments, the mixture contains approximately 1.67 mg/ml of a neuroactive steroid.

In some embodiments, the mixture contains from about 0.1 to about 1 mg/ml of a neuroactive steroid. In some embodiments, the mixture contains from about 0.25 to about 0.75 mg/ml of a neuroactive steroid. In some embodiments, the mixture contains approximately 0.5 mg/ml of a neuroactive steroid.

In some embodiments, the mixture contains from about 1% to about 20% wt./mass. cyclodextrin, for example sulfoalkyl ether-β-cyclodextrin. In some embodiments, the mixture contains from about 2.5% to about 15% wt./mass. cyclodextrin, for example sulfoalkyl ether-β-cyclodextrin. In some embodiments, the mixture contains from about 5% to about 15% wt./mass. cyclodextrin, for example sulfoalkyl ether-β-cyclodextrin. In some embodiments, the mixture contains from about 5% to about 10% wt./mass. cyclodextrin, for example sulfoalkyl ether-β-cyclodextrin. In some embodiments, the mixture contains approximately 8.3% wt./mass. cyclodextrin, for example sulfoalkyl ether-β-cyclodextrin.

In some embodiments, the mixture contains from about 0.1% to about 10% wt./mass. cyclodextrin, for example sulfoalkyl ether-β-cyclodextrin. In some embodiments, the mixture contains from about 0.5% to about 7.5% wt./mass. cyclodextrin, for example sulfoalkyl ether-β-cyclodextrin. In some embodiments, the mixture contains from about 0.5% to about 5% wt./mass. cyclodextrin, for example sulfoalkyl ether-β-cyclodextrin. In some embodiments, the mixture contains from about 1% to about 5% wt./mass. cyclodextrin, for example sulfoalkyl ether-β-cyclodextrin. In some embodiments, the mixture contains approximately 2.5% w/w. cyclodextrin, for example sulfoalkyl ether-β-cyclodextrin.

In some embodiments, the mixture contains from about 1 to about 3 mg/ml of neuroactive steroid and from about 1% to about 20% wt./wt. cyclodextrin, for example sulfoalkyl ether-β-cyclodextrin. In some embodiments, the mixture contains from about 1.2 to about 2.5 mg/ml of neuroactive steroid and from about 2.5% to about 15% wt./wt. cyclodextrin, for example sulfoalkyl ether-β-cyclodextrin. In some embodiments, the mixture contains from about 1.4 to about 2.0 mg/ml of neuroactive steroid and from about 5% to about 15% wt./mass. cyclodextrin, for example sulfoalkyl ether-β-cyclodextrin. In some embodiments, the mixture contains from about 1.6 to about 1.7 mg/ml of neuroactive steroid and from about 5% to about 10% wt./wt. cyclodextrin, for example sulfoalkyl ether-β-cyclodextrin. In some embodiments, the mixture contains approximately 1.67 mg/ml of neuroactive steroid and approximately 8.3% w/w. cyclodextrin, for example sulfoalkyl ether-β-cyclodextrin.

In some embodiments, the mixture contains from about 0.1 to about 1 mg/ml of neuroactive steroid and from about 0.1% to about 10% wt./wt. cyclodextrin, for example sulfoalkyl ether-β-cyclodextrin. In some embodiments, the mixture contains from about 0.25 to about 0.75 mg/ml of neuroactive steroid and contains from about 0.5% to about 5% wt./wt. cyclodextrin, for example sulfoalkyl ether-β-cyclodextrin. In some embodiments, the mixture contains approximately 0.5 mg/mL of neuroactive steroid and approximately 2.5% w/w. cyclodextrin, for example sulfoalkyl ether-β-cyclodextrin.

In some embodiments, the mixture contains a buffer as described herein, eg, citrate buffer, phosphate buffer. In some embodiments, the buffer is present in an amount of about 1 to about 500 mM (e.g., about 1 to about 250 mM, about 1 to about 200 mM, about 1 to about 150 mM, about 1 to about 100 mM, from about 1 to about 50 mM). In some embodiments, the buffer is at or near physiological pH. Preferably, the pH of the mixture is from about 3 to about 8 (for example, from about 5 to about 7, from about 5.5 to about 6.5, from about 5.9 to about 6.1) or any particular value within the specified range. In some embodiments, the pH of the mixture is from about 5 to about 6.5, or any specific value within that range (e.g., 5.5, 5.6, 5.7, 5.8, 5.9, 6, 6.1 , 6.2, 6.3, 6.4). In some embodiments, the pH of the mixture is about 6. In some embodiments, the buffer is a citrate buffer and the pH is from about 3 to about 7.4. In some embodiments, the buffer is a citrate buffer and the pH is from about 5.5 to about 6.2. In some embodiments, the buffer is a phosphate buffer and the pH is from about 6.2 to 8.2, preferably about 7.4.

In some embodiments, the mixture contains one part buffered neuroactive steroid solution (buffered neuroactive steroid solution as described herein) to two parts diluent (eg, WFI).

In some embodiments, the mixture contains one part buffered neuroactive steroid solution (buffered neuroactive steroid solution as described herein) to nine parts diluent (eg, saline, WFI).

In some embodiments, the mixture is isotonic. In some embodiments, the mixture is hypotonic. In some embodiments, the tonicity of the mixture is adjusted, for example, by tonicity enhancers, to produce solutions that become approximately 300 mOsm/L or less.

Buffers

The aqueous solution or mixture of a neuroactive steroid described herein contains a buffer (e.g., a buffer at a pH of from about 3 to about 8 (e.g., from about 5 to about 7, from about 5.5 to about 6.5, from about 5 .9 to about 6.1) As used herein, the term “buffer,” “buffer system,” or “buffering component” refers to a compound that typically, in combination with at least one other compound, provides a chemical system in solution that has buffering capacity, that is, the ability to neutralize, within certain limits, the pH-lowering or pH-raising effects of strong acids or bases (alkalis), respectively, with relatively little or no change in the original pH value (for example, the pH before exposure to, for example, a strong acid or base).For example, the buffer described herein maintains or adjusts the pH of a solution within a specific pH range. For example, "buffering capacity" may refer to the millimoles (mM) of strong acid or base (or hydrogen or hydroxide ions, respectively) required to change the pH by one unit when added to one liter (standard unit) of a buffer solution. From this definition it can be seen that the smaller the change in pH in a solution caused by the addition of a certain amount of acid or alkali, the greater the buffering capacity of the solution. See, for example, Remington: The Science and Practice of Pharmacy, Mack Publishing Co., Easton, Pennsylvania (19th edition, 1995), chapter 17, pages 225-227. The buffering capacity will depend on the type and concentration of the buffering components.

In some embodiments, the buffering components are present in solutions at a concentration of 1 mM, 2 mM, 5 mM, 10 mM, 20 mM, 50 mM, 75 mM, 100 mM, 150 mM, 200 mM, 250 mM, or more.

Preferred buffers include 4-2-hydroxyethyl-1-piperazineethanesulfonic acid (HEPES), 2-{[tris(hydroxymethyl)methyl]amino}ethanesulfonic acid (TES), 3-(N-morpholino)propanesulfonic acid (MOPS), piperazine- N,N'-bis(2-ethanesulfonic acid) (PIPES), dimethylarsinic acid (cacodylate), citrate (eg, sodium citrate, potassium citrate, ammonium citrate saline), 2-(N-morpholino)ethanesulfonic acid (MES) , phosphate (i.e. PBS, D-PBS), succinate (i.e. 2(R)-2-(methylamino)succinic acid), acetate, dimethyl glutarate, maleate, imidazole, N-(2-acetamido)-2- aminoethanesulfonic acid (ACES), N,N-bis(2-hydroxyethyl)-2-aminoethanesulfonic acid (BES), bicine, Bis-Tris, borate, N-cyclohexyl-3-aminopropanesulfonic acid (CAPS), glycine, 3-[ 4-2-hydroxyethyl)-1-piperazinyl]propanesulfonic acid (HEPPS or EPPS), N-[tris(hydroxymethyl)methyl]-3-aminopropanesulfonic acid, [(2-hydroxy-1,1-bis(hydroxymethyl)ethyl) amino]-1-propanesulfonic acid (TAPS), tricine, Tris, Tris base, Tris buffer, Tris-glycine, Tris-HCl, collidine, veronal acetate, N-(2-acetamido)iminodiacetic acid; N-(carbamoylmethyl)iminodiacetic acid (ADA), β-hydroxy-4-morpholinepropanesulfonic acid, 3-morpholino-2-hydroxypropanesulfonic acid (MOPSO), cholamine chloride, 3-(N,N-bis[2-hydroxyethyl]amino)- 2-hydroxypropanesulfonic acid (DIPSO), acetamidoglycine, 3-{[1,3-dihydroxy-2-(hydroxymethyl)-2-propanyl]amino}-2-hydroxy-1-propanesulfonic acid (TAPSO), piperazine-N,N -bis(2-hydroxypropanesulfonic acid) (POPSO), N-(2-hydroxyethyl)piperazine-N'-(2-hydroxypropanesulfonic acid) (HEPPSO), N-cyclohexyl-2-aminoethanesulfonic acid (CHES), 2-amino- methyl 1,3-propanediol (AMPd) and glycinamide.

In some embodiments, the buffer contains a monoprotic acid. In some embodiments, the buffer contains a polybasic acid (eg, citrate or phosphate). In some embodiments, the buffer is a solution of one or more substances (eg, a salt of a weak acid and a weak base, a mixture of a weak acid and a salt of a weak acid and a strong base). In some embodiments, the buffer contains piperazine (eg, PIPES, HEPES, POPSO, EPPS).

In some embodiments, the buffer contains a non-metal complexing compound (eg, MES, MOPS, PIPES).

In some embodiments, the buffer contains a metal complexing compound (ie, a metal chelating agent). In some embodiments, the metal chelating agent is citrate.

In some embodiments, the buffer is a citrate buffer. In some embodiments, the buffer is a phosphate buffer. In some embodiments, the buffer is a histidine buffer.

In some embodiments, the buffer is present at a concentration of about 0.01, 0.05, 0.1, 0.5, 1, 5, 10, 20, 50, 100, 200, 250, 500 mM, or more. In some embodiments, the buffer is present at a concentration of from about 1 to about 500 mM, from about 1 to about 300 mM, from about 1 to about 200 mM, from about 1 to about 100 mM, from about 1 to about 50 mM, from about 10 to about 500 mM, about 10 to about 300 mM, about 10 to about 200 mM, about 10 to about 100 mM, about 10 to about 50 mM.

In some embodiments, the buffer is present at a concentration of from about 0.01 to about 10 mM, from about 0.05 to about 5 mM, from about 0.05 to about 5 mM, from about 0.1 to about 5 mM, from about 0.1 to approximately 3.5 mM.

In some embodiments, the pH of the aqueous solution is at or near physiological pH. Preferably, the pH of the aqueous solution is from about 3 to about 8 (for example, from about 5 to about 7, from about 5.5 to about 6.5, from about 5.9 to about 6.1) or any specific value within the specified range . In some embodiments, the pH of the aqueous solution is from about 5 to about 6.5, or any specific value within that range (e.g., 5.5, 5.6, 5.7, 5.8, 5.9, 6, 6, 1, 6.2, 6.3, 6.4). In some embodiments, the pH of the aqueous solution is approximately 6. One skilled in the art will appreciate that the pH can be adjusted to a more optimal pH depending on the stability of the neuroactive steroids and sulfoalkyl ether-β-cyclodextrin included in the solution. The pH can be adjusted, for example, using hydrochloric acid, phosphoric acid or organic acids such as citric acid, lactic acid, malic acid, tartaric acid, acetic acid, gluconic acid, succinic acid and combinations thereof. In some embodiments, the pH is adjusted using a base (eg, 1N sodium hydroxide) or acid (eg, 1N hydrochloric acid).

In some embodiments, the buffer is a citrate buffer and the pH is from about 3 to about 8. In some embodiments, the buffer is a citrate buffer and the pH is from about 3 to about 7.4. In some embodiments, the buffer is a citrate buffer and the pH is from about 5.5 to about 6.2.

In some embodiments, the buffer is a phosphate buffer and the pH is from about 3 to about 9. In some embodiments, the buffer is a phosphate buffer and the pH is from about 6.2 to about 8.2. In some embodiments, the buffer is a phosphate buffer and the pH is approximately 7.4.

Neuroactive steroids

The aqueous solutions or mixtures described herein contain the neuroactive steroid described herein. Neuroactive steroids (or neurosteroids) are natural, synthetic, or semi-synthetic steroids that rapidly alter neuronal excitability through interaction with neurotransmitter-gating ion channels. Neuroactive steroids bind to membrane-bound receptors such as inhibitory and/or excitatory neurotransmitters, including GABA _A , NMDA and sigma receptors.

Steroids, which can be classified into functional groups according to chemical structure and physiological activity, include estrogen hormones, progestational hormones, and androgenic hormones. Of particular interest are progestational hormones, referred to herein as “progestins” or “progestogens,” as well as their derivatives and bioactive metabolites. Members of this broad family include the steroid hormones described in Remington's Pharmaceutical Sciences, Gennaro et al., Mack Publishing Co. (18th ed., 1990), 990-993. As with all other classes of steroids, stereoisomerism is of fundamental importance for sex hormones. Various progestins (eg, progesterone) and their derivatives may be used as used herein, including both synthetic and natural products, as well as progestin metabolites such as progesterone.

As used herein, the term "progesterone" refers to a member of the progestin family and includes a 21-carbon steroid hormone. Progesterone is also known as D4-pregnene-3,20-dione; Δ4-pregnene-3,20-dione; or pregn-4-ene-3,20-dione. As used herein, the term “synthetic progestin” is a molecule whose structure is related to that of progesterone, is synthetically produced, and retains the biological activity of progesterone.

Typical synthetic progestins include, but are not limited to, substitutions at position 17 of progesterone to introduce a hydroxyl, acetyl, hydroxyacetyl, aliphatic, nitro or heterocyclic group, modifications to produce 17α-OH esters (eg, 17α-hydroxyprogesterone caproate), and modifications that introduce 6-methyl, 6-ene and 6-chloro substituents into progesterone (eg, medroxyprogesterone acetate, megestrol acetate and chloromadinone acetate), and which retain the biological activity of progesterone. Such progestin derivatives include 5-dehydroprogesterone, 6-dehydro-retroprogesterone (dydrogesterone), allopregnanolone (allopregnan-3α or 3β-ol-20-one), ethynodiol diacetate, hydroxyprogesterone caproate (pregn-4-ene-3,20-dione, 17-(1-oxohexy)oxy); levonorgestrel, norethindrone, norethindrone acetate (19-norpregn-4-en-20-in-3-one, 17-(acetyloxy)-, (17α)-); norethinodrel, norgestrel, pregnenolone, ganaxolone (also called CCD-1042 or INN) and megestrol acetate. In some embodiments, the neuroactive steroid is ganaxolone.

Suitable progestins may also include allopregnon-3α or 3β, 20α or 20β-diol (see Merck catalog 258-261); allopregnan-3β,21-diol-11,20-dione; allopregnan-3β,17α-diol-20-one; 3,20-allopregnanedione, allopregnan, 3β,11β,17α,20β,21-pentol; allopregnan-3β,17α,20β,21-tetrol; allopregnan-3α or 3β,11β,17α,21-tetrol-20-one, allopregnan-3β,17α or 20β-triol; allopregnan-3β,17α,21-triol-11,20-dione; allopregnan-3β,11β,21-triol-20-one; allopregnan-3β,17α,21-triol-20-one; allopregnan-3α or 3β-ol-20-one; pregnanediol; 3,20-pregnandione; pregnan-3α-ol-20-one; 4-pregnene-20,21-diol-3,11-dione; 4-pregnen-11β,17α,20β,21-tetrol-3-one; 4-pregnene-17α,20β,21-triol-3,11-dione; 4-pregnen-17α,20β,21-triol-3-one and pregnenolone methyl ester. Other progestin derivatives include esters with non-toxic organic acids such as acetic acid, benzoic acid, maleic acid, malic acid, caproic acid and citric acid and inorganic salts such as hydrochloride, sulfate, nitrate, bicarbonate and carbonate salts. Other suitable progestins include alfaxalone (also referred to as INN, alfaxolone and alfaxolone), alfadolone (also referred to as alfadolone), hydroxydione and minaxolone. In some embodiments, the neuroactive steroid is alfaxolone.

Additional suitable neuroactive steroids are disclosed in WIPO Publication Nos. WO2013/188792, WO2013/056181, WO2015/010054, WO2014/169832, WO2014/169836, WO2014/169833, WO2014/169831, WO20 15/027227, WO 2014/100228 and US Patent No. 5,232,917, US 8,575,375 and US 8,759,330, which are incorporated herein by reference to the neuroactive steroids described therein.

In specific embodiments, the steroids are one or more of a series of sedative-hypnotic 3 alpha-hydroxy reduced ring A pregnane steroids, which include the major metabolites of progesterone and deoxycorticosterone, 3alpha-hydroxy-5alpha-pregnan-20-one (allopregnanolone) and 3alpha ,21-dihydroxy-5alpha-pregnan-20-one (allotetrahydroDOC), respectively. These 3 alpha-hydroxysteroids do not interact with classical intracellular steroid receptors, but bind stereoselectively and with high affinity to receptors for the major inhibitory neurotransmitter in the brain, gamma-aminobutyric acid (GABA).

In some embodiments, the neuroactive steroids are progesterone, pregnanolone, allopregnanolone, alfadalone, ganxolone, alfaxolone, or other progesterone analogues. In a specific embodiment, the neuroactive steroid is allopregnanolone or a derivative thereof. In some embodiments, the neuroactive steroid is allopregnanolone. Examples of derivatives include, but are not limited to, (20R)-17beta-(1-hydroxy-2,3-butadienyl)-5alpha-androstane-3alpha-ol (HBAO). Additional derivatives are described in WO 2012/127176.

In some embodiments, the neuroactive steroid is allopregnanolone. In some embodiments, the neuroactive steroid is ganaxolone. In some embodiments, the neuroactive steroid is alfaxolone.

The lipophilic nature of a neuroactive steroid (eg, pregnanolone, allopregnanolone, alfadalone, ganxolone, or alfaxolone) can be modified to produce a formulation for in vivo administration. As discussed above, a neuroactive steroid (eg, pregnanolone, allopregnanolone, alfadalone, ganxolone, or alfaxolone) can be combined with a carrier such as cyclodextrin to improve solubility. Alternatively or additionally, a neuroactive steroid (eg, pregnanolone, allopregnanolone, alfadalone, gangsolone, or alfaxolone) may be modified to improve solubility. For example, polar groups can be introduced at the 16α position to increase the water solubility, brain availability, and potency of neuroactive steroids, as described in Kasal et al., J. Med. Chem., 52(7), 2119-215 (2009).

Cyclodextrins

The aqueous solution or mixture of neuroactive steroid described herein contains cyclodextrin. The solubility of neuroactive steroids can be improved using cyclodextrins. Steroid-cyclodextrin complexes are known in the art. See, for example, US Patent No. 7,569,557 to Backensfeld et al. and the publication of US patent application No. US 2006/0058262 Zoppetti et al.

Cyclodextrins are cyclic oligosaccharides consisting of or containing six (α-cyclodextrin), seven (β-cyclodextrin), eight (γ-cyclodextrin) or more α(1,4)-linked glucose residues. The hydroxyl groups of the cyclodextrins face the outside of the ring, while the glucosidic oxygen and two rings of non-exchangeable hydrogen atoms face the inside of the cavity.

Neuroactive steroid-cyclodextrin complexes are preferably prepared from a cyclodextrin selected from the group consisting of β-cyclodextrin and its derivatives. The cyclodextrin can be chemically modified such that some or all of the primary or secondary hydroxyl groups of the macrocycle, or both, are functionalized with a side group. Suitable side groups include, but are not limited to, sulfinyl, sulfonyl, phosphate, acyl and C ₁ -C ₁₂ alkyl groups, optionally substituted with one or more (for example, 1, 2, 3 or 4) hydroxy, carboxy, carbonyl, acyl, oxy, oxo; or combinations thereof. Methods for modifying these alcohol moieties are known in the art, and many cyclodextrin derivatives are commercially available, including sulfobutyl ether β-cyclodextrins available under the trade name CAPTISOL® from Ligand Pharmaceuticals (La Jolla, Calif.).

Preferred cyclodextrins include, but are not limited to, alkylcyclodextrins, hydroxyalkylcyclodextrins such as hydroxypropyl β-cyclodextrin, carboxyalkylcyclodextrins and sulfoalkyl ethers of cyclodextrins such as β-cyclodextrin sulfobutyl ether.

In specific embodiments, the cyclodextrin is a beta-cyclodextrin having a plurality of charges (eg, negative or positive) on the surface. In more specific embodiments, the cyclodextrin is a β-cyclodextrin consisting of or containing multiple functional groups that are negatively charged at physiological pH. Examples of such functional groups include, but are not limited to, carboxylic acid (carboxylate) groups, sulfonate (RSO ^3- ) groups, phosphonate groups, phosphinate groups, and amino acids, which are negatively charged at physiological pH. The charged functional groups can be linked directly to the cyclodextrins or can be linked via a spacer such as an alkylene chain. The number of carbon atoms in the alkylene chain may vary, but is typically from about 1 to 10 carbon atoms, preferably 1 to 6 carbon atoms, more preferably 1 to 4 carbon atoms. Highly sulfonated cyclodextrins are described in US Patent No. 6,316,613.

In one embodiment, the cyclodextrins are β-cyclodextrin functionalized with multiple sulfobutyl ether groups. Such cyclodextrins are sold under the trade name CAPTISOL®.

CAPTISOL® is a polyanionic derivative of beta-cyclodextrin with a sodium sulfonate salt separated from the lipophilic cavity by a butyl ether spacer group or sulfobutyl ether (SBE). CAPTISOL® is not a single chemical compound, but is composed of a variety of polymer structures of varying degrees of substitution and positional/spatial isomers, conditioned and controlled to obtain a uniform composition using a proprietary manufacturing process that is repeatedly put into practice and improved to control impurities.

CAPTISOL® contains six to seven sulfobutyl ether groups per cyclodextrin molecule. Due to the very low pKa of sulfonic acid groups, CAPTISOL® carries many negative charges at physiologically compatible pH values. The four-carbon butyl chain, combined with the repulsion of negative charges on the end group, allows the cyclodextrin cavity to "expand". This often results in stronger binding to drug candidates than can be achieved using other modified cyclodextrins. This also allows for ionic charge interactions between the cyclodextrin and the positively charged drug molecule. In addition, these derivatives provide the molecule with significant solubility and safety for parenteral administration. Compared to beta-cyclodextrin, CAPTISOL® provides better interaction characteristics and superior water solubility of more than 100 g/100 ml, a 50-fold improvement.

Preferably, the cyclodextrin is present in an amount of from about 0.1% to about 40% w/w. of the total solution (e.g., a buffered solution of a neuroactive steroid), preferably from about 5% to about 40% w/w, more preferably from about 10% to about 40% w/w, most preferably from about 10% to approximately 35% w/w In some embodiments, the concentration of cyclodextrins is from about 15% to about 35% w/w, preferably from about 20% to about 35% w/w, more preferably from about 20% to about 30% w/w . In some embodiments, the concentration of cyclodextrins is approximately 25% w/w.

In one embodiment, the formulation contains from about 1 to about 2, preferably about 1.5 mg of a neuroactive steroid (eg, pregnanolone, allopregnanolone, alfadalone, ganaxolone, alfaxolone) per ml of cyclodextrin, eg, CAPTISOL®. In some embodiments, the cyclodextrin, e.g., sulfoalkyl ether-β-cyclodextrin, is present in the aqueous solution described herein in an amount of 0.1, 0.2, 0.3, 0.5, 0.7, 1.2, 1.5, 1.8, 2, 2.5, 3, 4, 5, 6, 7, 8, 10, 11, 12 mg/ml or more.

In some embodiments, the cyclodextrin, for example, sulfoalkyl ether-β-cyclodextrin, is present in the aqueous solution described herein in an amount of 1, 2, 3, 5, 7, 10, 12, 20, 25, 30, 40 wt%. /mass. or more.

In some embodiments, the cyclodextrin, e.g., sulfoalkyl ether-β-cyclodextrin, is present in the aqueous solution described herein in an amount of at least 0.1, 0.2, 0.3, 0.5, 0.7, 1 ,2, 1.5, 2, 3, 4, 5, 6, 7, 8, 10 mg/l or more.

In some embodiments, the molar ratio of neuroactive steroid to cyclodextrin, such as sulfoalkyl ether-β-cyclodextrin, is about 0.1, 0.05, 0.03, 0.02, 0.01, 0.008, 0.005 or less.

Tonicity Enhancers

The aqueous solution or mixture of the neuroactive steroid described herein may further contain a tonicity enhancer. Tonicity is the equivalent effective osmotic pressure or relative concentration of solutions, which determines the direction and extent of diffusion. Tonicity can be adjusted if necessary, usually with tonicity-enhancing agents. Such agents may, for example, be of the ionic and/or non-ionic type. Examples of ionic tonicity enhancers are alkali or alkali metal halides, such as, for example, CaCl ₂ , KBr, KCl, LiCl, NaI, NaBr or NaCl, Na ₂ SO ₄ or boric acid. Nonionic agents that enhance tonicity are, for example, urea, glycerin, sorbitol, mannitol, propylene glycol or dextrose. The aqueous solutions described are typically adjusted to isotonicity with tonicity agents (eg, about 270 to about 300 mOsm/L, about 275 to about 295 mOsm/L). In some embodiments, the described aqueous solutions are adjusted with tonicity agents to an osmolarity in the range of about 150 to about 320 mOsm/L (eg, about 200 to about 300 mOsm/L). In some embodiments, the aqueous solutions are less than about 320 mOsm/L (eg, less than about 300, 290, 280, 270, 260, 250 mOsm/L).

In some embodiments, the described aqueous solutions are hypertonic. For example, aqueous solutions may be hypertonic (eg, from about 900 to about 1000 mOsm/L). In some embodiments, aqueous solutions are diluted with Water for Injection ("WFI", e.g., highly purified water without any added components; a sterile, nonpyrogenic preparation of distilled water for injection without solutes), for example, to produce an isotonic or hypotonic solution. In some embodiments, the mixture is diluted with a NaCl solution (eg, saline).

Preservatives

The aqueous solution or mixture of a neuroactive steroid described herein may contain preservatives. Typical preservatives include antimicrobials (eg, tissue plasminogen activator, sargramostim, interleukins, phenol, benzyl alcohol, metacresol, parabens (methyl, propyl, butyl), benzalkonium chloride, chlorobutanol, thimerosal, phenylmercuric salts (acetate, borate, nitrate)), benzalkonium chloride, benzethonium chloride, chlorobutanol, myristyl gamma-picolinium chloride, 2-phenoxyethanol, thiomersal, methylparaben, propylparaben, butylparaben, ethylenediamine, formaldehyde.

The aqueous solution or mixture of a neuroactive steroid described herein may contain antioxidants. Typical antioxidants include sodium bisulfite, sulfuric acid salts, ascorbic acid and its salts, acetylcysteine, monothioglycerol), EDTA, cryoprotectants and lyoprotectants (e.g., sugars (e.g., sucrose, trehalose), amino acids (e.g., glycine, lysine), polymers ( e.g. liquid polyethylene glycol or dextran), polyols (e.g. mannitol, sorbitol).

Sterilization

The aqueous solution or mixture of a neuroactive steroid described herein may require sterilization, for example, before administration. The compositions described herein provide stability (eg, chemical stability, physical stability) in the case of sterilization processes. In some embodiments, the buffered solution or mixture of the neuroactive steroid is sterile. In some embodiments, the aqueous solution or mixture of the neuroactive steroid is sterilized through an aseptic processing process (eg, aseptic filling, aseptic filtration). In some embodiments, the aqueous solution or mixture of the neuroactive steroid is sterilized by terminal sterilization (e.g., heat (e.g., dry heat or steam autoclave) or irradiation (e.g., gamma irradiation). The compositions described herein (e.g., compositions containing buffer as described herein) provide stability (eg, chemical stability, physical stability) in the event of terminal sterilization (eg, temperature cycles from about 120°C to about 124°C, eg, 121°C) or irradiation.

Mixing

The aqueous solution or mixture of the neuroactive steroid described herein may require mixing, for example, to obtain homogeneous solutions or mixtures. In some embodiments, making a buffered solution or emulsion of a neuroactive steroid requires vigorous, high intensity, high shear mixing (stirring). Stirring can be supplemented with heating or without it. In some embodiments, heating the mixture while mixing can help achieve mixing efficiency and reduce the time required for dissolution or emulsification. The amount of heat (mixture temperature) depends on the system being mixed; but may be limited by equipment operation and physicochemical stability of the mixture. In some embodiments, a temperature of approximately 40°C has been found to be suitable for facilitating product preparation.

Mixing can be provided by devices such as high shear paddle mixers, rotor-stator mixers, homogenizers, ultrasonic devices or microfluidizers. Vigorous, high-intensity or high-shear mixing is used to stir and mix two mutually insoluble liquids or to facilitate the dissolution of solid particles in a carrier to make them uniform or homogeneous throughout. High shear mixers are designed to induce fluid movement at a different velocity relative to the fluid in the adjacent area. Dissolution or emulsification can be achieved by converting one of the phases of the product into a state consisting of extremely fine particles distributed evenly throughout the volume of the other liquid.

Mixing with high shear paddles may provide sufficient mixing to dissolve or emulsify the neuroactive steroid solution in some embodiments. However, in some embodiments, the mixing time may be too long for practical production runs. Agitation provided by rotor-stator mixers, homogenizers, ultrasonic devices or microfluidizers can speed up and facilitate dissolution to achieve practical production cycle times. In some embodiments, heating the mixture while mixing can help achieve mixing efficiency and reduce the time required for dissolution or emulsification. The amount of heat (mixture temperature) depends on the system being mixed; but may be limited by equipment operation and physicochemical stability of the mixture. In some embodiments, a temperature of approximately 40°C facilitates product preparation.

High shear mixers, such as rotor-stator mixers, can provide sufficient mixing to dissolve or emulsify the neuroactive steroid solution in some embodiments. High shear rotors/stators use a spinning blade or high-speed rotor, usually driven by an electric motor. The rotor rotates at very high speed (eg 2000 to 18000 rpm) in the mixture inside a stationary ring (stator) to create flux and shear. Suction is created by the high-speed rotation of the rotor blades inside the stator, drawing the mixture into the center of the rotor/stator module. High-speed centrifugal force moves the mixture towards the periphery of the rotor towards the stator, where it is crushed due to the limited clearance between the rotor and stator. The mixture is forced to undergo intense hydraulic shear at high speed through perforations in the stator into the mixing vessel. The influence of horizontal (radial) displacement and suction of the mixture into the rotor/stator creates a circulation mode in the mixing vessel. Rotor design and stator design vary among equipment types and designs; and one skilled in the art may find that numerous combinations of rotor and stator structures can operate acceptably. The size of the rotor/stator module will be determined depending on the batch size and the desired processing time. The location of the rotor/stator module will vary depending on the equipment design, but some embodiments may use a rotor/stator module mounted on or near the bottom of the mixing vessel. A top-mounted rotor/stator module designed to be immersed in the mixture may be used. A rotor/stator module mounted on the outside of a mixing vessel into which the mixture is introduced and can be initiated to flow or recirculate through the rotor/stator head. The desired rotor speed within the stator is typically variable and can be tuned to provide the desired flow and high shear mixing over practical production runs. Those skilled in the art will appreciate that the rotor peripheral speed can be used to facilitate scaling of the rotor/stator module size as the lot size increases. In some embodiments, heating the mixture during mixing can help achieve high shear mixing efficiency and reduce the time required for dissolution or emulsification. The amount of heat (mixture temperature) depends on the system being mixed; but may be limited by equipment operation and physicochemical stability of the mixture. In some embodiments, a temperature of approximately 40°C has been found to be suitable for facilitating the preparation of a product (eg, an aqueous solution or mixture as described herein).

High shear mixing devices, such as homogenizers, can provide sufficient mixing to dissolve or emulsify the neuroactive steroid solution in some embodiments. Homogenizers provide high shear because they work to pump a mixture at high pressure (e.g., 1000-5000 psi) into a small chamber consisting of a valve seat, impact ring, and valve. The mixture flows under high pressure through the area between the valve and the valve seat at high speed with the help and under a rapid pressure difference. The rapid pressure difference breaks up the mixture by cavitation and the shock generated when the cavitation bubble is destroyed. The mixture then hits the impact ring, causing further disruption and shear in the mixture. The mixture is discharged into the main volume of the solution. Various valve modules, relative location of emulsifier to product batch, multiple valve modules, and equipment with a wide range of capacities can be used. In some embodiments, heating the mixture during mixing can help achieve high shear mixing efficiency and reduce the time required for dissolution or emulsification. The amount of heat or temperature control of the mixing process (mixture temperature) depends on the system being mixed; but may be limited by equipment operation and physicochemical stability of the mixture. In some embodiments, a temperature of approximately 40°C has been found to be suitable for facilitating the preparation of a product (eg, an aqueous solution or mixture as described herein).

High shear mixing devices, such as microfluidizers, can provide sufficient mixing to dissolve or emulsify the neuroactive steroid solution in some embodiments. High shear mixing by microfluidizers is caused by pumping the mixture at extremely high speeds at high pressures (e.g., 2,000 to 40,000 psi) through small channels into an interaction chamber. In the interaction chamber, the mixture is subjected to high shear, turbulence, shock and cavitation. All of these forces can make it easier to achieve high shear mixing efficiency and reduce the time required for dissolution or emulsification. Various interaction chamber modules, relative positioning of the microfluidizer to the product batch, and equipment with a wide range of capacities can be used. The amount of heat or temperature control of the mixing process (mixture temperature) depends on the system being mixed; but may be limited by equipment operation and physicochemical stability of the mixture. In some embodiments, a temperature of approximately 40°C has been found to be suitable for facilitating product preparation.

High shear mixing devices that utilize ultrasonic energy can provide sufficient agitation to dissolve or emulsify the neuroactive steroid solution in some embodiments. The high shear displacement due to the results of ultrasonic energy is due to cavitation and the rapid destruction of small bubbles formed by cavitation. These forces can make it easier to achieve high shear mixing efficiency and reduce the time required for dissolution or emulsification. Various ultrasonic modules can be used, the relative location of the ultrasonic processing module to the product batch, and equipment with a wide range of powers. The amount of heat or temperature control of the mixing process (mixture temperature) depends on the system being mixed; but may be limited by equipment operation and physicochemical stability of the mixture. In some embodiments, a temperature of approximately 40°C has been found to be suitable for facilitating product preparation.

Containers

Also described herein are containers that contain the aqueous solution or mixture described herein. Examples of containers include bags (eg, plastic or polymer bags such as PVC), vials (eg, glass vial), bottles, or syringes. In one embodiment, the container is configured to deliver the solution or mixture parenterally (eg, intramuscularly or intravenously).

In some embodiments, the injectable product is packaged in an appropriately sized sealed glass container. In some embodiments, the product is intended to be reconstituted prior to infusion and is packaged in a pharmaceutical vial or bottle (eg, a suitable size, suitable glass or plastic vial or bottle). In some embodiments, the product may be prepared ready for injection and may be packaged in a prefilled syringe or other injection device (e.g., a suitable size, suitable glass or plastic packaging) or a large volume container (e.g., a suitable size, suitable glass or plastic container) intended for infusion. In some embodiments, the product is provided in a container that does not leach (e.g., introduce (or allow growth of) solids or contaminants into the solution.

Neurodegenerative diseases and disorders

The solutions or mixtures described herein may be used in a method described herein, for example, in the treatment of a disorder described herein, such as a neurodegenerative disease.

The term "neurodegenerative disease" includes diseases and disorders that are associated with the gradual loss of neuronal structure or function, or the death of neurons. Neurodegenerative diseases and disorders include, but are not limited to, Alzheimer's disease (including associated symptoms of mild, moderate or severe cognitive impairment); amyotrophic lateral sclerosis (ALS); anoxic and ischemic damage; ataxia and seizures (including for the treatment and prevention and prevention of epileptiform seizures caused by schizoaffective disorder or drugs used to treat schizophrenia); moderate forgetfulness; cerebral edema; cerebellar ataxia, including McLeod neuroacanthocytosis syndrome (MLS); closed skull injury; to whom; concussion injuries (eg, spinal cord injury and head injury); dementia, including multi-infarct dementia and senile dementia; disorders of consciousness; Down syndrome; drug-induced or treatment-induced Parkinsonism (eg, neuroleptic-induced acute akathisia, acute dystonia, Parkinsonism or tardive dyskinesia, neuroleptic malignant syndrome, or treatment-induced postural tremor); epilepsy; fragile X syndrome; Gilles de la Tourette syndrome; head injury; hearing impairment and loss; Huntington's disease; Lennox syndrome; levodopa-induced dyskinesia; mental retardation; movement disorders, including akinesia and akinetic (rigid) syndromes (including calcification of the basal ganglia of the brain, corticobasal degeneration, multiple system atrophy, ALS-Parkinsonism-dementia complex, Parkinson's disease, postencephalitic parkinsonism and progressive supranuclear palsy); muscle spasms and disorders associated with muscle spasticity or weakness, including chorea (such as benign hereditary chorea, drug-induced chorea, hemiballismus, Huntington's disease, neuroacanthocytosis, Sydenham's chorea and symptomatic chorea), dyskinesia (including tics, such as complex tics, simple tics as well as symptomatic tics), myoclonus (including generalized myoclonus and focal cycloclonus), tremor (eg, residual tremor, postural tremor, and intention tremor), and dystonia (including axial dystonia, dystonic graphospasm, hemiplegic dystonia, paroxysmal dystonia, and focal dystonia such as blepharospasm, oromandibular dystonia and spasmodic dysphonia and torticollis); neuronal damage, including vision damage, retinopathy, or macular degeneration of the eye; neurotoxic injury that follows cerebral stroke, thromboembolic stroke, hemorrhagic stroke, cerebral ischemia, cerebral vasospasm, hypoglycemia, amnesia, hypoxia, anoxia, perinatal asphyxia and cardiac arrest; Parkinson's disease; seizure; status epilepticus; stroke; noise in ears; tubular sclerosis, neurodegeneration induced by viral infection (for example, caused by acquired immunodeficiency syndrome (AIDS) and encephalopathies). Neurodegenerative diseases also include, but are not limited to, neurotoxic damage that follows cerebral stroke, thromboembolic stroke, hemorrhagic stroke, cerebral ischemia, cerebral vasospasm, hypoglycemia, amnesia, hypoxia, anoxia, perinatal asphyxia and cardiac arrest. Methods for treating or preventing a neurodegenerative disease also include treating or preventing the loss of neuronal function characteristic of the neurodegenerative disorder.

Mood disorders

The solutions or mixtures described herein can be used in a method described herein, for example, in the treatment of a disorder described herein, such as a mood disorder.

Clinical depression is also known as major depression, major depressive disorder (MDD), major depression, unipolar depression, unipolar disorder and recurrent depression, and refers to a mental disorder characterized by acutely and persistently depressed mood, accompanied by low self-esteem and loss of interest or pleasure in a normally enjoyable activity. Some people with clinical depression have trouble sleeping, lose weight, and generally feel anxious and irritable. Clinical depression affects the way a person feels, thinks and behaves, and can lead to a variety of emotional and physical problems. People with clinical depression may have problems with everyday activities and it makes the person feel as if life is not worth living.

Postnatal depression (PND) is also referred to as postpartum depression (PPD) and refers to a type of clinical depression that negatively affects women after childbirth. Symptoms may include sadness, fatigue, changes in sleeping and eating habits, decreased sexual desire, episodes of crying, restlessness and irritability. In some embodiments, PND is treatment-resistant depression (eg, treatment-resistant depression as described herein). In some embodiments, PND is refractory depression (eg, refractory depression as described herein).

Atypical depression (AD) is characterized by mood reactivity (eg, paradoxical anhedonia) and positivity, significant weight gain, or increased appetite. Patients suffering from AD may also have increased sleep duration or drowsiness (hypersomnia), a feeling of heavy limbs, and significantly reduced socialization due to hypersensitivity to perceived interpersonal rejection.

Melancholic depression is characterized by loss of pleasure (anhedonia) in most or all activities, failure to respond to pleasant stimuli, depressed mood more pronounced than with sadness or loss, excessive weight loss, or excessive guilt.

Psychotic major depression (PGD) or psychotic depression refers to a major depressive episode, particularly of a melancholic nature, in which a person experiences psychotic symptoms such as delusions and hallucinations.

Catatonic depression refers to major depression characterized by disturbances in motor behavior and other symptoms. The person may become mute and stuporous, and is either motionless or exhibits aimless or bizarre movements.

Seasonal affective disorder (SAD) is a type of seasonal depression where a person has a seasonal pattern of depressive episodes that occur in the fall or winter.

Dysthymia refers to a condition associated with unipolar depression, where the same physical and cognitive problems are expressed. They are not as severe and tend to last longer (eg at least 2 years).

Double depression refers to a fairly depressed mood (dysthymia) that lasts for at least 2 years and is interrupted by periods of major depression.

Depressive personality disorder (DPD) refers to a personality disorder with depressive features.

Recurrent brief depression (RTD) refers to a condition in which people have depressive episodes approximately once a month, with each episode lasting 2 weeks or less and usually lasting less than 2-3 days.

Minor depressive disorder or minor depression refers to depression in which at least 2 symptoms are present for 2 weeks.

Bipolar disorder or manic depressive disorder causes extreme mood swings that include emotional highs (mania or hypomania) and lows (depression). During periods of mania, a person may feel or act abnormally happy, energetic, or irritable. They often make poorly thought out decisions with little regard for the consequences. The need for sleep usually decreases. During periods of depression, crying, poor eye contact with others, and a negative outlook on life may occur. The risk of suicide among those with the disorder is high, over 6% over 20 years, while self-harm occurs in 30-40%. Other mental health problems, such as anxiety disorder and substance use disorder, are commonly associated with bipolar disorder.

Chronic illness depression refers to depression caused by chronic illnesses such as cancer or chronic pain, chemotherapy, chronic stress.

Treatment-resistant depression refers to a condition in which people have been treated for depression but there is no relief from symptoms. For example, antidepressants or physiological counseling (psychotherapy) do not improve depressive symptoms in people with treatment-resistant depression. In some cases, people with treatment-resistant depression experience relief from their symptoms but return to their previous symptoms. Refractory depression occurs in depressed patients who are resistant to standard pharmacological treatments, including tricyclic antidepressants, MAOIs, SSRIs, and double and triple neurotransmitter reuptake inhibitors and/or anxiolytic drugs, as well as non-pharmacological treatments (eg, psychotherapy, electroconvulsive therapy, vagus nerve stimulation and/or transcranial magnetic stimulation).

Suicidality, suicidal thoughts, and suicidal behavior refer to a person's tendency to commit suicide. Suicidal ideation refers to thoughts or unusual preoccupation with suicide. The range of suicidal ideation varies greatly, for example, from fleeting thoughts to large-scale thoughts, detailed planning, imitative actions, incomplete attempts. Symptoms include talking about suicide, obtaining means to commit suicide, withdrawing from social contact, preoccupation with death, feeling trapped or hopeless about the situation, increasing alcohol or drug use, doing risky or suicidal things, saying goodbye to people as if there were more of them they won't see.

Premenstrual dysphoric disorder (PMDD) refers to a severe, sometimes disabling extension of premenstrual syndrome (PMS). PMDD causes excessive mood changes, with symptoms that typically begin seven to ten days before a woman's menstrual cycle begins and continue through the first few days of a woman's menstrual cycle. Symptoms include sadness or hopelessness, anxiety or tension, extreme mood swings, and extreme irritability or anger.

Symptoms of depression include persistent anxious or sad feelings, feelings of helplessness, hopelessness, pessimism, feelings of worthlessness, low energy, restlessness, irritability, fatigue, loss of interest in enjoyable activities or hobbies, lack of positive thoughts or plans, excessive sleepiness, overeating, loss of appetite, insomnia, self-harm, suicidal thoughts and suicide attempts. The presence, severity, frequency and duration of symptoms may vary from case to case. Symptoms of depression and its relief can be determined by a doctor or psychologist (for example, through a mental state examination).

Anxiety disorders

The solutions or additives described herein may be used in a method described herein, for example, in the treatment of a disorder described herein, such as an anxiety disorder.

Anxiety disorder is an umbrella term that covers several different forms of abnormal and pathological fear and anxiety. Current psychiatric diagnostic criteria identify a wide variety of anxiety disorders.

Generalized anxiety disorder is a common chronic condition characterized by long-term anxiety that is not focused on any one object or situation. Individuals suffering from generalized anxiety experience nonspecific, persistent fear and worry and become overly preoccupied with daily activities. Generalized anxiety disorder is the most common anxiety disorder affecting older adults.

In panic disorder, a person suffers from short-term attacks of intense terror and apprehension, often marked by tremors, trembling, confusion, dizziness, nausea, and difficulty breathing. These panic attacks, defined by the APA as fear or discomfort that suddenly arises and peaks in less than ten minutes, can last several hours and can be caused by stress, fear, or even exercise; although the specific reason is not always obvious. In addition to recurrent unexpected panic attacks, a diagnosis of panic disorder also requires that said attacks have chronic consequences: either worry about the potential consequences of the attacks, persistent fear of future attacks, or significant changes in behavior associated with the attacks. Accordingly, those who suffer from panic disorder experience symptoms even outside of specific panic episodes. Often, panic sufferers notice normal changes in their heartbeat, which makes them think that something is wrong with their heart or that they are having another panic attack. In some cases, increased attention (hypervigilance) to the functioning of the body occurs during panic attacks, with any perceived physiological change interpreted as a possible life-threatening illness (eg, extreme hypochondriasis).

Obsessive-compulsive disorder is a type of anxiety disorder primarily characterized by recurring obsessions (anxious, persistent, and intrusive thoughts or images) and compulsions (urges to perform specific actions or rituals). The OCD mindset can be compared to superstition because it involves believing in a causal relationship where in reality there is none. Often the process is completely illogical; for example, pressure to walk in a certain way may be used to relieve obsessions about imminent negative affect. And in many cases the compulsion is completely inexplicable, just a desire to complete the ritual caused by nervousness. In a minority of cases, OCD sufferers may experience only obsession, without overt compulsions; far fewer sufferers experience only compulsions.

The largest category of anxiety disorders is the category of phobias, which includes all cases where fear and anxiety are caused by a specific stimulus or situation. Sufferers typically expect horrifying consequences from encountering the object of their fear, which can be anything from an animal to a place to a bodily fluid.

Post-traumatic stress disorder , or PTSD , is an anxiety disorder that occurs as a result of a traumatic experience. PTSD can occur as a result of an emergency situation such as combat, rape, hostage taking, or even a serious accident. It may also result from long-term (chronic) exposure to a severe stressor, such as soldiers who endure isolated battles but cannot cope with continuous combat. Common symptoms include flashbacks, avoidance reactions, and depression.

Eating disorders

The solutions or mixtures described herein may be used in a method described herein, for example, in the treatment of a disorder described herein, such as an eating disorder. Eating disorders are characterized by disturbances in eating behavior and weight regulation and are associated with a wide range of adverse psychological, physical and social consequences. A person with an eating disorder may begin to eat less or more food, but at some point their desire to eat less or more becomes out of control. Eating disorders may be characterized by severe distress or preoccupation with body weight or shape, or extreme efforts to regulate weight or food intake. Eating disorders include anorexia nervosa, bulimia nervosa, binge eating disorder, cachexia, and variants thereof.

Individuals with anorexia nervosa typically see themselves as overweight even when they are underweight. Individuals with anorexia nervosa may become obsessed with eating, eating, and weight control. Individuals with anorexia nervosa typically weigh themselves constantly, carefully portion out food, and eat very small amounts of only certain foods. Individuals with anorexia nervosa may engage in binge eating followed by crash dieting, excessive exercise, self-induced vomiting, or abuse of laxatives, diuretics, or enemas. Symptoms include extremely low body weight, severe dietary restriction, relentless pursuit of thinness and reluctance to maintain a normal or healthy weight, intense fear of gaining weight, a distorted sense of body image and self-esteem that is heavily influenced by subjective assessment of weight and body shape, or denial of seriousness low body weight, absence of menstruation among girls and women. Other symptoms include thinning bones, brittle hair and nails, dry and yellowish skin, fine hair growth throughout the body, mild anemia, muscle loss and weakness, severe constipation, low blood pressure or slow breathing and pulse, abnormal heart structure and function, damage brain, multiple organ failure, decreased core body temperature, lethargy, lethargy and infertility.

People with bulimia nervosa have recurrent and frequent episodes of eating unusually large amounts of food and a lack of control over these episodes. Binge eating is followed by behaviors that compensate for the overeating, such as forced vomiting, excessive use of laxatives or diuretics, fasting, excessive exercise, or a combination of these behaviors.

Unlike anorexia nervosa, people with bulimia nervosa usually maintain what is considered a healthy or normal weight, while some are overweight. But like people with anorexia nervosa, they tend to be afraid of gaining weight, desperate to lose weight, and unhappy with their body size and shape. Bulimic behavior usually occurs in secret because it is often accompanied by feelings of disgust or shame. The cycle of overeating and laxative use can occur anywhere from a few times a week to many times a day. Other symptoms include chronic inflammation and sore throat, swollen salivary glands in the neck and jaw, worn tooth enamel and increasingly sensitive and rotting teeth as a result of exposure to stomach acid, acid reflux and other gastrointestinal diseases, intestinal upset and irritation from overuse laxatives, severe dehydration due to purging of fluids, electrolyte imbalance (which can lead to heart attack or stroke).

People with compulsive overeating disorder lose control over eating. Unlike bulimia nervosa, periods of binge eating are not accompanied by compensatory behaviors such as laxative use, excessive exercise, or fasting. People with binge eating disorder are often overweight or obese. Obese people with binge eating disorder are at higher risk of developing cardiovascular disease and high blood pressure. They also experience guilt, shame, and distress from their overeating, which can lead to more overeating.

Cachexia is also known as “wasting syndrome” and is a food-related problem that many cancer patients experience. People with cachexia can continue to eat normally, but their body may refuse to use the vitamins and nutrients it ingests, or they may lose their appetite and stop eating. When a person experiences loss of appetite and stops eating, they may be considered to have advanced anorexia nervosa.

Epilepsy

The solutions or mixtures described herein can be used in the method described herein, for example, in the treatment of a disorder described herein, such as epilepsy, status epilepticus or seizure disorder, for example, as described in WO2013/112605 and WO/2014/031792, the contents of which are included in this application in its entirety.

Epilepsy is a brain disorder characterized by recurrent seizures over time. Types of epilepsy may include, but are not limited to, generalized epilepsy such as childhood absence epilepsy, juvenile myoclonic epilepsy, epilepsy with generalized tonic-clonic seizures on awakening, West syndrome, Lennox-Gastaut syndrome, partial epilepsy such as temporal lobe epilepsy lobes, frontal lobe epilepsy, benign focal epilepsy in children.

Status epilepticus (SE)

Status epilepticus (SE) may include, for example, convulsive status epilepticus, eg, early status epilepticus, persistent status epilepticus, refractory status epilepticus, super-refractory status epilepticus; non-convulsive status epilepticus, eg generalized status epilepticus, complex partial status epilepticus; generalized periodic epileptiform discharges; and periodic lateral epileptiform discharges. Convulsive status epilepticus is characterized by the presence of convulsive status epilepticus seizures and can include early status epilepticus, persistent status epilepticus, refractory status epilepticus, and super-refractory status epilepticus. Early status epilepticus is treated with first-line therapy. Persistent status epilepticus is characterized by status epileptic seizures that persist despite treatment with first-line therapy and require the introduction of second-line therapy. Refractory status epilepticus is characterized by status epileptic seizures that persist despite treatment with first-line and second-line therapy and are usually administered with a general anesthetic. Super-refractory status epilepticus is characterized by status epileptic seizures that persist despite treatment with first-line therapy, second-line therapy, and general anesthesia for 24 hours or more.

Non-convulsive status epilepticus may include, for example, focal non-convulsive status epilepticus, eg, complex partial non-convulsive status epilepticus, simple partial non-convulsive status epilepticus, mild non-convulsive status epilepticus; generalized nonconvulsive status epilepticus, such as late-onset absence nonconvulsive status epilepticus, atypical absence nonconvulsive status epilepticus, or typical absence nonconvulsive status epilepticus.

The compositions described herein may also be administered as a prophylactic agent to a subject having a CNS disorder, e.g., traumatic brain injury, status epilepticus, e.g., convulsive status epilepticus, e.g., early status epilepticus, persistent status epilepticus, refractory status epilepticus, super-refractory status epilepticus; non-convulsive status epilepticus, eg generalized status epilepticus, complex partial status epilepticus; generalized periodic epileptiform discharges; and periodic lateral epileptiform discharges; before the onset of a seizure.

Seizure

A seizure is a physical or behavioral change that occurs after an episode of abnormal electrical activity in the brain. The term "seizure" is often used interchangeably with "convulsion." Convulsions are when a person's body shakes quickly and involuntarily. During seizures, a person's muscles constantly contract and relax.

Based on the type of behavior and brain activity, seizures are divided into two broad categories: generalized and partial (also called localized or focal). Classifying the type of seizure helps doctors diagnose whether a patient has epilepsy.

Generalized seizures are generated by electrical impulses throughout the brain, whereas partial seizures are generated (at least initially) by electrical impulses in a relatively small part of the brain. The part of the brain that generates seizures is sometimes called the focus.

There are six types of generalized seizures. The most common and significant, and therefore the best known, is the generalized seizure, also called grand mal seizure. In this type of seizure, the patient becomes unconscious and usually falls. Loss of consciousness is accompanied by a generalized stiffening of the body (called the "tonic" phase of the seizure) for 30-60 seconds, then a sharp jerking ("clonic" phase) for 30-60 seconds, after which the patient goes into deep sleep ("post-convulsive" or postconvulsant phase). During grand mal seizures, injuries and accidents such as tongue biting and urinary incontinence can occur.

Absence seizures cause a brief loss of consciousness (just a few seconds) with few or no symptoms. The patient, most often a child, usually stops activity and stares blankly. These attacks begin and end suddenly and may occur several times a day. Patients usually do not know that they are having an attack, except that they may be aware of the "loss of time."

Myoclonic seizures consist of sporadic jerking movements, usually on both sides of the body. Patients sometimes describe the twitching as short-term electrical shocks. When severe, these seizures can lead to falling or involuntarily throwing objects.

Clonic seizures are repetitive, rhythmic jerking movements that involve both sides of the body at the same time.

Tonic seizures are characterized by stiffening of the muscles.

Atonic seizures consist of a sudden and general loss of muscle tone, especially in the arms and legs, often leading to falls.

The seizures described herein may include epileptic seizures; acute recurrent seizures; cluster seizures; continuous seizures; incessant seizures; prolonged convulsive seizures; recurrent seizures; status epileptic seizures, for example, refractory convulsive status epilepticus, non-convulsive status epileptic seizures; refractory seizures; myoclonic seizures; tonic seizures; tonic-clonic seizures; simple partial seizures; complex partial seizures; secondary generalized seizures; atypical absence seizures; absence seizures; atonic seizures; benign Rolandic seizures; febrile seizures; emotional seizures; focal seizures; epileptic seizures of laughter; convulsive seizures with generalized onset; infantile spasms; Jacksonian epilepsy; massive bilateral myoclonic seizures; multifocal seizures; seizures with neonatal onset; nocturnal seizures; seizures of the occipital lobe of the brain; post-traumatic seizures; mild convulsive seizures; Sylvan's seizures; visual reflex epileptic seizures; or seizures caused by withdrawal symptoms.

Tremor

The solutions or mixtures described herein can be used in a method described herein, for example, in the treatment of a disorder described herein, such as tremor.

Tremor is an involuntary, sometimes rhythmic, muscle contraction and relaxation that may involve vibrations or jerking of one or more parts of the body (eg, arm, arm, eye, face, head, vocal cords, torso, legs).

Cerebellar tremor , or intention tremor, is a slow, wide tremor of the limbs that occurs after a purposeful movement. Cerebellar tremor is caused by cerebellar metastases or damage to the cerebellum, such as due to a tumor, stroke, disease (eg, multiple sclerosis, hereditary degenerative disorder).

Dystonic tremor occurs in individuals who have dystonia, a movement disorder in which prolonged involuntary muscle contractions cause twisting and repetitive movements and/or painful and abnormal body positions or postures. Dystonic tremors can affect any muscle in the body. Dystonic tremors occur irregularly and can often be relieved by complete rest.

Essential tremor or benign essential tremor is the most common type of tremor. Essential tremor may be mild and non-progressive in some and may progress slowly starting on one side of the body but affecting both sides within 3 years. The hands are most often involved, but the head, voice, tongue, legs and torso may also be involved. The frequency of tremors may decrease as a person ages, but the severity may increase. Increased emotion, stress, fever, physical exhaustion, or low blood sugar can cause tremors and/or increase their severity.

Orthostatic tremor is characterized by rapid (eg, greater than 12 Hz) rhythmic muscle contractions that occur in the legs and trunk immediately after standing. Cramps are felt in the thighs and legs, and the patient may tremble involuntarily when asked to stand in one place. Orthostatic tremor may occur in patients with essential tremor.

Parkinsonian tremors are caused by damage to structures in the brain that control movement. Parkinsonian tremor is often a precursor to Parkinson's disease and is usually seen as a "pill rolling" type of hand action that can also involve the chin, lips, legs and torso. The onset of parkinsonian tremor usually begins after age 60. The movement begins with one limb or one side of the body and may progress to involve the other side.

Physiological tremor can occur in normal individuals and is not clinically significant. This can be seen in all voluntary muscle groups. Physiological tremors can be caused by certain drugs, alcohol, or medical conditions, including an overactive thyroid and hypoglycemia. Tremor classically has a frequency of approximately 10 Hz.

Psychogenic tremor or hysterical tremor can occur at rest or during postural or kinetic movement. A patient with psychogenic tremor may have conversion disorder or another mental illness.

Rubral tremor is characterized by a large-wave slow tremor that can be present at rest, in body position and with intention. Tremors are associated with conditions that affect the red nucleus in the midbrain, classic unusual tremors.

Anesthesia/sedation

The solutions or mixtures described herein can be used in the method described herein, for example, to induce anesthesia or sedation. Anesthesia is a pharmacologically induced and reversible state of amnesia, analgesia, loss of responsiveness, loss of skeletal muscle reflexes, decreased stress response, or all of these. These effects can be obtained from a single drug, which by itself provides the right combination of effects, or sometimes from a combination of drugs (eg, hypnotics, sedatives, paralytics, analgesics) to achieve very specific combinations of results. Anesthesia allows patients to undergo surgeries and other procedures without the distress and pain they might otherwise experience.

Sedation is the reduction of irritability or agitation by the administration of a pharmacological agent, usually to facilitate a medical or diagnostic procedure.

Sedation and analgesia involve a range of states of consciousness, ranging from minimal sedation (anxiolysis) to general anesthesia.

Minimal sedation is also known as anxiolysis. Minimal sedation is a drug-induced state during which the patient typically responds to verbal commands. Cognitive function and coordination may be impaired. Respiratory and cardiovascular functions are usually not affected.

Moderate sedation/analgesia (conscious sedation) is a drug-induced depression of consciousness during which the patient specifically responds to a verbal command, either independently or with the application of mild tactile stimulation. Usually no intervention is required to maintain an open airway. Spontaneous ventilation is usually sufficient. Cardiovascular function is usually preserved.

Deep sedation/analgesia is a drug-induced depression of consciousness during which the patient cannot readily respond to external stimuli, but responds purposefully (not a reflexive withdrawal from a painful stimulus) after repeated or painful stimulation. Independent respiratory function may be impaired and the patient may require assistance in maintaining an open airway. Spontaneous ventilation may be insufficient. Cardiovascular function is usually preserved.

General anesthesia is a drug-induced loss of consciousness during which the patient does not respond to external stimuli, even painful stimuli. The ability to maintain independent respiratory function is often impaired and assistance is often required to maintain an open airway. Positive pressure ventilation may be required due to inhibition of spontaneous ventilation or drug-induced depression of neuromuscular function. Cardiovascular function may be impaired.

Sedation in the intensive care unit (ICU) can reduce patients' perception of the environment and reduce their response to external stimuli. It can play a role in the treatment of a critically ill patient and covers a wide range of symptom control that will vary among patients and individuals over the course of their illness. Heavy sedation in intensive care has been used to facilitate endotracheal tube tolerance and ventilator synchronization, often with the use of neuromuscular blockers.

In some embodiments, sedation (eg, long-term sedation, continuous sedation) is induced and maintained in the ICU for an extended period of time (eg, 1 day, 2 days, 3 days, 5 days, 1 week, 2 weeks, 3 weeks, 1 month , 2 months). Long-term sedation agents may have a longer duration of action. ICU sedatives may have a short half-life.

Procedural sedation and analgesia, also referred to as conscious sedation, is a method of administering sedatives or dissociative agents with or without analgesics to induce a state that allows the subject to tolerate unpleasant procedures while maintaining cardiorespiratory function.

Methods of administration

An aqueous solution or mixture described herein containing a therapeutically effective amount of a neuroactive steroid, cyclodextrin and a buffer can be administered parenterally (e.g., intranasally, buccally, intravenously, or intramuscularly, e.g., intramuscular injection (IM) or IV).

In one embodiment, the aqueous solution or mixture containing the neuroactive steroid is administered at a dose equivalent to the parenteral administration of the neuroactive steroid from about 0.1 ng to about 100 g per kg body weight, from about 10 ng to about 50 g per kg body weight, about 100 ng to about 1 g per kg body weight, about 1 μg to about 100 mg per kg body weight, about 10 μg to about 10 mg per kg body weight, about 100 μg to about 5 mg per kg body weight body, from about 250 mcg to about 3 mg per kg body weight, from about 500 mcg to about 2 mg per kg body weight, from about 1 mcg to about 50 mg per kg body weight, from about 1 mcg to about 500 mcg per kg body weight; and from about 1 μg to about 50 μg per kg of body weight. Alternatively, the amount of an aqueous solution or mixture containing the neuroactive steroid administered to achieve a therapeutic effective dose is approximately 0.1 ng, 1 ng, 10 ng, 100 ng, 1 μg, 10 μg, 100 μg, 1 mg, 1.5 mg, 2 mg, 3 mg, 4 mg, 5 mg, 6 mg, 7 mg, 8 mg, 9 mg, 10 mg, 11 mg, 12 mg, 13 mg, 14 mg, 15 mg, 16 mg, 17 mg, 18 mg, 19 mg, 20 mg, 30 mg, 40 mg, 50 mg, 60 mg, 70 mg, 80 mg, 90 mg, 100 mg, 500 mg per kg body weight or more neuroactive steroid.

In one embodiment, an aqueous solution or mixture containing a neuroactive steroid is administered as an intravenous bolus infusion at a dose equivalent to a parenteral administration of the neuroactive steroid from about 0.1 ng to about 100 g per kg of body weight, from about 10 ng to about 50 g per kg body weight, from about 100 ng to about 1 g per kg body weight, from about 1 μg to about 100 mg per kg body weight, from about 1 μg to about 50 mg per kg body weight, from about 10 μg to about 5 mg per kg body weight, from about 100 mcg to about 500 mcg per kg body weight, from about 100 mcg to about 400 mcg per kg body weight, from about 150 mcg to about 350 mcg per kg body weight, from about 250 mcg up to approximately 300 mcg per kg body weight. In one embodiment, an aqueous solution or mixture containing a neuroactive steroid is administered as an intravenous bolus infusion at a dose equivalent to a parenteral administration of the neuroactive steroid from about 100 to about 400 μg/kg. In some embodiments, an aqueous solution or mixture containing a neuroactive steroid is administered as an intravenous bolus infusion containing about 150 to about 350 μg/kg of neuroactive steroid. In some embodiments, an aqueous solution or mixture containing a neuroactive steroid is administered as an intravenous bolus infusion containing about 250 to about 300 μg/kg of neuroactive steroid. In specific embodiments, an aqueous solution or mixture containing a neuroactive steroid is administered as an intravenous bolus infusion at a dose equivalent to approximately 100 mcg/kg, 125 mcg/kg, 150 mcg/kg, 175 mcg/kg, 200 mcg/kg, 225 mcg/kg, 250 mcg/kg, 260 mcg/kg, 270 mcg/kg, 280 mcg/kg, 290 mcg/kg, 300 mcg/kg, 325 mcg/kg, or 350 mcg/kg neuroactive steroid.

In one embodiment, an aqueous solution or mixture containing a neuroactive steroid is administered as an intravenous bolus infusion at a dose equivalent to a parenteral administration of the neuroactive steroid from about 0.1 nmol/L to about 100 μmol/L per kg body weight, from about 1 nmol /L to approximately 10 µmol/L per kg body weight, from approximately 10 nmol/L to approximately 10 µmol/L per kg body weight, from approximately 100 nmol/L to approximately 10 µmol/L per kg body weight, from approximately 300 nmol/L to about 5 μmol/L per kg body weight, from about 500 nmol/L to about 5 μmol/L per kg body weight, and from about 750 nmol/L to about 1 μmol/L per kg body weight. Alternatively, the amount of an aqueous solution or mixture containing the neuroactive steroid administered to achieve a therapeutic effective dose is approximately 0.1 ng, 1 ng, 10 ng, 100 ng, 1 μg, 10 μg, 100 μg, 1 mg, 1.5 mg, 2 mg, 3 mg, 4 mg, 5 mg, 6 mg, 7 mg, 8 mg, 9 mg, 10 mg, 11 mg, 12 mg, 13 mg, 14 mg, 15 mg, 16 mg, 17 mg, 18 mg, 19 mg, 20 mg, 21 mg, 22 mg, 23 mg, 24 mg, 25 mg, 26 mg, 27 mg, 28 mg, 29 mg, 30 mg, 40 mg, 50 mg, 60 mg, 70 mg , 80 mg, 90 mg, 100 mg, 500 mg per kg body weight or more of a neuroactive steroid.

In some embodiments, the aqueous solution or mixture containing the neuroactive steroid may be administered one or more times per day. The duration of treatment may be, for example, once a day for about 1, 2, 3, 4, 5, 6, 7 days or more. In some embodiments, the implementation is administered either as a single dose as an individual unit dose, or multiple smaller unit doses, or through multiple administrations of divided doses at specified intervals. For example, a unit dose may be administered over a period of time from about 0 hour to about 1 hour, from about 1 hour to about 24 hours, from about 1 hour to about 72 hours, from about 1 hour to about 120 hours, or from about 24 hours to at least approximately 120 hours after injury. Alternatively, a unit dose may be administered at approximately 0.5, 1, 1.5, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 30, 40, 48, 72, 96, 120 hours or more after injury. Subsequent unit doses may be administered at any time after the initial administration so as to achieve a therapeutic effect. For example, additional unit doses may be administered to protect the subject from a secondary wave of swelling that may occur during the first few days after injury.

In some embodiments, administration of an aqueous solution or mixture containing a neuroactive steroid includes a period of time during which administration is withdrawn.

As used herein, the term "withdrawal" or "withdrawal dose" refers to an administration protocol that reduces the dose administered to a patient and thus results in a gradual reduction and eventual removal of an aqueous solution or mixture containing a neuroactive steroid, either over a fixed period of time. , or for a period of time determined empirically by a physician's assessment based on regular observation of the subject's therapeutic response. The withdrawal period may be approximately 12, 24, 36, 48 hours or longer. Alternatively, the period of withholding may be from about 1 to 12 hours, from about 12 to about 48 hours, or from about 24 to about 36 hours. In some embodiments, the withdrawal period is approximately 24 hours.

The applied failure may be a "linear" failure. For example, a "10%" linear failure of 500 mg would be 500, 450, 400, 350, 300, 250, 200, 150, 100, 50. Alternatively, exponential failure could be used, which if the example above is used program, exponential failure would be, for example, 500, 450, 405, 365, 329, 296, 266, 239, etc. Accordingly, approximately 5%, 10%, 15%, 20%, 25%, 30%, 35% or 40% linear or exponential failure can be used in the methods of the invention. In addition, a linear or exponential rejection of about 1 to 5%, about 6% to 10%, about 11% to 15%, about 16% to 20%, about 21% to 25%, about from 26% to 30%, from about 31% to 35%, from about 36% to 40%.

In other embodiments, administration of an aqueous solution or mixture containing a neuroactive steroid includes a finite period of time during which the administration of the neuroactive steroid is gradually reduced.

As used herein, the terms “taper administration,” “taper dose,” and “gradual dose reduction” refer to an administration protocol that reduces the dose administered to a patient and thereby provides a gradual reduction and eventual removal of an aqueous solution or mixture containing a neuroactive steroid, either for a fixed period of time or for a period of time determined empirically by a physician's assessment based on regular observation of the subject's therapeutic response. The taper period may be approximately 12, 24, 36, 48 hours or more. Alternatively, the taper period may be from about 1 to 12 hours, from about 12 to about 48 hours, or from about 24 to about 36 hours. In some embodiments, the taper period is approximately 24 hours.

The ramp applied may be a "linear" ramp. For example, a linear "10%" taper from 500 mg would be 500, 450, 400, 350, 300, 250, 200, 150, 100, 50 mg. An alternative is to use exponential fade-out, which if the above program is used as an example, exponential fade-out would be, for example, 500, 450, 405, 365, 329, 296, 266, 239, etc. Accordingly, 5%, 10%, 15%, 20%, 25%, 30%, 35% or 40% linear or exponential reduction can be used in the methods of the invention. In addition, a linear or exponential reduction of about 1 to 5%, about 6% to 10%, about 11% to 15%, about 16% to 20%, about 21% to 25%, can be used. about 26% to 30%, about 31% to 35%, about 36% to 40%. In some embodiments, the drug dose taper is approximately a 25% linear taper.

In one embodiment, an aqueous solution or mixture containing a neuroactive steroid is administered as an intravenous infusion in an amount of neuroactive steroid/time unit from about 20 to about 5000 μg/kg/hour. In some embodiments, the neuroactive steroid in a maintenance cycle is administered as an intravenous infusion in an amount of neuroactive steroid/time unit from about 20 to about 2500 μg/kg/hour. In some embodiments, the neuroactive steroid in a maintenance cycle is administered as an intravenous infusion in an amount of neuroactive steroid/time unit from about 20 to about 500 μg/kg/hour. In some embodiments, the neuroactive steroid is administered as an intravenous infusion at a rate of from about 20 to about 250 mcg/kg/hour. In some embodiments, the neuroactive steroid is administered as an intravenous infusion in an amount of neuroactive steroid/time unit from about 20 to about 200 μg/kg/hour. In some embodiments, the neuroactive steroid is administered as an intravenous infusion in an amount of neuroactive steroid/time unit from about 20 to about 150 μg/kg/hour. In some embodiments, the neuroactive steroid is administered as an intravenous infusion in an amount of neuroactive steroid/time unit from about 50 to about 100 μg/kg/hour. In some embodiments, the neuroactive steroid is administered as an intravenous infusion in an amount of neuroactive steroid/time unit from about 70 to about 100 μg/kg/hour. In specific embodiments, the neuroactive steroid is administered as an intravenous infusion at an amount of neuroactive steroid/time unit of approximately 25 mcg/kg/hour, 50 mcg/kg/hour, 75 mcg/kg/hour, 80 mcg/kg/hour, 85 mcg/hour. kg/hour, 86 mcg/kg/hour, 87 mcg/kg/hour, 88 mcg/kg/hour, 89 mcg/kg/hour, 90 mcg/kg/hour, 100 mcg/kg/hour, 125 mcg/kg /hour, 150 mcg/kg/hour or 200 mcg/kg/hour.

In one embodiment, an aqueous solution or mixture containing a neuroactive steroid is administered as an intravenous infusion at a dose equivalent to a parenteral administration of the neuroactive steroid from about 0.1 ng to about 100 g per kg of body weight, from about 10 ng to about 50 g per kg body weight, from about 100 ng to about 1 g per kg body weight, from about 1 μg to about 100 mg per kg body weight, from about 1 μg to about 50 mg per kg body weight, from about 10 μg to about 5 mg per kg body weight; and from about 100 μg to about 1000 μg per kg of body weight. In one embodiment, an aqueous solution or mixture containing a neuroactive steroid is administered as an intravenous infusion at a dose equivalent to a parenteral administration of the neuroactive steroid from about 0.1 nmol/L to about 100 μmol/L per kg body weight, from about 1 nmol/L L to approximately 10 µmol/L per kg body weight, from approximately 10 nmol/L to approximately 10 µmol/L per kg body weight, from approximately 100 nmol/L to approximately 10 µmol/L per kg body weight, from approximately 300 nmol /L to about 5 µmol/L per kg body weight, from about 500 nmol/L to about 5 µmol/L per kg body weight and from about 750 nmol/L to about 5 µmol/L per kg body weight. Alternatively, the amount of an aqueous solution or mixture containing the neuroactive steroid administered to achieve a therapeutic effective dose is approximately 0.1 ng, 1 ng, 10 ng, 100 ng, 1 μg, 10 μg, 100 μg, 1 mg, 1.5 mg, 2 mg, 3 mg, 4 mg, 5 mg, 6 mg, 7 mg, 8 mg, 9 mg, 10 mg, 11 mg, 12 mg, 13 mg, 14 mg, 15 mg, 16 mg, 17 mg, 18 mg, 19 mg, 20 mg, 21 mg, 22 mg, 23 mg, 24 mg, 25 mg, 26 mg, 27 mg, 28 mg, 29 mg, 30 mg, 40 mg, 50 mg, 60 mg, 70 mg , 80 mg, 90 mg, 100 mg, 500 mg neuroactive steroid per kg body weight or higher.

As used herein, the term "about" means approximately plus or minus ten percent.

EXAMPLES

Example 1 Degradation Pathway of Allopregnanolone in SBECD Formulations

Figure 1 shows the two main degradation pathways found for allopregnanolone in SBECD formulations. Based on the data described in Figures 3-5 and 8 and Tables 1-11 and 16 , the major degradation pathway observed at pH ~6 or less is the epimerization of allopregnanolone to compound 1269 . Based on the data presented in Figures 3-5 and 8 and Tables 1-11 and 16 , the major degradation pathway observed at pH ~6 or greater is the oxidation of allopregnanolone to compound 136 .

The solubility of allopregnanolone was determined in sulfobutylether-β-cyclodextrin without buffer. A graphical representation of allopregnanolone versus cyclodextrin is shown in Figure 2 .

Example 2: Allopregnanolone in sulfobutylether-β-cyclodextrin without buffer.

A formulation of allopregnanolone (5 mg/mL) in 250 mL/mL sulfobutylether-β-cyclodextrin was prepared without buffer and packaged in a type I glass vial.

Specifically, the formulation was prepared by dissolving the required amount of Betadex sodium sulfobutyl ether (i.e., sulfobutyl ether-β-cyclodextrin) in approximately 80% of the required amount of sterile water for injection (SWI) in a suitable vessel using a standard paddle stirrer at 35-40° C. Allopregnanolone was added to an unbuffered solution of Betadex sodium sulfobutyl ether (ie, sulfobutyl ether-β-cyclodextrin) and mixed until dissolved using a high shear mixer. High shear stirring at 35-40°C was continued until the solution became noticeably clear, indicating dissolution of the allopregnanolone drug substance. The bulk solution was brought to final volume using SWI and mixed. The solution was filtered through a 0.45 µm pre-filter and aseptically filtered through suitable duplicate sterile 0.2 µm filters (such as Millipore PVDF) into a pre-sterilized filling vessel. The sterile solution was aseptically filled into pre-sterilized vials, sealed with pre-sterilized stoppers, and the stoppers were secured to the vials using an aluminum crimp cap. Filled vials were checked 100% for visible particles and defects in container closure, samples were taken to control the finished product and stored at 2-8°C.

Stability results showed a decreasing trend in pH and signs of degradation (formation of compounds 136 and 1269 ) that occurred faster at higher temperatures. The presence of degradation products at higher temperatures makes the allopregnanolone formulation chemically unstable under these conditions. Unstable composition limits shelf life for use in human clinical trials and potential commercial applications.

In Table 1, formulations of allopregnanolone (5 mg/ml) in 250 mg/ml sulfobutylether-β-cyclodextrin without buffer were monitored for 9 months at 25°C/60% relative humidity. The pH, the content of the main substance, the amount of impurities and mechanical inclusions were recorded.

Composition stability

Table 1 . Formulation of 5 mg/ml allopregnanolone in 250 mg/ml SBECD, 20 ml vials, unbuffered, non-autoclaved and stored at 25°C/60% RH for 9 months

Index Original 1 month 3 months 4 months 6 months 7 months 9 months Appearance Compliant Compliant Compliant Compliant Compliant Compliant Compliant pH 5.4 5.4 4.8 4.5 4.3 4.1 4.1 Main substance content 102.7 102.2 101.8 101.2 101.0 102.2 100.8 Related substances according to HPLC
Known impurities ¹
(square %) 136 Not detected 0.17 0.44 0.56 0.74 0.93 1.26 1269 Not detected Not detected Not detected Not detected 0.14 0.14 0.20 Mechanical inclusions Quantity ≥ 10 µm:
Quantity ≥ 25 µm: 35
7 eleven
0 48
16 22
0 52
3 49
5 Opaque

In Table 2, formulations of allopregnanolone (5 mg/ml) in 250 mg/ml sulfobutylether-β-cyclodextrin without buffer were monitored for 3 months at 40°C/75% relative humidity. The pH, the content of the main substance, the amount of impurities and mechanical inclusions were recorded.

Table 2 . Formulation of 5 mg/ml allopregnanolone in 250 mg/ml SBECD, in 20 ml vials, unbuffered, non-autoclaved and stored at 40°C/75% RH for 3 months

Index Original 1 month 3 months Appearance Compliant Compliant Compliant pH 5.4 5.4 4.7 Main substance content 102.7 101.4 99.9 Related substances according to HPLC
Known impurities (area %) 136 Not detected 0.58 2.87 1269 Not detected 0.10 0.42 Mechanical inclusions Quantity ≥ 10 µm
Quantity ≥ 25 µm 35
7 23
1 51
3

In Table 3 , formulations of allopregnanolone (5 mg/ml) in 250 mg/ml sulfobutylether-β-cyclodextrin without buffer were monitored for 6 months at 40°C/75% relative humidity. The pH, the content of the main substance, the amount of impurities and mechanical inclusions were recorded.

Table 3 . Formulation of 5 mg/ml allopregnanolone in 250 mg/ml SBECD, in 20 ml vials, unbuffered, non-autoclaved and stored at 40°C/75% RH for 6 months

Index Original 1 month 3 months 6 months Appearance Compliant Compliant Compliant Compliant pH 5.6 5.2 4.9 4.3 Main substance content 98.2 98.0 98.3 96.9 Related substances according to HPLC
Known impurities (area %) 136 Not detected 0.15 0.31 0.57 1269 Not detected Not detected 0.14 0.48 Mechanical inclusions Quantity ≥ 10 µm
Quantity ≥ 25 µm 115
3 80
7 80
eleven 78
4

Example 3 Allopregnanolone in sulfobutylether-β-cyclodextrin with buffer.

A formulation of allopregnanolone (5 mg/mL) in 250 mg/mL sulfobutylether-β-cyclodextrin was prepared using citrate buffer and packaged in a type I glass vial.

Seven allopregnanolone solutions were prepared as described in Table 4 . Samples of each of the seven solutions were autoclaved at 121°C for 30, 60 and 90 minutes. The solutions were stored at room temperature until testing. Table 5 shows the initial pH values for solutions.

Table 4 . Compositions of allopregnanolone formulations obtained for testing

Component Control 5 mM
pH 5.5 5 mM
pH 6.0 5 mM
pH 6.5 10 mM
pH 5.5 10 mM
pH 6.0 10 mM
pH 6.5 Hello
(g/l) 5 Citric acid monohydrate (g/l) Not applicable 0.25 0.13 0.05 0.51 0.27 0.11 Sodium citrate dihydrate (g/l) Not applicable 1.12 1.28 1.40 2.23 2.57 2.79 Sodium hydroxide or citric acid
how much is needed Until pH is reached Captisol® (g/l) 250 Water for injections How much is needed up to 1 liter

Table 5 . Summary of pH of starting buffers

Buffer preparation Buffer concentration Target pH pH after adding citric acid/sodium citrate Final adjusted pH value 5 5.5 5.73 5.50 5 6.0 6.24 5.96 5 6.5 6.40 6.50 10 5.5 5.62 5.51 10 6.0 6.12 6.02 10 6.5 6.60 6.60

A comparison of base content values for non-autoclaved and autoclaved samples is shown in Table 6 . The data shows that the content of the main substance (%) was kept constant for all autoclaving times studied.

Table 6 . The effect of autoclaving on the content of the main substance of the product

Content of the main substance (% by LC) after autoclaving (121°C) Prototype Target pH Initial content of the main substance* 30 min 60 min 90 min Control Not applicable 101.0 99.9 100.4 99.5 Buffered 5 mM 5.5 101.9 101.4 101.2 100.6 6.5 101.3 101.6 102.6 101.4 Buffered 10 mM 5.5 101.8 100.0 99.2 99.7 6.5 100.6 99.9 100.3 99.5

*Content of main substance at time zero, non-autoclaved sample

Table 7 summarizes the composition of impurities, in particular the amount of compounds 136 and 1269 formed during short-term exposure to high temperatures.

An oxidative degradation product (compound 136 ) was observed at 0.13% in the control sample after autoclaving for 90 minutes. Similar levels were found in 90-minute buffered samples at pH 5.5 (0.15% for the 5 mM sample and 0.11% for the 10 mM sample). The 90-minute pH 6.5 buffered samples contained slightly lower levels of oxidative degradation product (0.05% for the 5 mM sample and 0.03% for the 10 mM sample).

Under the conditions of the present study, there is a noticeable and non-significant improvement in the absence of formation of compound 136 when using 10 mM buffer compared to 5 mM buffer.

It is important to note that during the thermal stress study, no epimerization was observed (formation of compound 1269) - (not detected) for the buffered formulations compared to 0.15% for the unbuffered control at the 90 minute time point.

Table 7 . Composition of impurities after autoclaving

Composition of impurities after autoclaving for 30, 60 or 90 minutes Prototype Target pH 30 min 60 min 90 min Control 136 Not applicable 0.10 0.10 0.13 1269 Not detected 0.11 0.15 Buffered 5 mM 136 5.5 0.06 0.11 0.15 1269 Not detected Not detected Not detected 136 6.5 0.02 0.03 0.05 1269 Not detected Not detected Not detected Buffered 10 mM 136 5.5 0.04 0.08 0.11 1269 Not detected Not detected Not detected 136 6.5 0.01 0.02 0.03 1269 Not detected Not detected Not detected

Table 8 summarizes the initial pH values, initial content of the main substance and impurities for each batch. The table includes non-autoclaved control samples along with initially autoclaved samples at T=0. Samples were analyzed for pH after storage for approximately 3 months at room temperature and for base and impurity content after approximately 4 months at room temperature.

Table 8 . Summary of Initial Major and Impurity Contents - Autoclaved vs. Non-autoclaved

Description Initial pH Initial content of the main substance, % by LC Total impurity content, % area pH Content of the main substance, % by LC Total impurity content, % area Autoclavable control 5.7 101.1 0.79 5.1 101.2 0.93 Non-autoclaved control 6.3 101.0 0.83 5.2 101.4 1.10 5 mM pH 5.5 autoclaved 5.1 101.6 0.84 5.0 101.5 0.94 5 mM pH 6.0 autoclaved 5.4 ** ** 5.4 102.0 1.25 5 mM pH 6.5 autoclaved 5.9 102.9 0.85 5.9 102.3 1.02 10 mM pH 5.5 autoclaved 5.1 101.5 0.83 5.1 100.6 0.84 10 mM pH 6.0 autoclaved 5.5 ** ** 5.6 99.6 0.84 10 mM pH 6.5 autoclaved 6.1 100.7 0.85 6.1 100.2 0.84 5 mM pH 5.5 non-autoclaved 5.1 101.9 0.84 5.1 101.1 0.84 5 mM pH 6.0 non-autoclaved 5.4 ** ** 5.4 102.1 1.04 5 mM pH 6.5 non-autoclaved 5.9 101.3 0.85 5.9 102.4 0.96 10 mM pH 5.5 non-autoclaved 5.1 101.8 0.88 5.1 100.4 0.84 10 mM pH 6.0 non-autoclaved 5.5 ** ** 5.5 99.8 0.84 10 mM pH 6.5 non-autoclaved 6.1 100.6 0.84 6.1 100.9 0.97

**Not studied at baseline

The pH of the unbuffered non-autoclaved control dropped by 1.1 pH units after 3 months of storage at room temperature, and the pH of the unbuffered non-autoclaved control dropped by 0.6 pH units after 3 months of storage at room temperature

The pH of the buffered solutions did not change significantly (the largest pH change recorded was 0.1 pH unit).

Both buffer concentrations of 5 and 10 mM provided good pH control after autoclaving and storage.

The initial data (T=0) for the content of the main substance (%) and the total content of impurities in general for the test samples showed a stable range of 100.6-102.9% and 0.79-0.85%, respectively. The content of the main substance in the samples at T=4 months corresponded to the samples at T=0 and did not show any signs of degradation. This was also the case for the total impurity content.

Large batches of allopregnanolone (5 mg/mL) in 250 mL/mL sulfobutylether-β-cyclodextrin were prepared using citrate buffer and packaged in a Type I glass vial.

Specifically, the formulation was prepared by dissolving the required amount of citric acid monohydrate (CAM) and sodium citrate dihydrate (SCD) in approximately 80% of the required amount of sterile water for injection (SWI) in a suitable vessel using a standard paddle mixer at 35-40°C. The required amount of Betadex sodium sulfobutyl ether (ie, sulfobutyl ether-β-cyclodextrin) was added to the buffer solution and stirred to dissolve. The pH of the product was checked and, if necessary, adjusted with hydrochloric acid or sodium hydroxide to pH 6.0 +/- 0.2. Allopregnanolone was added to Betadex sodium sulfobutyl ether (ie sulfobutyl ether-β-cyclodextrin) buffer solution and stirred until dissolved using a high shear mixer. High shear stirring at 35-40°C was continued until the solution became noticeably clear, indicating dissolution of the allopregnanolone drug substance. The pH of the product was checked and, if necessary, adjusted with hydrochloric acid or sodium hydroxide to ensure a pH of 6.0 +/- 0.1 of the product. The bulk solution was brought to final volume with SWFI and mixed. The solution was filtered through a 0.45 µm pre-filter and aseptically filtered through suitable duplicate sterile 0.2 µm filters (such as Millipore PVDF) into a pre-sterilized filling vessel. The sterile solution was aseptically filled into pre-sterilized vials, sealed with pre-sterilized stoppers, and the stoppers were secured to the vials using an aluminum crimp cap (component described in Table 9 ). Filled vials were checked 100% for visible particles and defects in container closure, samples were taken to control the finished product and stored at 2-8°C.

Table 9 . Types of packaging for formulations

Bottle Bottle number according to specification PC 3196 Description of the bottle USF Type 1 borosilicate glass bottle, 20 ml, 20 mm neck Manufacturer Schott Cork Plug number according to specification PC 4078 Description of the plug Item 19700021 or 19700022 in S10-F451 chlorobutyl B2-40 with FluroTec coating Manufacturer West Cap Description of the plug Aluminum cap 20 mm Manufacturer West Pharmaceutical Services Manufacturer Not applicable*

*Not related to the product. Different colors were used to differentiate between formulations.

In Table 10 , formulations of allopregnanolone (5 mg/ml) in 250 mg/ml sulfobutylether-β-cyclodextrin in 10 mM citrate buffer pH 6 were monitored for 6 months at 40°C/75% relative humidity. The pH, the content of the main substance (for example, the declared percentage), the amount of impurities and mechanical inclusions were recorded.

Table 10 . Injection of 5 mg/ml allopregnanolone in 250 mg/ml SBECD, in 20 ml vials, in 10 mM citrate buffer pH=6, stored at 40°C/75% relative humidity for 6 months

Index Original 1 month 3 months 6 months Appearance Compliant Compliant Compliant Compliant pH 5.8 5.7 5.8 5.9 Main substance content (%) 99.5 98.8 99.0 98.4 Related substances according to HPLC (wt %) 136 Not detected Not detected Not detected <0.10 1269 <0.10 <0.10 <0.10 0.12 Mechanical inclusions ≥ 10 µm 76 163 319 38 ≥ 25 µm 7 0 12 1

In Table 11 , formulations of allopregnanolone (5 mg/ml) in 250 mg/ml sulfobutylether-β-cyclodextrin in 10 mM citrate buffer pH 6 were monitored for 12 months at 25°C/60% relative humidity. The pH, the content of the main substance (for example, the declared percentage), the amount of impurities and mechanical inclusions were recorded.

Table 11 . Injection of 5 mg/ml allopregnanolone in 250 mg/ml SBECD, in 20 ml vials, in 10 mM citrate buffer pH=6, stored at 25°C/60% relative humidity for 12 months

Index Original 1 month 3 months 6 months 9 months 12 months Appearance Compliant Compliant Compliant Compliant Compliant Compliant pH 5.8 5.7 5.8 5.8 5.8 5.8 Main substance content (%) 99.5 99.5 99.6 97.6 98.6 99.3 Related substances according to HPLC (wt %) 136 Not detected Not detected Not detected Not detected Not detected Not detected 1269 <0.10 <0.10 <0.10 0.10 <0.10 0.10 Mechanical inclusions ≥ 10 µm 76 89 69 66 37 40 ≥ 25 µm 7 0 1 18 1 4

Example 4 Terminal Sterilization of Allopregnanolone Injection, 5 mg/ml in 250 mg/ml cyclodextrin (10 mM citrate buffer, pH 6.0) in 20 ml vials

Experiments were conducted to demonstrate that the sterilization process for injection of 5 mg/ml allopregnanolone in 250 mg/ml Captisol® (10 mM citrate buffer, pH=6.0, 20 ml/vial) ensures temperature uniformity and biodestruction throughout load volume using a Finn-Aqua steam sterilizer, including demonstrating that there is no increase in the known microbiological load of Geobacillus stearothermophilus .

The protocol established and validated the sterilization process, and determined where sterilizer load cells could be placed during normal product handling. Three (3) maximum load sterilizer cycles and three (3) minimum load sterilizer cycles were performed for each vial size using a Finn-Aqua steam sterilizer, model 91515-DP-RP-GMP-S7, serial number C0A41043. The Finn-Aqua steam sterilizer was a double door unit controlled by a Siemens Simatic S7-300 programmable logic controller (PLC). The sterilizer was powered by a user interface operated by an OP27 panel operator. The inner chamber dimensions were (w × h × d) 37 inches × 61 inches × 61 inches, for a total internal volume of 75 cu. ft. There was one cart that could be equipped with up to 15 shelves. Each shelf contained 8 trays of bottles (each tray held 162 20 ml bottles). "D-value" refers to the time required at temperature (T) to reduce the specific microbial population by 90% or the time required to reduce the number of survivors by a factor of 10 (1 log).

The maximum autoclave batch size of 259 L could hold approximately 12,690 vials. The minimum load for validation was 3 L based on the minimum batch size of a single tray autoclave.

The product was aseptically filled into the sterile working chamber of the production equipment, which was supported by simulating the aseptic processing process (media filling). These aseptic processing evaluations confirmed that the product had a Sterility Assurance Level (SAL) of 10 ^-3 . Bioburden was measured in samples taken after filling and before terminal sterilization. It was intended to measure a zero (0) CFU/10 mL level and a warning level of >1 CFU/10 mL.

Validation was performed using a dwell time equal to the proposed standard dwell time to demonstrate the ability of the process to achieve an 8 log reduction in spore load (6 log+2 log safety factor). The D-value of the product was determined to be 3.5 minutes for a 20 ml bottle and 4.5 for a 50 ml bottle. To combine both vial sizes in one run, the highest D-value was selected. Assuming that complete destruction of the BI requires a 6 log reduction, the resulting proposed exposure (destruction) time for the validation cycle would be:

Thus, the verification cycle was defined as:

Calculated times that result in a decimal value are rounded to the next minute. Additionally, to maintain the product temperature above 121.1°C for sterilization, the sterilizer set point during exposure was 122.2°C.

The effectiveness of the terminal sterilization process was determined by temperature uniformity and demonstrated a reduction of at least 6 log in the viable number of G. stearothermophilus spores applied at levels ranging from 1 × 10 ⁶ to 5 × 10 ⁶ spores per vial. Based on successful demonstration of biological kill during the validation cycle, exposure time in the production run will have an exposure time of 40 minutes (at a verified exposure temperature of 122.2°C ± 1.0°C) to match the estimated required exposure time for inoculated crop growth product was determined over a D-value time. For a 20 ml vial size, three (3) full load sterilizer cycles were performed consisting of 10 minute, 15 minute and 20 minute exposure times. Once these three (3) experimental run cycles were completed, the optimal run cycle was selected and tested by performing an additional two (2) sterilizer run cycles.

Validation consisted of two parts. Conduct three (3) maximum load sterilizer cycles with temperature measuring devices and biological indicators distributed throughout the chamber, with particular attention to locations identified through empty chamber cycles performed during the annual autoclave requalification (biological indicator locations will be placed in the same place for each cycle). Three (3) minimum load cycles of the sterilizer were performed using one (1) tray located on the top shelf of the chamber (the sterilizer was loaded sequentially from the top shelf of the chamber, so any sterilizer loads with less than the maximum number of trays will always have trays on top shelf of the sterilizer).

A biological indicator (BI) was placed adjacent to each load control/immersion contact probe (LPC/PP). The term "sensor" in the context of this section refers to a device for measuring temperature. All immersion sensors, sterilizer load cells, contact load control sensors and biological indicators were placed in vials containing the product composition; the remainder of the load consisted of bubbles containing an equivalent amount of water. The use of water bottles was acceptable because the product was formulated as an aqueous solution and its thermal properties were essentially identical to pure water.

Load test - minimum and maximum camera load

Purpose : Demonstrate temperature uniformity and biodestruction throughout the entire vial load.

Admission Criteria :

1) All identified biological indicators (BI) should not show an increase.

2) All positive controls must show growth at the end of incubation.

3) All negative controls must have a negative growth test at the end of incubation.

4) All immersion sensors and contact load control sensors must maintain a temperature range of 122.2°C±1.0°C during exposure.

Example 5 Characteristics of Compound 1269

The signal assignments of the ¹ H and ¹³ C NMR spectra for 1269 are given in Table 12 .

Table 12 . Assignment of ¹ H and ¹³ C NMR spectra signals for 1269 (CDCl ₃ )

Structure Position δ _H
(m.d.) Multiplicity ¹
J _H (Hz) Number of protons δ _C
(m.d.) gradient-selective heteronuclear single-quantum correlation (gHSQC)
¹ J Gradient-selective heteronuclear correlation with distant atoms (gHMBC)
^2.3 J 1 1.43
1.27 overlap m
overlap m 1H
1H 32.36 32.36-1.43
32.36-1.27 32.36-0.76: ³ JC₁-H₁₉ 2 1.64 overlap m 2H 29.17 29.17-1.64 - 3 4.02 ear quintet, J =2.4 Hz 1H 66.64 66.64-4.02 - 3a 1.30 w.s. 1H - - - 4 1.50
1.38 overlap m
overlap m 1H
1H 36.08 36.08-1.50
36.08-1.38 - 5 1.50 overlap m 1H 39.16 39.16-1.50 39.16-0.76: ³ JC₅-H₁₉
39.16-1.43: ³ JC₅-H₁ 6 1.16 overlap m 2H 28.72 28.72-1.16 -
- 7 1.68
1.01 overlap m
m 1H
1H 32.41 32.41-1.68
32.41-1.01 1.01-50.55: ³ J H ₇ -C ₁₄ 8 1.30 overlap m 1H 35.91 35.91-1.30 - 9 0.72 ddd, J =12.5, 10.2, 4.1 Hz 1H 53.77 53.77-0.72 53.77-0.76: ³ JC₉-H₁₉
53.77-1.13: ³ JC₉-H₁₂ 10 - - - 36.28 - 36.28-0.76: ² JC₁₀-H₁₉ eleven 1.58
1.28 overlap m
overlap m 1H
1H 20.91 20.91-1.58
20.91-1.28 20.91-1.13: ² J C_eleven-H₁₂
20.91-0.72: ² J C_eleven-H₉ 12 1.73
1.13 overlap m
overlap m 1H
1H 35.57 35.57-1.73
35.57-1.13 35.57-2.78: ³ J C ₁₂ -H ₁₇
35.57-0.90: ³ J C ₁₂ -H ₁₈ 13 - - - 46.01 - 46.01-2.78: ² J C ₁₃ -H ₁₇
46.01-0.90: ² J C ₁₃ -H ₁₈ 14 1.25 overlap m 1H 50.55 50.5-1.25 50.55-0.90: ³ J C ₁₄ -H ₁₈
50.55-2.78: ³ J C ₁₄ -H ₁₇ 15 1.78
1.20 overlap m
overlap m 1H
1H 26.06 26.06-1.78
26.06-1.20 26.06-2.78: ³ J C ₁₅ -H ₁₇ 16 1.90
1.70 overlap m
overlap m 1H
1H 24.47 24.47-1.90
24.47-1.70 24.47-2.78: ² J C ₁₆ -H ₁₇ 17 2.78 dd, J =8.4, 2.5 Hz 1H 61.59 61.59-2.78 61.59-0.90: ³ J C ₁₇ -H ₁₈
61.59-2.12: ³ J C ₁₇ -H ₂₁ 18 0.90 With 3H 21.15 21.15-0.90 21.15-2.78: ³ J C ₁₈ -H ₁₇ 19 0.76 With 3H 11.33 11.33-0.76 11.33-0.72: ³ J C ₁₉ -H ₉ 20 - - - 212.96 - 212.96-2.12: ² JC₂₀-H₂₁
212.96-2.78: ² JC₂₀-H₁₇ 21 2.12 With 3H 32.99 32.99-2.12 - Chemical shifts ¹ H for overlapping resonances (overlapping multiplets) were taken from HSQC data.

LC-MS analysis of 1269 is presented in Figure 6 and Table 13 .

Table 13 . Assignment of mass spectrum signals of compound 1269

Designation Weight [M+H] ⁺ 303.36 [M+HH ₂ O] ⁺ 285.36 [M+H+MeOH] ⁺ 335.41 [2M+H] ⁺ 605.70

Example 6 . Connection Characteristics 136

Table 14 . Assignment of ¹ H and ¹³ C NMR spectra signals for 136 (CDCl ₃ )

Structure Position δ _H
(m.d.) Multiplicity
J _H (Hz) Number of protons δ _C
(m.d.) gradient-selective heteronuclear single-quantum correlation (gHSQC)
¹ J Gradient-selective heteronuclear correlation with distant atoms (gHMBC)
^2.3 J 1 2.03
1.36 overlap m
overlap m 1H
1H 38.73 38.73-2.03
38.73-1.36 38.73-1.02: ³ JC₁-H₁₉ 2 2.39
2.30 m
overlap m 1H
1H 38.32 38.32-2.39
38.32-2.30 - 3 - - - 212.01 - 212.01-2.39: ² JC₃-H₂
212.01-2.27: ² JC₃-H₄ 4 2.27
2.09 t, J =14.2 Hz
overlap m 1H
1H 44.83 44.83-2.27
44.83-2.09 - 5 1.54 m 1H 46.85 46.85-1.55 46.85-1.02: ³ JC₅-H₁₉
46.85-2.27: ³ JC₅-H₄ 6 1.32 overlap m 2H 29.01 29.01-1.32 29.01-1.72: ² JC₆-H₇
29.01-0.94: ² JC₆-H₇
29.01-2.27: ² JC₆-H₄ 7 1.72
0.94 m
m 1H
1H 31.83 31.83-1.72
31.83-0.94 31.83-1.17: ² JC₇-H₁₄ 8 1.43 overlap m 1H 35.55 35.55-1.43 - 9 0.79 m 1H 53.85 53.85-0.79 53.85-1.02: ³ JC₉-H₁₉ 10 - - - 35.87 - 35.87-2.27: ³ JC₁₀-H₄
35.87-1.02: ² JC₁₀-H₁₉ eleven 1.65
1.39 overlap m
overlap m 1H
1H 21.62 21.62-1.65
21.62-1.39 21.62-0.79: ² JC_eleven-H₉
21.62-2.04: ² JC_eleven-H₁₂ 12 2.04
1.44 overlap m
overlap m 1H
1H 39.12 39.12-2.04
39.12-1.44 39.12-2.53: ³ J C ₁₂ -H ₁₇
39.12-0.64: ³ J C ₁₂ -H ₁₈ 13 - - - 44.36 - 44.36-2.53: ² J C ₁₃ -H ₁₇
44.36-0.64: ² J C ₁₃ -H ₁₈ 14 1.17 overlap m 1H 56.64 56.64-1.17 56.64-2.53: ³ J C ₁₄ -H ₁₇
56.64-0.64: ³ J C ₁₄ -H ₁₈ 15 1.69
1.23 overlap m
overlap m 1H
1H 24.60 24.60-1.69
24.60-1.39 24.60-1.17: ² J C ₁₅ -H ₁₄
24.60-1.65: ² J C ₁₅ -H ₁₆ 16 2.17
1.65 m
overlap m 1H
1H 23.02 23.02-2.17
23.02-1.65 23.02-2.53: ² J C ₁₆ -H ₁₇
209.66-2.53: ³ J H ₁₆ -C ₂₀ 17 2.53 t, J =9.0 Hz 1H 63.92 63.92-2.53 63.92-0.64: ³ J C ₁₇ -H ₁₈ 18 0.64 With 3H 13.63 13.63-0.64 13.63-2.53: ³ J C ₁₈ -H ₁₇ 19 1.02 With 3H 11.65 11.65-1.02 11.65-0.79: ³ J C ₁₉ -H ₉ 20 - - - 209.66 - 209.66-2.12: ² JC₂₀-H₂₁
209.66-2.53: ² JC₂₀-H₁₇ 21 2.12 With 3H 31.70 31.70-2.12 - Chemical shifts ¹ H for overlapping resonances (overlap m) were taken from HSQC data.

LC-MS analysis 136 is presented in Figure 7 and Table 15 .

Table 15 . Assignment of mass spectrum signals of compound 136

Designation Weight [M+H] ⁺ 317.33 [M+H+CH ₃ CN] ⁺ 358.49 [2M+H] ⁺ 633.71

Example 6 : pH Stability of Allopregnanolone Formulations in SBECD

A formulation of allopregnanolone (5 mg/mL) in 250 mL/mL sulfobutylether-β-cyclodextrin was prepared at different pH values and packaged in a type I glass vial. The content of the formulation was measured after 12 weeks at 40°C ( Figure 8A ). The content of the main substance of the composition was measured after 12 weeks at 60°C ( Figure 8B ).

Example 7 . Comparison of the effects of different buffers

Figures 3A-B illustrate the purity of the formulation measured after 12 weeks at 40°C in phosphate buffer. Figures 4A-B illustrate the purity of the formulation measured after 12 weeks at 40°C in citrate buffer. Figure 5 illustrates the formation of 136 over time at 40°C in different buffers.

Example 8 . Stability of the formulation with allopregnanolone at low temperatures

The stability of a formulation of allopregnanolone (5 mg/ml) in 250 mg/ml sulfobutylether-β-cyclodextrin in 10 mM citrate buffer pH 6, which was stored at 2-8°C for 12 months, was determined. Stability study data are shown in Table 16.

Table 16. Stability of allopregnanolone formulation stored at 2-8°C for 12 months

Index Original 1 month 3 months 6 months 9 months 12 months Appearance Compliant Compliant Compliant Compliant Compliant Compliant pH 5.8 5.7 5.8 5.8 5.8 5.8 Main substance content (%) 99.5 99.2 99.1 98.2 98.8 97.5 Related substances according to HPLC (wt.%) 136 Not detected Not detected Not detected Not detected Not detected Not detected 1269 <0.10 <0.10 <0.10 0.10 <0.10 <0.10 Mechanical inclusions ≥ 10 µm 76 69 38 214 25 163 ≥ 25 µm 7 0 3 16 1 34

Claims (7)

1. A pharmaceutically acceptable diluted aqueous solution of allopregnanolone, containing (i) a first heat-treated solution containing allopregnanolone, beta-cyclodextrin sulfobutyl ether and citrate buffer, and (ii) a diluent; wherein the dilute aqueous solution is prepared by mixing the first heat-treated solution with a diluent, and where the first solution is autoclaved by subjecting it to heat sterilization cycles for at least 10, 15, 20, 30 or 40 minutes at a temperature of from 110° C. to 150 °C. 2. A dilute aqueous solution according to claim 1, wherein the dilute aqueous solution is homogeneous. 3. A diluted aqueous solution according to claim 1, where the diluent contains sterile water for injection or saline solution. 4. A diluted aqueous solution according to claim 1, where the autoclaving temperature is from 121°C to 123°C. 5. The diluted aqueous solution according to claim 1, wherein the first solution, subjected to heat treatment, contains 5 mg/ml allopregnanolone and 250 mg/ml sulfobutyl ether-β-cyclodextrin. 6. The dilute aqueous solution of claim 1, wherein the sulfobutyl ether-β-cyclodextrin comprises a sulfobutyl ether-β-cyclodextrin salt. 7. The dilute aqueous solution of claim 1, wherein the sulfobutyl ether-β-cyclodextrin comprises the sodium salt of sulfobutyl ether-β-cyclodextrin.

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