Dantrolene prodrugs and methods of use thereof

Abstract

The present disclosure relates to dantrolene prodrugs, compositions thereof, and their use in the treatment of disease.

Description

[0001] Cross-reference to related applications

[0002] This application claims the benefit of U.S. Provisional Application No. 62 / 575,124, filed October 20, 2017, and U.S. Provisional Application No. 62 / 674,422, filed May 21, 2018, the entirety of which is incorporated herein by reference. Technical Field

[0003] This disclosure relates to dantrolene prodrugs, compositions thereof, and methods of using them in the treatment of diseases. Background Technology

[0004] Prodrugs are typically alternative forms of active pharmaceutical ingredients that have been reversibly chemically modified or derivatized by chemical groups that render the prodrug inactive, impart solubility, stability, or bioavailability, or alter other properties of the active pharmaceutical ingredient. Generally, the chemical groups of a prodrug are cleaved from the prodrug by the action of heat, cavitation, pressure, pH changes, reduction-oxidation, and / or enzymatic activity, thereby releasing the active pharmaceutical ingredient. The cleavage of the chemical groups of the prodrug can occur before drug delivery to the subject, but generally takes place in vivo via an enzymatic process.

[0005] Dantrolin (1-{[5-(4-nitrophenyl)-2-furanyl]methyleneamino}imidazolidine-2,4-dione) has the structure of formula (1):

[0006]

[0007] Dantrolene is the first-line treatment for malignant hyperthermia (“MH”) and is widely used in most cases of anesthetic administration. First synthesized in 1967, dantrolene was initially used to treat muscle rigidity in 1975 and later approved by the FDA for the treatment of MH in 1979. Dantrolene is considered a potent muscle relaxant and a treatment for neurogenic rigidity. Since its initial discovery, dantrolene has been studied for the prevention and treatment of other life-threatening conditions, such as overdose of recreational drugs like ecstasy (N-methyl-3,4-methylenedioxyphenylisopropylamine), heatstroke, malignant syndrome caused by antipsychotic drugs, and ischemic damage to the peripheral nervous system. It may also have significant implications for the prevention of sudden infant death syndrome (SIDS).

[0008] Dantril is poorly soluble in water. This poor solubility significantly impairs its application. For example, DANTRIUM TMDantrolene sodium is provided in 20 mg vials and must be reconstituted with 60 mL of sterile water before intravenous administration. The recommended dose of dantrolene for treating hemorrhagic muscular dystrophy (MH) is 1 mg / kg to approximately 10 mg / kg. Therefore, a subject weighing 80 kg would require a rapid infusion of up to 2400 mL to treat MH.

[0009] In addition to poor solubility, dantrialin solutions also have a high pH. TM The pH is approximately 9.5. An improved dantrolene sodium formulation, which can be reconstituted to 50 mg / mL, significantly improves the rate of dantrolene administration. However, the reconstituted... It also has a high pH of approximately 10.3. Due to this high pH, ​​current dantrolene formulations cannot be administered subcutaneously or intramuscularly—they can only be administered intravenously. In fact, care must be taken to prevent extravasation into surrounding tissues to avoid tissue necrosis.

[0010] While dantrialeline prodrugs may help address the drug's solubility and pH challenges, identifying a suitable prodrug moiety becomes complex due to several factors inherent in the dantrialeline molecule. For example, the poor solubility of dantrialeline is presumably attributed to its extended aromatic system, which may participate in hydrophobic π-stacking behavior. Even the charged nitro moiety of dantrialeline does not improve the compound's solubility in water.

[0011] Dantrolene includes a hydantoin moiety, which is found in other drug compounds such as phenytoin. However, while prodrug strategies have been reported to improve the water solubility of other hydantoin-containing compounds, it is unclear whether a similar strategy could be successfully applied to dantrolene given its unique chemical structure and physical properties.

[0012] There is a need for new dantrolene formulations with appropriate concentrations and pH levels to make them suitable for intramuscular or subcutaneous use, as well as oral, transmucosal (e.g., intranasal) and intraosseous administration. Summary of the Invention

[0013] This disclosure relates to compounds of formula I and pharmaceutically acceptable salts thereof.

[0014]

[0015] Where R is -P(O)(OH)2 or -P(O)(OR1)(OR2); R1 is H, -C 1-26 Alkyl, aryl, C 1-6 Alkane C(O)OC 1-26 Alkyl, -C1 alkyl OC(O)C 1-26 Alkyl or C1 alkyl OC(O)OC 1-26 Alkyl; and R2 is -C 1-26 Alkyl, aryl, C1-6 Alkane C(O)OC 1-26 Alkyl, -C1 alkyl OC(O)C 1-26 Alkyl or C1 alkyl OC(O)OC 1-26 Alkyl groups. Pharmaceutical compositions comprising compounds of formula I are also described, as well as methods of using them.

[0016] This disclosure also relates to compounds of formula II and their pharmaceutically acceptable salts.

[0017]

[0018] Where R3 is H, -C(O)-ZN(R4)(R5), -C(O)ZC(O)-OH, or -C(O)-NH-Y-CH2-OC(O)-ZC(O)-OH; Z is C 1-6 Alkane; Y is arylene; C 1-6 Alkyl; R5 is H or C 1-6 Alkyl groups; or R4 and R5, together with the nitrogen to which they are attached, to form heterocyclic alkyl groups. Pharmaceutical compositions comprising compounds of formula II are also described, as well as methods of using them. Attached Figure Description

[0019] Figure 1 The peak area over time was depicted for the conversion of the prodrug of this disclosure to dantrolene via alkaline phosphatase at 25°C.

[0020] Figure 2 The peak area over time was depicted for the conversion of the prodrug (2a) of this disclosure to dantrolene via alkaline phosphatase at 25°C.

[0021] Figure 3 The peak area over time for the conversion of the prodrug of this disclosure to dantrolene via rat plasma at 22°C is depicted.

[0022] Figure 4 The peak area over time for the conversion of the prodrug of this disclosure to dantrolene via rat plasma at 37°C is depicted.

[0023] Figure 5 The conversion of compound 2a to dantrolene in rat plasma was depicted. Rat plasma was incubated with 100 μg / mL of compound 2a at 37 °C. The areas under the prodrug peak and the dantrolene peak in the 385 nm chromatogram were plotted against reaction time. Circles represent the prodrug, and triangles represent dantrolene.

[0024] Figure 6 The mean concentration of dantrolene in rat plasma from animals administered 7.5 mg / kg prodrug 2a was depicted (n = 5 ± SEM (standard error of mean)). Quantification was performed by absorbance at 385 nm.

[0025] Figure 7 The mean concentration of dantrolene in whole blood of rats (n=3) from animals administered 7.5 mg / kg prodrug 2a was depicted. Quantification was performed by absorbance at 385 nm.

[0026] Figure 8 The mean concentration of dantrolene in rat plasma (n = 5 ± SEM) from animals administered 7.5 mg / kg prodrug 2a was depicted. Quantification was performed by absorbance at 385 nm.

[0027] Figure 9 The mean concentration of dantrolene in whole blood of rats administered with 7.5 mg / kg prodrug 2a was depicted (n = 5 ± SEM). Quantification was performed by absorbance at 385 nm.

[0028] Figure 10 The conversion of compound 2b to dantrolene in rat plasma was depicted. Rat plasma was incubated with 100 μg / mL of compound 2b at 37 °C. The areas under the prodrug peak and the dantrolene peak in the 385 nm chromatogram were plotted against reaction time. Circles represent the prodrug, and triangles represent dantrolene.

[0029] Figure 11 The mean concentration of dantrolene in rat plasma (n = 5 ± SEM) from animals administered 2b at 10.6 mg / kg was depicted. Quantification was performed by absorbance at 385 nm.

[0030] Figure 12 The conversion of 10c to dantrolene in rat plasma was depicted. Rat plasma was incubated with 100 μg / mL of compound 10c at 37 °C. The areas under the prodrug peak and the dantrolene peak in the 385 nm chromatogram were plotted against reaction time. Circles represent the prodrug, and triangles represent dantrolene.

[0031] Figure 13 The conversion of 12a to dantrolene in rat plasma was depicted. Rat plasma was incubated with 100 μg / mL of compound 12a at 37 °C. The areas under the prodrug peak and the dantrolene peak in the 385 nm chromatogram were plotted against reaction time. Circles represent the prodrug, and triangles represent dantrolene.

[0032] Figure 14 The mean concentrations of dantrolene in whole blood from animals administered 4 mg / kg prodrug 12a were depicted (n = 3 ± SEM). Quantification was performed by absorbance at 385 nm.

[0033] Figure 15The conversion of 17b to dantrolene in rat plasma was depicted. Rat plasma was incubated with 100 μg / mL 17b at 37°C. The areas under the prodrug peak and the dantrolene peak in the 385 nm chromatogram were plotted against reaction time. Circles represent the prodrug, and triangles represent dantrolene.

[0034] Figure 16 The conversion of 22c to dantrolene in rat plasma was depicted. Rat plasma was incubated with 100 μg / mL 22c at 37 °C. The areas under the prodrug peak and the dantrolene peak in the 385 nm chromatogram were plotted against reaction time. Circles represent the prodrug, and triangles represent dantrolene.

[0035] Figure 17 The mean concentrations of dantrolene in whole blood from animals administered 4 mg / kg prodrug 22c were depicted (n = 3 ± SEM). Quantification was performed by absorbance at 385 nm. Invention Details

[0037] This disclosure will be more readily understood by referring to the following detailed description in conjunction with the accompanying drawings and embodiments that form a part of this disclosure. It should be understood that this disclosure is not limited to the specific components, apparatus, methods, applications, conditions, or parameters described and / or shown herein, and that the terminology used herein is for illustrative purposes only and is not intended to limit the scope of the claimed disclosure.

[0038] The use of references without a specified quantity in the description, including the claims, includes the plurality of the references, and references to a particular value include at least that particular value, unless the context clearly requires otherwise.

[0039] When expressing a range of values, exemplary implementations include from one specific value and / or to another specific value. All ranges are inclusive and composable. Furthermore, references to values ​​stated as ranges include all and every value within that range. When a value is expressed as an approximation using the preposition “about,” it should be understood that the specific value forms another implementation. As used herein, the term “about,” when referring to a measurable value such as quantity, duration, etc., means including reasonable variation of that value, for example, from ±10% of the specified value. For example, the phrase “about 50%” could include 50 ± 10%, or from 45% to 55%, including 50%.

[0040] It should be understood that, for clarity, certain features of this disclosure described herein in the context of separate embodiments may also be provided in combination in a single embodiment. Conversely, for simplicity, various features of this disclosure described in the context of a single embodiment may also be provided separately or in any sub-combination.

[0041] As used in this article, whether alone or in combination with other terms, it should be understood that the phrase “treatment method” can be used interchangeably with the phrase “for the treatment of” a specific disease.

[0042] As used herein, whether alone or in combination with other terms, “pharmaceutical acceptable” indicates that the indicated entity, such as a pharmaceutically acceptable excipient, is substantially chemically and / or physically compatible with the other components in the composition, and / or substantially physiologically compatible with its recipient.

[0043] As used herein, a “pharmaceutical composition” means a composition prepared by combining any formulation, including the suspensions or dispersions described herein, with one or more pharmaceutically acceptable excipients.

[0044] "Pharmaceutical-acceptable excipients" means diluents, adjuvants, excipients, or carriers that are administered with the compounds disclosed herein. "Pharmaceutical-acceptable excipients" means nontoxic, biologically tolerable, and otherwise biologically suitable for administration to a target, such as inert substances, which are added to or otherwise used as a medium, carrier, or diluent to facilitate and are compatible with the administration of the pharmaceutical preparation. Examples of excipients are cited, for example, in Remington's Pharmaceutical Sciences, 17th edition, Mack Publishing Co. (1985).

[0045] As used herein, whether alone or in combination with other terms, “object,” “individual,” and “patient” refer to mammals, including humans. The term “human” refers to and includes human children, adolescents, or adults.

[0046] As used herein, whether alone or in combination with other terms, "treatment" means and includes improving, alleviating, and / or curative use and results, or any combination thereof. In other embodiments, the methods described herein may be used preventively. It should be understood that "prevention" or preventative use or results does not involve or require absolute or complete prevention (i.e., 100% preventative or protective use or results). As used herein, prevention or preventative use or results means that the application of a compound or composition reduces or lessens the severity of the particular condition, symptom, symptom, or disease described herein; reduces or lessens the likelihood of experiencing the particular condition, symptom, symptom, or disease described herein; or delays the onset or recurrence (recurrence) of the particular condition, symptom, symptom, or disease described herein; or any combination thereof.

[0047] As used herein, whether alone or in combination with other terms, "therapeutic" and "therapeuticly effective amount" refer to an amount of a compound or composition that (a) treats the specific condition, symptom, ailment, or disease described herein; (b) alleviates, improves, or eliminates one or more symptoms of the specific condition, ailment, or disease described herein; or (c) delays the onset or recurrence (recurrence) of the specific condition, symptom, ailment, or disease described herein. It should be understood that the terms "therapeutic" and "therapeuticly effective" include any of the foregoing effects (a)-(c), alone or in combination with any of the other (a)-(c).

[0048] The term "C1-C6 alkane" refers to an aliphatic linker having 1, 2, 3, 4, 5, or 6 carbon atoms, and includes, for example, -CH2-, -CH(CH3)-, -CH(CH3)-CH2-, and -C(CH3)2-. The term "-C0 alkane-" refers to a bond.

[0049] The term "alkyl" refers to a group having 1 to 12 carbon atoms ("C1-C12"). 12 The alkyl group is preferably a straight-chain or branched hydrocarbon group with 1 to 6 carbon atoms (“C1-C6”). Examples of alkyl groups include methyl (Me, C1 alkyl), ethyl (Et, C2 alkyl), n-propyl (C3 alkyl), isopropyl (C3 alkyl), butyl (C4 alkyl), isobutyl (C4 alkyl), sec-butyl (C4 alkyl), tert-butyl (C4 alkyl), pentyl (C5 alkyl), isopentyl (C5 alkyl), tert-pentyl (C5 alkyl), hexyl (C6 alkyl), isohexyl (C6 alkyl), etc.

[0050] The term "heterocyclic alkyl" refers to any three- to ten-membered monocyclic or bicyclic saturated ring structure containing at least one heteroatom selected from O, N, and S. Examples of suitable heterocyclic alkyl groups include, but are not limited to, aziridine, aziridine, pyrrolidinyl, piperazine, piperidinyl, morpholinyl, thiomorpholinyl, etc.

[0051] The term "aryl," when used alone or as part of a substituent group, refers to a monocyclic or bicyclic aromatic hydrocarbon ring structure having 6 or 10 carbon atoms in the ring. Preferred aryl moieties include phenyl and naphthyl groups.

[0052] The term "arylene" refers to a monocyclic or bicyclic aromatic hydrocarbon ring structure having 6 or 10 carbon atoms in the ring. Preferred arylene moieties include phenylene and naphthylene. The compounds disclosed herein can be chiral and therefore can exist as a single enantiomer or a mixture of enantiomers. This disclosure contemplates all enantiomers and mixtures thereof.

[0053] Isotopic variants of the compounds of Formulas I and II are also within the scope of this disclosure. As used herein, the term "isotopic variant" refers to a compound that contains a higher proportion of isotopes than naturally occurring on one or more of the atoms constituting such a compound. For example, an "isotopic variant" of a compound may be radiolabeled, i.e., containing one or more radioactive isotopes, or may be made of non-radioactive isotopes such as deuterium (…). 2 H or D), carbon-11 ( 11 C), Carbon-13 ( 13 C), Nitrogen-15 ( 15 N), Fluorine-18 ( 18 F) and other markings. It should be understood that in compounds that have undergone such isotopic substitution, the following atoms present may be different, such that, for example, any hydrogen may be... 2 H / D, any carbon may be 11 C or 13 C, any nitrogen may be 15 N, or any fluorine (if present), may be 18 F, and the presence and location of such atoms can be determined within the scope of the art.

[0054] Compounds of Formula I and II are converted to dantrolene in vivo. In some aspects, compounds of Formula I and II are converted to dantrolene in vivo with a half-life of about 1 second or less to about 1 minute to 90 minutes. In some aspects, compounds of Formula I and II are converted to dantrolene in vivo with a half-life of less than 1 second. In other respects, the compounds of Formulas I and II are converted in vivo to dantrolene with a half-life of several seconds, i.e., a half-life of less than one minute, for example, a half-life of about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, or about 59 seconds. In other aspects, the compounds of formulas I and II are converted to dantrolene in vivo with a half-life of about 1 to about 5 minutes, for example, about 1, 2, 3, 4, or about 5 minutes. In other aspects, the compounds of formulas I and II are converted to dantrolene in vivo with a half-life of about 1 to about 10 minutes. In other aspects, the compounds of formulas I and II are converted to dantrolene in vivo with a half-life of about 5 to about 10 minutes. In some aspects, the compounds of formulas I and II are converted to dantrolene in vivo with a half-life of about 1 minute to about 60 minutes. In some aspects, the compounds of formulas I and II are converted to dantrolene in vivo with a half-life of about 1 minute to about 45 minutes. In some aspects, the compounds of formulas I and II are converted to dantrolene in vivo with a half-life of about 1 minute to about 30 minutes. In some aspects, the compounds of formulas I and II are converted to dantrolene in vivo with a half-life of about 1 minute to about 20 minutes. In some respects, the compounds of formulas I and II are converted in vivo to dantrolene with a half-life of about 1, 2, 3, 4, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, or 90 minutes.

[0055] This disclosure relates to dantrolene prodrug of Formula I or a pharmaceutically acceptable salt thereof:

[0056]

[0057] Where R is -P(O)(OH)2 or -P(O)(OR1)(OR2);

[0058] R1 is H, -C 1-26 Alkyl, aryl, C 1-6 Alkane C(O)OC 1-26 Alkyl, -C1 alkyl OC(O)C 1-26 Alkyl or C1 alkyl OC(O)OC1-26 Alkyl; and

[0059] R2 is -C 1-26 Alkyl, aryl, C 1-6 Alkane C(O)OC 1-26 Alkyl, -C1 alkyl OC(O)C 1-26 Alkyl or C1 alkyl OC(O)OC 1-26 alkyl.

[0060] In some respects, the dantrolene prodrug of this disclosure is one in which R is -P(O)(OH)2 and has the formula IA:

[0061]

[0062] Pharmaceutically acceptable salts of compounds of formula IA are also within the scope of this disclosure. Preferred salts include, for example, sodium salts of compounds of formula IA. Lithium, magnesium, calcium, and potassium salts of said compounds of formula IA are also within the scope of this disclosure. Alternative salt forms include ammonium salts, choline salts, and tromethamine salts. One preferred salt of said compound IA is a monosodium salt. Another preferred salt of said compound IA is a disodium salt. Another preferred salt of said compound IA is a monoaminobutyrate salt. Another preferred salt of said compound IA is a diaminobutyrate salt. Pharmaceutically acceptable organic salts of compounds of formula IA are also within the scope of this disclosure.

[0063] In some respects, the dantrolene prodrugs disclosed herein are those in which R is -P(O)(OR1)(OR2) and has the formula IB:

[0064]

[0065] In some respects, R1 is H. In other respects, R2 is -C. 1-26 Alkyl, aryl, C 1-6 Alkane C(O)OC 1-26 Alkyl, -C1 alkyl OC(O)C 1-26 Alkyl or C1 alkyl OC(O)OC 1-26 Alkyl groups. Pharmaceutically acceptable salts of such formula IB compounds are also within the scope of this disclosure. Preferred salts include, for example, sodium salts of formula IB compounds. Other salts include lithium, magnesium, calcium, and potassium salts of said formula IB compounds. Alternative salt forms include ammonium salts, choline salts, and tromethamine salts. Pharmaceutically acceptable organic salts of formula IB compounds are also within the scope of this disclosure.

[0066] In some aspects of compounds of formula IB, R1 is H and R2 is -C. 1-26 Alkyl group. For example, in some respects, R1 is H and R2 is -C. 1-6Alkyl group. In other respects, R1 is H and R2 is -C. 1-12 Alkyl group. In other respects, R1 is H and R2 is -C. 13-26 Alkyl group. In other respects, R1 is H and R2 is -C. 18-26 Alkyl group. In other respects, R1 is H and R2 is -C. 20-26 Alkyl. In some respects, R1 is H and R2 is a -C1 alkyl. In some respects, R1 is H and R2 is a -C2 alkyl. In some respects, R1 is H and R2 is a -C3 alkyl. In some respects, R1 is H and R2 is a -C4 alkyl. In some respects, R1 is H and R2 is a -C5 alkyl. In some respects, R1 is H and R2 is a -C6 alkyl. In some respects, R1 is H and R2 is a -C7 alkyl. In some respects, R1 is H and R2 is a -C8 alkyl. In some respects, R1 is H and R2 is a -C9 alkyl. In some respects, R1 is H and R2 is a -C 10 Alkyl group. In some respects, R1 is H and R2 is -C. 11 Alkyl group. In some respects, R1 is H and R2 is -C. 12 Alkyl group. In some respects, R1 is H and R2 is -C. 13 Alkyl group. In some respects, R1 is H and R2 is -C. 14 Alkyl group. In some respects, R1 is H and R2 is -C. 15 Alkyl group. In some respects, R1 is H and R2 is -C. 16 Alkyl group. In some respects, R1 is H and R2 is -C. 17 Alkyl group. In some respects, R1 is H and R2 is -C. 18 Alkyl group. In some respects, R1 is H and R2 is -C. 19 Alkyl group. In some respects, R1 is H and R2 is -C. 20 Alkyl group. In some respects, R1 is H and R2 is -C. 21 Alkyl group. In some respects, R1 is H and R2 is -C. 22 Alkyl group. In some respects, R1 is H and R2 is -C. 23 Alkyl group. In some respects, R1 is H and R2 is -C. 24 Alkyl group. In some respects, R1 is H and R2 is -C. 25 Alkyl group. In some respects, R1 is H and R2 is C. 26 alkyl.

[0067] In some aspects of compounds of formula IB, R1 is H and R2 is aryl. For example, in some aspects of compounds of formula IB, R1 is H and R2 is phenyl.

[0068] In some aspects of compounds of formula IB, R1 is H and R2 is C. 1-6 Alkane C(O)OC 1-26 Alkyl. For example, in some respects, R1 is H and R2 is a C1 alkane C(O)OC. 1-26 Alkyl group. In other respects, R1 is H and R2 is a C2 alkane (C(O)OC). 1-26 Alkyl group. In other respects, R1 is H and R2 is a C3 alkane (C(O)OC). 1-26 Alkyl group. In other respects, R1 is H and R2 is a C4 alkane (C(O)OC). 1-26 Alkyl group. In other respects, R1 is H and R2 is a C5 alkane (C(O)OC). 1-26 Alkyl group. In other respects, R1 is H and R2 is a C6 alkane (C(O)OC). 1-26 Alkyl group. In other respects, R1 is H and R2 is C. 1-6 Alkane C(O)OC 1-6 Alkyl group. In other respects, R1 is H and R2 is C. 1-6 Alkane C(O)OC 1-12 Alkyl group. In other respects, R1 is H and R2 is C. 1-6 Alkane C(O)OC 13-26 Alkyl group. In other respects, R1 is H and R2 is C. 1-6 Alkane C(O)OC 18-26 Alkyl group. In other respects, R1 is H and R2 is C. 1-6 Alkane C(O)OC 20-26 alkyl.

[0069] In some aspects of compounds of formula IB, R1 is H and R2 is a -C1 alkane OC(O)C 1-26 Alkyl group. For example, in some respects, R1 is H and R2 is a -C1 alkane OC(O)C. 1-6 Alkyl group. In other respects, R1 is H and R2 is a -C1 alkane (OC(O)C). 1-12 Alkyl group. R1 is H and R2 is a -C1 alkane OC(O)C 13-16 Alkyl group. R1 is H and R2 is a -C1 alkane OC(O)C 18-26 Alkyl group. R1 is H and R2 is a -C1 alkane OC(O)C 20-26 alkyl.

[0070] In some aspects of compounds of formula IB, R1 is H and R2 is a -C1 alkane OC(O)OC 1-26 Alkyl. For example, in some respects, R1 is H and R2 is a -C1 alkane. 1-6 Alkyl group. In other respects, R1 is H and R2 is a -C1 alkane. 1-12Alkyl group. In some respects, R1 is H and R2 is a -C1 alkane. 13-16 Alkyl group. In some respects, R1 is H and R2 is a -C1 alkane. 18-26 Alkyl group. In some respects, R1 is H and R2 is a -C1 alkane. 20-26 alkyl.

[0071] In other respects of compounds of formula IB, R1 is -C 1-26 Alkyl, aryl, C 1-6 Alkane C(O)OC 1-26 Alkyl, -C1 alkyl OC(O)C 1-26 Alkyl or C1 alkyl OC(O)OC 1-26 Alkyl and R2 is -C 1-26 Alkyl, aryl, C 1-6 Alkane C(O)OC 1-26 Alkyl, -C1 alkyl OC(O)C 1-26 Alkyl or C1 alkyl OC(O)OC 1-26 alkyl.

[0072] In some aspects of compounds of formula IB, R1 is -C 1-26 Alkyl and R2 is -C 1-26 Alkyl, aryl, C 1-6 Alkane C(O)OC 1-26 Alkyl, -C1 alkyl OC(O)C 1-26 Alkyl or C1 alkyl OC(O)OC 1-26 Alkyl group. For example, in these respects, R1 can be -C 1-6 Alkyl group. In other respects, R1 is -C. 1-12 Alkyl group. In other respects, R1 is -C. 13-26 Alkyl group. In other respects, R1 is -C. 18-26 Alkyl group. In other respects, R1 is -C. 20-26 Alkyl group. In some aspects, R1 is a -C1 alkyl group. In some aspects, R1 is a -C2 alkyl group. In some aspects, R1 is a -C3 alkyl group. In some aspects, R1 is a -C4 alkyl group. In some aspects, R1 is a -C5 alkyl group. In some aspects, R1 is a -C6 alkyl group. In some aspects, R1 is a -C7 alkyl group. In some aspects, R1 is a -C8 alkyl group. In some aspects, R1 is a -C9 alkyl group. In some aspects, R1 is a -C 10 Alkyl group. In some respects, R1 is -C 11 Alkyl group. In some respects, R1 is -C 12 Alkyl group. In some respects, R1 is -C 13 Alkyl group. In some respects, R1 is -C 14 Alkyl group. In some respects, R1 is -C15 Alkyl group. In some respects, R1 is -C 16 Alkyl group. In some respects, R1 is -C 17 Alkyl group. In some respects, R1 is -C 18 Alkyl group. In some respects, R1 is -C 19 Alkyl group. In some respects, R1 is -C 20 Alkyl group. In some respects, R1 is -C 21 Alkyl group. In some respects, R1 is -C 22 Alkyl group. In some respects, R1 is -C 23 Alkyl group. In some respects, R1 is -C 24 Alkyl group. In some respects, R1 is -C 25 Alkyl group. In some respects, R1 is -C 26 alkyl.

[0073] In some aspects of compounds of formula IB, R1 is aryl and R2 is -C. 1-26 Alkyl, aryl, C 1-6 Alkane C(O)OC 1-26 Alkyl, -C1 alkyl OC(O)C 1-26 Alkyl or C1 alkyl OC(O)OC 1-26 Alkyl group. For example, in some respects, R1 is phenyl and R2 is -C. 1-26 Alkyl, aryl, C 1-6 Alkane C(O)OC 1-26 Alkyl, -C1 alkyl OC(O)C 1-26 Alkyl or C1 alkyl OC(O)OC 1-26 alkyl.

[0074] In some aspects of compounds of formula IB, R1 is C 1-6 Alkane C(O)OC 1-26 Alkyl and R2 is -C 1-26 Alkyl, aryl, C 1-6 Alkane C(O)OC 1-26 Alkyl, -C1 alkyl OC(O)C 1-26 Alkyl or C1 alkyl OC(O)OC 1-26 Alkyl. For example, in some respects, R1 is a C1 alkane C(O)OC. 1-26 Alkyl group. In other respects, R1 is a C2 alkane (C(O)OC). 1-26 Alkyl group. In other respects, R1 is a C3 alkane (C(O)OC). 1-26 Alkyl group. In other respects, R1 is a C4 alkane (C(O)OC). 1-26 Alkyl group. In other respects, R1 is a C5 alkane (C(O)OC). 1-26 Alkyl group. In other respects, R1 is a C6 alkane (C(O)OC).1-26 Alkyl group. In other respects, R1 is C1. 1-6 Alkane C(O)OC 1-6 Alkyl group. In other respects, R1 is C1. 1-6 Alkane C(O)OC 1-12 Alkyl group. In other respects, R1 is C1. 1-6 Alkane C(O)OC 13-26 Alkyl group. In other respects, R1 is C1. 1-6 Alkane C(O)OC 18-26 Alkyl group. In other respects, R1 is C1. 1-6 Alkane C(O)OC 20-26 alkyl.

[0075] In some aspects of compounds of formula IB, R1 is a -C1 alkane OC(O)C 1-26 Alkyl and R2 is -C 1-26 Alkyl, aryl, C 1-6 Alkane C(O)OC 1-26 Alkyl, -C1 alkyl OC(O)C 1-26 Alkyl or C1 alkyl OC(O)OC 1-26 Alkyl group. For example, in some respects, R1 is a C1 alkane OC(O)C. 1-6 Alkyl group. In other respects, R1 is a -C1 alkane OC(O)C. 1-12 Alkyl group. In other respects, R1 is a -C1 alkane OC(O)C. 13-16 Alkyl group. In other respects, R1 is a -C1 alkane OC(O)C. 18-26 Alkyl group. In other respects, R1 is a -C1 alkane OC(O)C. 20-26 alkyl.

[0076] In some aspects of compounds of formula IB, R1 is a -C1 alkane OC(O)OC. 1-26 Alkyl and R2 is -C 1-26 Alkyl, aryl, C 1-6 Alkane C(O)OC 1-26 Alkyl, -C1 alkyl OC(O)C 1-26 Alkyl or C1 alkyl OC(O)OC 1-26 Alkyl. For example, in some respects, R1 is a C1 alkane OC(O)OC. 1-6 Alkyl group. In other respects, R1 is a -C1 alkane (OC(O)OC). 1-12 Alkyl group. In other respects, R1 is a -C1 alkane (OC(O)OC). 13-16 Alkyl group. In other respects, R1 is a -C1 alkane (OC(O)OC). 18-26 Alkyl group. In other respects, R1 is a -C1 alkane (OC(O)OC). 20-26 alkyl.

[0077] In some respects, R1 is -C 1-26 Alkyl and R2 is -C 1-26 Alkyl group. For example, in some respects, R1 and R2 are each independently -C. 1-6 Alkyl, -C 1-12 Alkyl, -C 13-26 Alkyl, -C 18-26 Alkyl, -C 20-26 Alkyl, -C1 alkyl, -C2 alkyl, -C3 alkyl, -C4 alkyl, -C5 alkyl, -C6 alkyl, -C7 alkyl, -C8 alkyl, -C9 alkyl, -C 10 Alkyl, -C 11 Alkyl, -C 12 Alkyl, -C 13 Alkyl, -C 14 Alkyl, -C 15 Alkyl, -C 16 Alkyl, -C 17 Alkyl, -C 18 Alkyl, -C 19 Alkyl, -C 20 Alkyl, -C 21 Alkyl, -C 22 Alkyl, -C 23 Alkyl, -C 24 Alkyl, -C 25 Alkyl, or -C 26 alkyl.

[0078] In some respects, R1 is aryl (e.g., phenyl) and R2 is aryl (e.g., phenyl).

[0079] In some respects, R1 is C 1-6 Alkane C(O)OC 1-26 Alkyl and R2 is C 1-6 Alkane C(O)OC 1-26 Alkyl group. For example, in some respects, R1 and R2 are each independently a C1 alkane (C(O)OC). 1-26 Alkyl, C2 alkyl C(O)OC 1-26 Alkyl, C3 alkyl C(O)OC 1-26 Alkyl, C4 alkyl C(O)OC 1-26 Alkyl, C5 alkyl C(O)OC 1-26 Alkyl, C6 alkyl C(O)OC 1-26 Alkyl, C 1-6 Alkane C(O)OC 1-6 Alkyl, C 1-6 Alkane C(O)OC 1-12 Alkyl, C 1-6 Alkane C(O)OC 13-26 Alkyl, C 1-6Alkane C(O)OC 18-26 Alkyl, or C 1-6 Alkane C(O)OC 20-26 alkyl.

[0080] In some respects, R1 is a -C1 alkane OC(O)C 1-26 Alkyl and R2 is a -C1 alkane OC(O)C 1-26 Alkyl group. For example, in some respects, R1 and R2 are each independently a C1 alkane OC(O)C. 1-6 Alkyl, -C1 alkyl OC(O)C 1-12 Alkyl, -C1 alkyl OC(O)C 13-16 Alkyl, -C1 alkyl OC(O)C 18-26 Alkyl or -C1 alkyl OC(O)C 20-26 alkyl.

[0081] In some respects, R1 is a -C1 alkane OC(O)OC 1-26 Alkyl and R2 is a -C1 alkane OC(O)OC 1-26 Alkyl group. For example, in some respects, R1 and R2 are each independently a C1 alkane. 1-6 Alkyl, -C1 alkyl OC(O)OC 1-12 Alkyl, -C1 alkyl OC(O)OC 13-16 Alkyl, -C1 alkyl OC(O)OC 18-26 Alkyl or -C1 alkyl OC(O)OC 20-26 alkyl.

[0082] Compounds of Formula I, including those of Formulas IA and IB, may exist as pharmaceutically acceptable salts where applicable. These salts include sodium salts. Potassium, lithium, calcium, and magnesium salts are also contemplated. Alternative salt forms include ammonium salts, choline salts, and tromethamine salts.

[0083] Also within the scope of this disclosure are dantrolene prodrug of formula II or a pharmaceutically acceptable salt thereof:

[0084]

[0085] in

[0086] R3 is H, -C(O)-ZN(R4)(R5), -C(O)ZC(O)-OH, or -C(O)-Y-CH2-OC(O)-ZC(O)-OH;

[0087] Z is C 1-6 alkyl;

[0088] Y is aryl;

[0089] R4 is H or C. 1-6alkyl;

[0090] R5 is H or C 1-6 alkyl;

[0091] Alternatively, R4 and R5, together with the nitrogen they are attached to, form heterocyclic alkyl groups.

[0092] In a preferred aspect, R3 is H and the compound of formula II is a compound of formula II-A or a pharmaceutically acceptable salt thereof:

[0093]

[0094] In other aspects of Formula II, R3 is C(O)-ZN(R4)(R5) and the compound of Formula II is a compound of Formula II-B or a pharmaceutically acceptable salt thereof:

[0095]

[0096] in

[0097] Z is C 1-6 alkyl;

[0098] R4 is H or C. 1-6 alkyl;

[0099] R5 is H or C 1-6 alkyl;

[0100] Alternatively, R4 and R5, together with the nitrogen they are attached to, form heterocyclic alkyl groups.

[0101] In these aspects of formula II-B, Z can be a C1 alkane, C2 alkane, C3 alkane, C4 alkane, C5 alkane, or C6 alkane. In some aspects, Z is C 1-2 Alkane. In some respects, Z is a C1 alkane.

[0102] In these respects of Equation II-B, R4 is H. In other respects, R4 is C. 1-6 Alkyl, such as C1 alkyl, C2 alkyl, C3 alkyl, C4 alkyl, C5 alkyl, or C6 alkyl. In a preferred aspect, R4 is methyl, ethyl, or isopropyl.

[0103] In these respects of Equation II-B, R5 is H. In other respects, R5 is C. 1-6 Alkyl, such as C1 alkyl, C2 alkyl, C3 alkyl, C4 alkyl, C5 alkyl, or C6 alkyl. In a preferred aspect, R5 is methyl, ethyl, or isopropyl.

[0104] In some of these aspects of Equation II-B, R4 is H and R5 is H. In other aspects, R4 is H and R5 is C. 1-6Alkyl groups, such as C1 alkyl, C2 alkyl, C3 alkyl, C4 alkyl, C5 alkyl, or C6 alkyl. In other respects, R4 and R5 are each independently C... 1-6 Alkyl groups, such as C1 alkyl, C2 alkyl, C3 alkyl, C4 alkyl, C5 alkyl, or C6 alkyl.

[0105] In some of these aspects of formula II-B, R4 and R5, together with the nitrogen to which they are attached, form a heterocyclic alkyl group. Preferred heterocyclic alkyl moieties include, for example, morpholino, piperazine, piperidinyl, pyrrolidinyl, azacyclic butyl, and azapropylidinyl.

[0106] Preferred compounds of formula II-B include, for example,

[0107]

[0108]

[0109] And its pharmaceutically acceptable salts.

[0110] In other respects of Formula II, R3 is C(O)-ZC(O)-OH and the compound of Formula II is a compound of Formula II-C or a pharmaceutically acceptable salt thereof.

[0111]

[0112] in

[0113] Z is C 1-6 alkyl.

[0114] In these aspects of formula II-C, Z can be a C1 alkane, C2 alkane, C3 alkane, C4 alkane, C5 alkane, or C6 alkane. In some aspects, Z is C 1-2 Alkane. In some respects, Z is a C1 alkane. In other respects, Z is a C2 alkane.

[0115] Preferred compounds of formula II-C are

[0116]

[0117] And its pharmaceutically acceptable salts.

[0118] In other respects of Formula II, R3 is -C(O)-NH-Y-CH2-OC(O)-ZC(O)-OH and the compound of Formula II is a compound of Formula II-D or a pharmaceutically acceptable salt thereof.

[0119]

[0120] in

[0121] Y is a aryl group; and

[0122] Z is C 1-6 alkyl.

[0123] In these aspects of formula II-D, Y can be phenylene or naphthylene, preferably phenylene.

[0124] In these aspects of formulas II-D, Z can be a C1 alkane, C2 alkane, C3 alkane, C4 alkane, C5 alkane, or C6 alkane. In some aspects, Z is C 1-2 Alkane. In some respects, Z is a C1 alkane. In other respects, Z is a C2 alkane.

[0125] The preferred compound of formula II-D is

[0126]

[0127] And its pharmaceutically acceptable salts.

[0128] In other respects, R3 is -C(O)-OY-CH2-OC(O)-ZC(O)-OH and the compounds of formula II are compounds of formula II-E or their pharmaceutically acceptable salts.

[0129]

[0130] in

[0131] Y is a aryl group; and

[0132] Z is C 1-6 alkyl.

[0133] In these aspects of formulas II-E, Y can be phenylene or naphthylene, preferably phenylene.

[0134] In these aspects of formulas II-E, Z can be a C1 alkane, C2 alkane, C3 alkane, C4 alkane, C5 alkane, or C6 alkane. In some aspects, Z is C 1-2 Alkane. In some respects, Z is a C1 alkane. In other respects, Z is a C2 alkane.

[0135] Compounds of Formula II, including compounds of Formulas II-A, II-B, II-C, II-D, and II-E, may exist as pharmaceutically acceptable salts where applicable. These salts include sodium salts. Potassium, lithium, calcium, and magnesium salts are also contemplated. Alternative salt forms include ammonium salts, choline salts, and tromethamine salts. Pharmaceutically acceptable organic salts of Formula II compounds are also within the scope of this disclosure.

[0136] Compounds of formulas I and II, including compounds of formulas IA, IB, II-A, II-B, II-C, II-D, and II-E, and their pharmaceutically acceptable salts, can be prepared into pharmaceutical compositions by mixing said compounds with pharmaceutically acceptable excipients. In some embodiments, said one or more additional pharmaceutically acceptable excipients are selected from preservatives, antioxidants, or mixtures thereof. In a further embodiment of this disclosure, said additional pharmaceutically acceptable excipients are preservatives, such as, but not limited to, phenol, cresol, parabens, chlorobutanol, or mixtures thereof. In a further embodiment of this disclosure, said additional pharmaceutically acceptable excipients are antioxidants, such as, but not limited to, ascorbic acid, sodium metabisulfite, palmitic acid, butylated hydroxyanisole, butylated hydroxytoluene, tocopherol, or mixtures thereof.

[0137] The pharmaceutical compositions disclosed herein can be provided as suspensions. In other embodiments, the pharmaceutical compositions disclosed herein can be provided as solutions.

[0138] The pharmaceutical compositions disclosed herein may contain the compound of the disclosed invention at a concentration of about 1 mg / mL to about 400 mg / mL, for example, 1 mg / mL to about 200 mg / mL, 1 mg / mL to about 300 mg / mL, preferably 5 mg / mL to about 125 mg / mL, preferably at physiological pH. In certain embodiments of the present disclosure, the compound of the disclosed invention is present at a concentration equal to or greater than about 5 mg / mL. In further embodiments, the compound of the disclosed invention is present at a concentration of about 10 to 25 mg / mL. In even further embodiments, the compound of the disclosed invention is present at concentrations of about 1 mg / mL, 5 mg / mL, 10 mg / mL, 15 mg / mL, 20 mg / mL, 25 mg / mL, 30 mg / mL, 35 mg / mL, 40 mg / mL, 45 mg / mL, or 50 mg / mL. In a further embodiment, the compounds of this disclosure are present at concentrations of about 125 mg / mL, 150 mg / mL, 175 mg / mL, 200 mg / mL, 225 mg / mL, 250 mg / mL, 275 mg / mL, 300 mg / mL, 325 mg / mL, 350 mg / mL, 375 mg / mL, or about 400 mg / mL.

[0139] In some embodiments, the compounds of this disclosure are present at a concentration equal to or greater than about 55 mg / mL. In further embodiments, the compounds of this disclosure are present at a concentration of about 55 to 125 mg / mL. In specific embodiments, the compounds of this disclosure are present at concentrations of about 75 mg / mL, 80 mg / mL, 85 mg / mL, 90 mg / mL, 95 mg / mL, 100 mg / mL, 105 mg / mL, 110 mg / mL, 115 mg / mL, 120 mg / mL, or 125 mg / mL. In other embodiments, the compounds of this disclosure are present at concentrations of about 75 mg / mL to 95 mg / mL, 80 mg / mL to 100 mg / mL, 90 mg / mL to 110 mg / mL, 95 mg / mL to 105 mg / mL, 95 mg / mL to 115 mg / mL, 100 mg / mL to 110 mg / mL, 110 mg / mL to 125 mg / mL, including all ranges and subranges therebetween.

[0140] In some embodiments, the pharmaceutical compositions of this disclosure may further comprise one or two or more stabilizers. In a further embodiment of this disclosure, the stabilizer is selected from surfactants, polymers, cross-linked polymers, buffers, electrolytes, and non-electrolytes. In a further further embodiment of this disclosure, the composition comprises a combination of two or more stabilizers selected from surfactants, polymers, cross-linked polymers, buffers, electrolytes, and non-electrolytes. In a further further embodiment of this disclosure, the stabilizer is a surfactant, such as, but not limited to, polyethylene oxide (PEO), PEO derivatives, polysorbate 80, polysorbate 20, poloxamer 188, polyethoxylated vegetable oil, lecithin, human serum albumin, and mixtures thereof. In a particular embodiment of this disclosure, the stabilizer is a polymer, such as, but not limited to, polyvinylpyrrolidone (e.g., but not limited to, povidone K12, povidone K17, and mixtures thereof), polyethylene glycol 3350, and mixtures thereof. In other embodiments of this disclosure, the stabilizer is an electrolyte, such as, but not limited to, sodium chloride, calcium chloride, and mixtures thereof. In other embodiments of this disclosure, the stabilizer is a non-electrolyte, such as, but not limited to, dextran, glycerol, mannitol, or mixtures thereof. In other embodiments of this disclosure, the stabilizer is a cross-linked polymer, such as, but not limited to, sodium carboxymethyl cellulose (CMC). In some embodiments of this disclosure, the stabilizer is CMC 7LF, CMC 7MF, CMC 7HF, or mixtures thereof.

[0141] In further embodiments of this disclosure, a combination of non-electrolyte stabilizers and electrolyte stabilizers may be used. In some embodiments, the combination of stabilizers may comprise two or more non-electrolyte stabilizers. In other embodiments, the combination of stabilizers may comprise two or more electrolyte stabilizers. In a further embodiment, the combination of stabilizers may comprise one or more non-electrolyte stabilizers and one or more electrolyte stabilizers. In an even further embodiment, the combination of stabilizers may comprise two or more of mannitol, dextrose, and sodium chloride.

[0142] In some embodiments of this disclosure, a combination of surfactant stabilizers and polymer stabilizers may be used. In some embodiments, the combination of stabilizers may comprise two or more surfactant stabilizers. In other embodiments, the combination of stabilizers may comprise two or more polymer stabilizers. In a further embodiment, the combination of stabilizers may comprise one or more surfactant stabilizers and one or more polymer stabilizers. In a still further embodiment, the combination of stabilizers may comprise two or more of polysorbate 80, polysorbate 20, and poloxamer 188. In a still further embodiment, the combination of stabilizers may comprise one or more of polysorbate 80, polysorbate 20, and poloxamer 188, and one or more of povidone K12, povidone K17, and polyethylene glycol 3350.

[0143] In some embodiments of this disclosure, the composition comprises one or more of the stabilizers at a concentration of about 0.2 mg / mL to about 75 mg / mL, and all ranges and subranges therebetween. In specific embodiments of this disclosure, the composition comprises one or more stabilizers at concentrations of about 0.2 to 0.7 mg / mL, 0.5 to 1 mg / mL, 1 to 5 mg / mL, 2 to 8 mg / mL, 5 to 6 mg / mL, 5 to 10 mg / mL, 8 to 12 mg / mL, 10 to 15 mg / mL, 15 to 20 mg / mL, 20 to 30 mg / mL, 30 to 40 mg / mL, 40 to 50 mg / mL, 45 to 55 mg / mL, 50 to 60 mg / mL, or 60 to 75 mg / mL, and all ranges and subranges therebetween. In a further embodiment of this disclosure, the composition comprises one or more stabilizers at a concentration of about 0.2 mg / mL, 0.5 mg / mL, 1 mg / mL, 2 mg / mL, 3 mg / mL, 4 mg / mL, 5 mg / mL, 5.5 mg / mL, 6 mg / mL, 7 mg / mL, 8 mg / mL, 9 mg / mL, 10 mg / mL, 12 mg / mL, 15 mg / mL, 17 mg / mL, 20 mg / mL, 25 mg / mL, 30 mg / mL, 35 mg / mL, 40 mg / mL, 45 mg / mL, 50 mg / mL, 55 mg / mL, 60 mg / mL, 65 mg / mL, 70 mg / mL, or 75 mg / mL.

[0144] In certain embodiments of this disclosure, the composition further comprises one or more buffers, such as, but not limited to, NaH₂PO₄·H₂O, NaH₂PO₄·2H₂O, anhydrous NaH₂PO₄, sodium citrate, citric acid, Tris, sodium hydroxide, HCl, or mixtures thereof. In some embodiments of this disclosure, the composition comprises one or more buffers of about 1 mM to 20 mM, and all ranges and subranges thereof. In certain embodiments of this disclosure, the composition comprises one or more buffers of about 1 to 2 mM, 1 to 3 mM, 1 to 5 mM, 2 to 8 mM, 5 to 6 mM, 5 to 10 mM, 8 to 12 mM, 10 to 15 mM, or 15 to 20 mM, and all ranges and subranges thereof. In a further embodiment of this disclosure, the composition comprises one or more buffers of about 1 mM, 2 mM, 3 mM, 4 mM, 5 mM, 6 mM, 7 mM, 8 mM, 9 mM, 10 mM, 11 mM, 12 mM, 13 mM, 14 mM, 15 mM, 16 mM, 17 mM, 18 mM, 19 mM, or 20 mM.

[0145] In some embodiments of this disclosure, the pH of the pharmaceutical composition is from about 3 to 10, for example, 3, 4, 5, 6, 7, 8, 9, or 10. In further embodiments of this disclosure, the pH of the composition is from about 5 to 9. In further embodiments of this disclosure, the pH of the composition is from about 6 to 9. In further embodiments of this disclosure, the pH of the composition is from about 6 to 7. In further embodiments of this disclosure, the pH of the composition is from about 6 to 8.5. In further embodiments of this disclosure, the pH of the composition is from about 7 to 8.5. In further embodiments of this disclosure, the pH of the composition is greater than 7 to 8.5. In some embodiments of this disclosure, the pH of the composition is from about 6.0 to 8.0. In specific embodiments of this disclosure, the pH of the composition is from about 6.0 to 7.0, 6.5 to 7.0, 6.5 to 7.5, 6.7 to 7.2, 7.0 to 7.2, 7.0 to 7.5, 7.0 to 8.0, or 7.0 to 8.5.

[0146] In some embodiments of this disclosure, the molar osmolarity of the pharmaceutical composition ranges from about 280 mOsm / L to about 310 mOsm / L, for example, about 280, 285, 290, 300, 305, or about 310 mOsm / L. In further embodiments of this disclosure, the molar osmolarity of the composition ranges from about 290 mOsm / L to about 300 mOsm / L. In still further embodiments of this disclosure, the molar osmolarity of the composition is about 290 mOsm / L. In some embodiments, the molar osmolarity can be selected by using an appropriate amount of one or more stabilizers acting as tonicators in the composition, such as, but not limited to, the non-electrolyte stabilizers and electrolyte stabilizers described herein. In some embodiments, the molar osmolarity can be selected by using an appropriate amount of one or more buffers acting as tonicators in the composition, such as, but not limited to, the buffers described herein.

[0147] The pharmaceutical compositions disclosed herein can be administered intravenously. Alternatively, the pharmaceutical compositions of the present invention can be administered intramuscularly. In other embodiments, the pharmaceutical compositions of the present invention are administered subcutaneously. The pharmaceutical compositions of the present invention can also be administered orally. In other embodiments, the pharmaceutical compositions of the present invention are administered via mucous membranes, such as intranasal administration. In other embodiments, the pharmaceutical compositions of the present invention are administered intraosseously.

[0148] The compounds and pharmaceutical compositions disclosed herein can be used to treat conditions that respond to dantrolene. For example, a therapeutically effective amount of the disclosed compound, or a salt thereof, can be administered to a subject requiring treatment. In other aspects, a therapeutically effective amount of the disclosed pharmaceutical composition, or a salt thereof, can be administered to a subject requiring treatment. In other aspects, a subject requiring treatment can be exposed to a therapeutically effective amount of the disclosed compound, such as compounds of formula IA, IB, II-A, II-B, II-C, II-D, II-E, or a pharmaceutically acceptable salt thereof. For example, a subject requiring treatment can be exposed to a therapeutically effective amount of the disclosed compound, such as a compound of formula II-A, or a pharmaceutically acceptable salt thereof.

[0149] Conditions that respond to dantrolene include, for example, malignant hyperthermia, chronic rigidity, exertional heatstroke, cardiac arrhythmias, tachycardia, atrial fibrillation, cardiac arrest, myocardial infarction, heart failure, myocardial injury, cardiomyopathy, central core disease, amyotrophic lateral sclerosis, rhabdomyolysis, Duchenne muscular dystrophy, ataxia, detrusor overactivity, overactive bladder, seizures, epilepsy, antipsychotic malignant syndrome, human stress disorder, Alzheimer's disease, Huntington's disease, multiple sclerosis, amyotrophic lateral sclerosis, Parkinson's disease, ischemia-reperfusion injury, neuronal reperfusion injury, hypoxia, cerebral aneurysm, subarachnoid hemorrhage, stroke, and hyperthermia associated with drug abuse or overdose.

[0150] In a preferred aspect, the compounds and / or pharmaceutical compositions disclosed herein are used to treat malignant hyperthermia in a subject.

[0151] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat chronic rigidity in subjects.

[0152] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat exertional heatstroke in subjects.

[0153] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat cardiac arrhythmias in subjects.

[0154] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat tachycardia in subjects.

[0155] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat atrial fibrillation in subjects.

[0156] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat cardiac arrest in subjects.

[0157] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat myocardial infarction in subjects.

[0158] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat heart failure in subjects.

[0159] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat myocardial injury in subjects.

[0160] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat cardiomyopathy in subjects.

[0161] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat central axial void disease in subjects.

[0162] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat subjects with amyotrophic lateral sclerosis (ALS).

[0163] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat rhabdomyolysis in subjects.

[0164] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat subjects with Duchenne muscular dystrophy.

[0165] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat ataxia in subjects.

[0166] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat subjects with detrusor muscle overactivity.

[0167] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat subjects with excessive bladder activity.

[0168] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat epileptic seizures in subjects.

[0169] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat epilepsy in the subject.

[0170] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat a subject with malignant antipsychotic syndrome.

[0171] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat human stress disorders in subjects.

[0172] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat subjects with Alzheimer's disease.

[0173] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat Huntington's disease in subjects.

[0174] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat multiple sclerosis in subjects.

[0175] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat Parkinson's disease in subjects.

[0176] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat ischemia-reperfusion injury in subjects.

[0177] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat neuronal reperfusion injury in subjects.

[0178] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat hypoxia in subjects.

[0179] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat cerebral aneurysms in subjects.

[0180] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat subarachnoid hemorrhage in subjects.

[0181] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat stroke in subjects.

[0182] In other respects, the compounds and / or pharmaceutical compositions disclosed herein are used to treat subjects with hyperthermia associated with drug abuse (e.g., ecstasy (3,4-methylenedioxymethamphetamine) abuse).

[0183] In other respects, the compounds and / or pharmaceutical compositions of this disclosure are used to treat subjects with hyperthermia associated with drug overdose (e.g., ecstasy (3,4-methylenedioxymethamphetamine) overdose).

[0184] In other respects, the compounds and / or pharmaceutical compositions of this disclosure are used to treat acetylcholine accumulation in a subject. In other respects, the compounds and / or pharmaceutical compositions of this disclosure are used to treat neurotoxic nerve agent exposure in a subject, such as nerve gas exposure (e.g., organophosphorus gases such as sarin, soman, and VX). See, for example, U.S. Provisional Application No. 62 / 554,049, filed September 5, 2017. As used herein, “neurotoxic nerve agent” or “nerve agent” refers to a compound that affects the transmission of nerve impulses in the nervous system. Nerve agents are organophosphorus compounds, i.e., they have the formula (R)3P(O), wherein each R group may be the same or different. "G" type nerve agents include: O-pinacol methylfluorophosphonate (Soman, GD), ethyl N,N-dimethylphosphatamidocyanate (Tabeng, GA), prop-2-methyl methylfluorophosphonate (Sarin, GB), cyclohexyl methylfluorophosphonate (Cyclosalin, GF), and 2-(dimethylamino)ethyl (GV). "V" type nerve agents include: O-cyclopentyl S-(2-diethylaminoethyl)methyl thiophosphonate (EA-3148), (S)-({{2-(diethylamino)ethyl]sulfonyl}(ethyl)phosphonate ethyl ester), for example (S)-{[2-(diethylamino)ethyl]thioalkyl}(ethyl)phosphonic acid ethyl ester (VE), O,O-diethyl S-[2-(diethylamino)ethyl]thiophosphate (VG), S-[2-(diethylamino)ethyl]O-ethylmethyl thiophosphonate (VM), N,N-diethyl-2-(methyl-(2-methylpropoxy)phosphoyl)thioalkylethylamine (VR), and ({2-[bis(propyl-2-yl)amino]ethyl}thioalkyl)(methyl)phosphonic acid ethyl ester (VX). The methods described herein can be used to treat subjects exposed to one nerve agent. The methods described herein can be used to treat subjects exposed to two or more nerve agents.

[0185] As used in this article, the phrases “caused by exposure to nerve agents” and “as a result of nerve agent exposure” refer to the effects as a direct consequence of nerve agent exposure, as well as the effects as a secondary consequence of nerve agent exposure.

[0186] In some aspects, this disclosure relates to a method of treating a subject exposed to a nerve agent with a pharmaceutical composition comprising an amount of a compound of Formula I as described herein or a pharmaceutically acceptable salt thereof. For example, in some aspects, the method prevents neurological damage secondary to nerve agent exposure. In other aspects, the method provides a neuroprotective effect after nerve agent exposure. In other aspects, the method improves brain tissue damage secondary to nerve agent exposure. In other aspects, the method improves brain tissue damage resulting from status epilepticus secondary to nerve agent exposure. In other aspects, the method prevents neuronal necrosis caused by nerve agent exposure. In other aspects, the method improves neuronal necrosis caused by nerve agent exposure. In other aspects, the method treats intracellular calcium overload caused by nerve agent exposure. In other aspects, the method improves intracellular calcium overload caused by nerve agent exposure. In other aspects, the method prevents intracellular calcium overload caused by nerve agent exposure.

[0187] Subjects described herein may be exposed to nerve agents through inhalation. In other respects, subjects may be exposed to the nerve agents through transdermal delivery. Still in other respects, subjects may be exposed to nerve agents by consuming liquids or food contaminated with the nerve agent. In still other respects, subjects may be exposed to the nerve agents by subcutaneous, intravenous, or intramuscular administration.

[0188] In some aspects, the method relates to protecting a subject from nerve necrosis after exposure to a nerve agent. In these embodiments, after exposure to a nerve agent, a pharmaceutical composition comprising an amount of a compound of formula I or a pharmaceutically acceptable salt thereof is administered to the subject. As used herein, “protection” from nerve necrosis includes mitigating the severity of the nerve agent effect or improving the effect of the nerve agent or reducing nerve damage caused by nerve agent exposure. In some aspects, “protection” from nerve necrosis includes preventing nerve necrosis in a subject already exposed to a nerve agent. That is, subjects “protected” from nerve necrosis by administration of the compounds and compositions described herein perform better in neurobehavioral tests compared to nerve agent-exposed subjects who have not been administered the compounds or compositions described herein.

[0189] In some embodiments, the entire central nervous system of the subject is protected from nerve necrosis. In some embodiments, the frontoparietal cortex, hippocampus, and / or thalamus are protected from nerve necrosis. In other aspects, the frontoparietal cortex is protected from nerve necrosis. In other aspects, the hippocampus is protected from nerve necrosis. In other embodiments, the thalamus is protected from nerve necrosis.

[0190] The presence and extent of nerve necrosis can be determined using methods known in the art, including neurobehavioral testing, radiological testing, and pathological evaluation.

[0191] This disclosure also relates to methods for protecting a subject from central nervous system dysfunction caused by exposure to a nerve agent. These methods include administering a pharmaceutical composition comprising an amount of a compound of formula I or a pharmaceutically acceptable salt thereof to the subject after the subject has been exposed to the nerve agent.

[0192] This disclosure also relates to methods for protecting a subject from central nervous system dysfunction caused by exposure to a nerve agent. These methods include administering, after the subject has been exposed to a nerve agent, a pharmaceutical composition comprising an amount of a compound of formula I as described herein or a pharmaceutically acceptable salt thereof to the subject.

[0193] This disclosure also relates to methods for treating behavioral changes in subjects caused by exposure to nerve agents. These methods include administering a pharmaceutical composition comprising an amount of a compound of formula I or a pharmaceutically acceptable salt thereof to the subject after the subject has been exposed to the nerve agent.

[0194] As used herein, “protection” against central nervous system dysfunction includes mitigating the severity of the central nervous system effects of nerve agents, improving the central nervous system effects of nerve agents, or reducing the central nervous system effects of nerve agents. That is, subjects “protected” against central nervous system dysfunction by administration of a composition containing compound I performed better in neurobehavioral tests compared to nerve agent-exposed subjects who were not administered the composition.

[0195] This disclosure also relates to methods for treating nerve agent-induced seizures in subjects exposed to nerve agents. In some aspects, the treated seizures are status epilepticus (SE). These methods include administering to the subject a pharmaceutical composition comprising an amount of a compound of formula I or II or a pharmaceutically acceptable salt thereof. As used herein, treatment of nerve agent-induced seizures results in a reduction in the severity or duration of the seizures. In other aspects, said treatment results in a reduction in both the severity and duration of the seizures.

[0196] The amount of the compound of formula I or II or a pharmaceutically acceptable salt thereof that is effective in treating a subject according to any of the methods described shall be determined by a practitioner experienced in the art. The therapeutically effective amount may be the amount required for a single dose to treat the subject. Alternatively, the therapeutically effective amount may be the cumulative amount of dantrolene required for the subject during long-term treatment.

[0197] In embodiments intended for human use, the effective amount of the compound of formula I or II is an amount equivalent to 1 mg / kg to 100 mg / kg of dantrolene, administered in one or more doses. In other aspects, the effective amount of the compound of formula I or II is equivalent to 1 mg / kg to about 90 mg / kg of dantrolene. In other aspects, the effective amount of the compound of formula I or II is equivalent to 1 mg / kg to about 80 mg / kg of dantrolene. In other aspects, the effective amount of the compound of formula I or II is equivalent to 1 mg / kg to about 70 mg / kg of dantrolene. In other aspects, the effective amount of the compound of formula I or II is equivalent to 1 mg / kg to about 60 mg / kg of dantrolene. In other aspects, the effective amount of the compound of formula I or II is equivalent to 1 mg / kg to about 50 mg / kg of dantrolene. In other aspects, the effective amount of the compound of formula I or II is equivalent to 1 mg / kg to about 40 mg / kg of dantrolene. In other aspects, the effective amount of the compound of formula I or II is equivalent to about 1 mg / kg to about 30 mg / kg of dantrolene. In other aspects, the effective amount of the compound of formula I or II is equivalent to about 1 mg / kg to about 20 mg / kg of dantrolene. In other aspects, the effective amount of the compound of formula I or II is equivalent to about 5 mg / kg to about 30 mg / kg of dantrolene. In other aspects, the effective amount of the compound of formula I or II is equivalent to about 10 mg / kg to about 30 mg / kg of dantrolene. In other aspects, the effective amount of the compound of formula I or II is equivalent to about 15 mg / kg to about 30 mg / kg of dantrolene. In other aspects, the effective amount of the compound of formula I or II is equivalent to about 20 mg / kg to about 30 mg / kg of dantrolene. In other aspects, the effective amount of the compound of formula I or II is equivalent to about 5 mg / kg to about 20 mg / kg of dantrolene. In other aspects, the effective amount of the compound of formula I or II is equivalent to about 5 mg / kg to about 15 mg / kg of dantrolene. In other aspects, the effective amount of the compound of formula I or II is equivalent to about 5 mg / kg to about 10 mg / kg of dantrolene. In other aspects, the effective amount of the compound of formula I or II is equivalent to about 10 mg / kg to about 20 mg / kg of dantrolene. In other aspects, the effective amount of the compound of formula I or II is equivalent to about 2 mg / kg to about 10 mg / kg, preferably from about 2 mg / kg to about 6 mg / kg of dantrolene. In other aspects, the effective amount of the compound of formula I or II is equivalent to about 15 mg / kg to about 20 mg / kg of dantrolene. In other aspects, the effective amount of the compound of formula I or II is equivalent to 10 mg / kg to 100 mg / kg of dantrolene. In other aspects, the effective amount of the compound of formula I or II is equivalent to 20 mg / kg to 100 mg / kg of dantrolene.In other aspects, the effective amount of the compound of formula I or II is equivalent to 30 mg / kg to 100 mg / kg of dantrolene. In other aspects, the effective amount of the compound of formula I or II is equivalent to 40 mg / kg to 100 mg / kg of dantrolene. In other aspects, the effective amount of the compound of formula I or II is equivalent to 50 mg / kg to 100 mg / kg of dantrolene. In other aspects, the effective amount of the compound of formula I or II is equivalent to 50 mg / kg to 75 mg / kg of dantrolene. In other aspects, the effective amount of the compound of formula I or II is equivalent to 25 mg / kg to 75 mg / kg of dantrolene. In some aspects, the effective amount of the compound of formula I or II is equivalent to 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, or about 34 mg / kg of dantrolene. In some aspects, the effective amount of the compound of formula I or II for treating human subjects is equivalent to 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or about 100 mg / kg of dantrolene. In other respects, the effective amount of the compound of formula I or II is the same as that of about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50. The following numbers are equivalent to approximately 100 mg / kg of dantrolene: 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99.

[0198] In some aspects of this disclosure, the timing of administration of the pharmaceutical composition comprising a compound of formula I or II or a pharmaceutically acceptable salt thereof to a subject exposed to a nerve agent may affect the amount of neurotoxic protection conferred upon the subject.

[0199] In some aspects of this disclosure, the timing of administration of the pharmaceutical composition comprising a compound of formula I or II or a pharmaceutically acceptable salt thereof to a subject exposed to a nerve agent may affect the amount of central nervous system dysfunction conferred upon the subject.

[0200] In some aspects of this disclosure, the timing of administration of the pharmaceutical composition comprising a compound of formula I or II or a pharmaceutically acceptable salt thereof to a subject exposed to a nerve agent may affect the treatment of nerve agent-induced seizures in the subject.

[0201] Regarding the timing of administration of pharmaceutical compositions comprising the compound of formula I or II or a pharmaceutically acceptable salt thereof, in some aspects, at least one dose of the pharmaceutical composition comprising the compound of formula I or II or a pharmaceutically acceptable salt thereof is administered to the subject 24 hours or less after exposure to the nerve agent. In some aspects, at least one dose of the pharmaceutical composition comprising the compound of formula I or II or a pharmaceutically acceptable salt thereof is administered to the subject 20 hours or less after exposure to the nerve agent. In some aspects, at least one dose of the pharmaceutical composition comprising the compound of formula I or II or a pharmaceutically acceptable salt thereof is administered to the subject 16 hours or less after exposure to the nerve agent. In some aspects, at least one dose of the pharmaceutical composition comprising the compound of formula I or II or a pharmaceutically acceptable salt thereof is administered to the subject 12 hours or less after exposure to the nerve agent. In some aspects, a pharmaceutical composition comprising a compound of formula I or II or a pharmaceutically acceptable salt thereof is administered to the subject 8 hours or less after exposure to a nerve agent. In some aspects, a pharmaceutical composition comprising a compound of formula I or II or a pharmaceutically acceptable salt thereof is administered to the subject 4 hours or less after exposure to a nerve agent. In some aspects, a pharmaceutical composition comprising a compound of formula I or II or a pharmaceutically acceptable salt thereof is administered to the subject 2 hours or less after exposure to a nerve agent. In some aspects, a pharmaceutical composition comprising a compound of formula I or II or a pharmaceutically acceptable salt thereof is administered to the subject 1 hour or less after exposure to a nerve agent. In some respects, at least one dose of a pharmaceutical composition comprising a compound of formula I or II or a pharmaceutically acceptable salt thereof is administered to the subject within approximately 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23 hours or approximately 24 hours after the subject has been exposed to a nerve agent.

[0202] While in some respects, the pharmaceutical composition comprising a compound of formula I or II or a pharmaceutically acceptable salt thereof may deliver an effective amount of the compound of formula I or II to a subject exposed to the nerve agent in a single dose, in other respects, two or more doses of the pharmaceutical composition may be required to deliver an effective amount of the compound of formula I or II to a subject exposed to the nerve agent. For example, 2, 3, 4, 5, 6, 7, 8, 9, or 10 doses of the pharmaceutical composition may be required to deliver an effective amount of the compound of formula I or II to a subject exposed to the nerve agent. These additional doses may be administered substantially simultaneously with the first dose. In other respects, the additional doses are time-separated from the first dose. In those respects where three or more doses are administered, each dose may be time-separated from the administration of any other dose. The interval between doses can be one or more hours, for example, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, or 24 hours. In other respects, the interval between doses can be one day or more.

[0203] According to this disclosure, administering a compound of formula I or II to a subject exposed to a nerve agent is an adjunctive therapy for nerve agent exposure. Subjects exposed to nerve agents may also be administered one or more nerve agent antidotes. One class of antidotes for nerve agent exposure is acetylcholinesterase reactivators, such as asoxime chloride (HI-6). Another class of antidotes for nerve agent exposure is inverse antagonists of acetylcholine receptors, such as metronidazole. Subjects exposed to nerve agents may also be administered anti-seizure drugs. Exemplary antiepileptic drugs include: aldehydes (e.g., metaldehyde), aromatic allyl alcohols (e.g., stearyl alcohol), benzodiazepines (e.g., clonazepam, clonazepam, diazepam, midazolam, lorazepam, nitrazepam, temazepam, nimesapam), barbiturates (e.g., phenobarbital, methylphenobarbital, barbitxalon), bromides (e.g., potassium bromide), carbamates (e.g., non-amino esters), carboxamides (e.g., carbamazepine, oxcarbazepine, eslicarbazepine acetate), fatty acids (e.g., valproic acid, sodium valproate, sodium divalproate, vigabatrin, progabi, tiagabine), topiramate, GABA analogs (e.g., gabapentin, pregabalin), and hydantoin derivatives (…). Examples include phenytoin, toluene-toin, fosphenytoin, oxadione, oxadione, oxadione, propionates (e.g., becqueramine), pyrimidine diones (e.g., primidone), pyrrolidines (e.g., brivaracetam, levetiracetam, cetrazetam), succinimides (e.g., ethosuximide, phenosuximide, methosuximide), sulfonamides (e.g., acetazolamide, sultiame, acetazolamide, zonisamide), triazines (e.g., lamotrigine), ureas (e.g., pheneturide, phenacemide), valproylamides (e.g., valproamide, penoxamide), perampanel, and combinations thereof. In some aspects, the antiepileptic drug is a benzodiazepine, such as midazolam. In other aspects, the antiepileptic drug is a barbiturate. In other aspects, the antiepileptic drug is hydantoin. In some aspects, the antiepileptic drug is metaldehyde. In other aspects, the antiepileptic drug is potassium bromide. In some aspects, the antiepileptic drug is a fatty acid. In other aspects, the antiepileptic drug is topiramate.

[0204] In those instances where an antidote is administered to a subject exposed to a nerve agent, the compound of formula I or II is administered after the antidote has been administered. For example, the compound of formula I or II may be administered after administration of an acetylcholinesterase reactivator and / or after administration of an inverse antagonist of an acetylcholine receptor.

[0205] In those instances of administering an antiepileptic drug to a subject exposed to a nerve agent, the compound of formula I or II may be administered simultaneously with the administration of the antiepileptic drug. The compound of formula I or II may be administered substantially simultaneously with the administration of the antiepileptic drug, for example, within approximately 5 minutes of the administration of the antiepileptic drug. In other embodiments, the compound of formula I or II is administered before the administration of the antiepileptic drug. In other embodiments, the compound of formula I or II is administered after the administration of the antiepileptic drug.

[0206] The pharmaceutical compositions comprising compounds of formula I or II, or pharmaceutically acceptable salts thereof, may be administered intravenously. In other aspects, the pharmaceutical compositions comprising compounds of formula I or II, or pharmaceutically acceptable salts thereof, may be administered transdermally. In other aspects, the pharmaceutical compositions comprising compounds of formula I or II, or pharmaceutically acceptable salts thereof, may be administered intramuscularly. In other aspects, the pharmaceutical compositions comprising compounds of formula I or II, or pharmaceutically acceptable salts thereof, may be administered intraosseously. In other aspects, the pharmaceutical compositions comprising compounds of formula I or II, or pharmaceutically acceptable salts thereof, may be administered subcutaneously.

[0207] Preferred pharmaceutical compositions for use in the method comprise a compound of formula I or II or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable excipients. Preferred pharmaceutical compositions comprise a compound of formula I or II or a pharmaceutically acceptable salt thereof, mannitol, polysorbate (e.g., polysorbate 80), povidone (e.g., povidone K12), optionally a pH adjuster (e.g., NaOH or HCl), and water.

[0208] According to this disclosure, dantrolene products in the reference catalogue, for example... In comparison, administration of compounds and / or pharmaceutical compositions as disclosed herein will produce substantially equivalent AUCs in subjects. In other respects, administration of compounds and / or pharmaceutical compositions as disclosed herein will produce substantially equivalent AUCs in subjects compared to comparative compositions. For example, in some aspects, the relative mean AUC of dantrolene in the disclosed pharmaceutical compositions after administration to subjects is... (0-t) and AUC (0-∞) The 90% confidence interval (CI) will be used for dantrolene products in the reference catalog, such as... The relative average AUC of Dantrolin (0-t) and AUC (0-∞) Within 80% to 125% (e.g., 80%, 85%, 90%, 95%, 100%, 105%, 110%, 115%, 120%, or 125%). In some aspects, after administration to a subject, the relative mean AUC of dantrolene in the disclosed pharmaceutical composition is [data missing]. (0-t) and AUC(0-∞) The 90% confidence interval (CI) will be calculated based on the relative mean AUC of dantrolene after administration of the comparison product. (0-t) and AUC (0-∞) Between 80% and 125% (e.g., 80%, 85%, 90%, 95%, 100%, 105%, 110%, 115%, 120% or 125%).

[0209] The following embodiments are provided to illustrate some of the concepts described in this disclosure. While each embodiment is considered to provide a specific individual implementation of this disclosure, none of these embodiments should be considered as limiting the more general implementations described herein. In the following embodiments, efforts have been made to ensure the accuracy of the figures used (e.g., quantities, temperatures, etc.), but some experimental errors and biases should be taken into account. Detailed Implementation

[0210] Example 1

[0211]

[0212] Dantrolin sodium (1 eq.) was dissolved in anhydrous dimethylformamide. Reagent 3 (1 eq.) was added, and the reaction mixture was stirred at 60 °C under nitrogen. After 4 h, another equivalent of reagent 3 was added, and the reaction was stirred overnight at 60 °C. The reaction mixture was then diluted with ethyl acetate and washed twice with saturated sodium chloride. The layers were separated. The organic layer was dried over sodium sulfate and concentrated under vacuum. The crude product was purified by silica gel chromatography. The target product was separated to a purity of 90-95%. 1 The H NMR results were consistent with the prediction of the target product.

[0213] Example 1, Method A: 1a was dried overnight with P2O5. 3 (0.84 mL, 3.72 mmol) was added to a mixture of 1a (500 mg, 1.48 mmol) and DMF (10 mL) at 0 °C, followed by the addition of NaI (245 mg, 1.63 mmol). The resulting mixture was stirred at room temperature for 64 h. The mixture was diluted with EtOAc (30 mL) and brine (20 mL). The organic layer was separated, washed with water (2 x 15 mL), dried over anhydrous Na2SO4, filtered, and evaporated. The crude residue was purified by rapid chromatography (twice), eluting with 0-10% MeOH / CH2Cl2, to give target compound 4 (355 mg, 45%) as a yellow solid.

[0214] Example 1, Method B1a was dried overnight with P2O5. 3 (6.5 mL, 28.79 mmol) was added to a mixture of 1a (8.0 g, 23.8 mmol) and DMF (160 mL) at room temperature, followed by the addition of NaI (4.28 g, 28.55 mmol). The resulting mixture was stirred at room temperature for 40 h. The mixture was diluted with EtOAc (250 mL) and brine (60 mL). The organic layer was separated, washed with water (2 x 75 mL), dried over anhydrous Na2SO4, filtered, and evaporated. The residue was ground with CH2Cl2-hexane to produce a yellow solid (~7 g). This solid was purified by rapid chromatography (twice, with deactivated SiO2), eluting with 0–10% MeOH / CH2Cl2, to give target compound 4 (1.92 g, 15%) as a yellow solid.

[0215] Example 2

[0216]

[0217] The sample of compound 4 was treated with 1 ml of a 9 / 1 mixture of trifluoroacetic acid / water for 20–30 min at ambient temperature. Excess TFA was immediately removed under high vacuum, and the resulting solid was collected by filtration, washed with water (5 ml), and dried. The starting materials, reaction mixture, and final product were analyzed by LC / MS to determine whether 2-reconverted to dantrialeline during deprotection. No reconversion of 2-reconverted to dantrialeline was observed. The product… 1 The H NMR results were consistent with the prediction of the target product.

[0218] Example 2, Method A: TFA (9 mL) was added to a mixture of 4 (886 mg, 1.65 mmol) in CH2Cl2 (9 mL). The resulting mixture was stirred at room temperature for 3 h. The solvent was evaporated to dryness on a rotary evaporator. The resulting residue was ground with hexane for 1 h, and the yellow solid was filtered and dried to yield target compound 2 (660 mg, 94%).

[0219] Example 3

[0220]

[0221] 50 mg of 2 was mixed with 3 ml of methanol (completely dissolved) and applied to a 1 g Na+ ion exchange column. The compound was eluted with methanol and lyophilized to produce 18 mg (36% recovery) of an orange solid. This substance was dissolved in water and carefully titrated to pH 8.5 with the addition of small aliquots of 0.1 M NaOH while stirring. The solution was then lyophilized to produce an orange solid, compound 2a. The LC / MS values ​​of the sample before and after lyophilization were consistent, indicating that no reversion to dantrolene occurred during ion exchange. The product...1 The H NMR results were consistent with the prediction of the target product.

[0222] Example 3, Method A: At room temperature, 0.1N NaOH (23.6 mL, 2.34 mmol) was added in 650 μL aliquots to a stirred suspension of 2 (500 mg, 1.17 mmol) in water (63 mL, HPLC grade) by rapid vortexing until pH 8.5 was reached. The solution was filtered, and the filtrate was lyophilized overnight to yield title compound 2a (530 mg, 96%) as a yellow solid. MS (CI) m / z = 424.9 [M] + . 1 H NMR (300MHz, D2O): δ8.08(d,J=8.8Hz,2H),7.72(d,J=8.8Hz,2H),7.59(s,1H),6 .98(d,J=3.6Hz,1H), 6.86(d,J=3.6Hz,1H), 5.19(d,J=6.0Hz,2H), 4.32(s,2H).

[0223] Example 4. Conversion of 2a to dantrolene by alkaline phosphatase at 25°C

[0224] Incubate with alkaline phosphatase

[0225] The prodrug 2a was incubated with purified alkaline phosphatase at 25°C. The final reaction mixture contained approximately 20 μg / mL of the prodrug and 50 μU / μL of alkaline phosphatase (from calf intestine, Sigma#11097075001) in 1X PBS at pH 7.4. A control mixture containing 20 μg / mL of the prodrug but without the enzyme was also prepared. The enzyme reaction mixture was stored at 25°C and analyzed by HPLC at 10 μL aliquots at 0.9 h, 3.2 h, 5.5 h, 7.7 h, and 19.9 h. The control mixture was also stored at 25°C and analyzed by HPLC at 10 μL aliquots at 1.5 h, 3.8 h, 6.6 h, 8.3 h, and 20.4 h.

[0226] Sample analysis by HPLC

[0227] Analysis was performed using a Waters 2695 Alliance system maintained at 25°C and equipped with a PDA detector and a Restek Ultra C18 column (5 μm, 250 x 4.6 mm). A gradient method was used, with mobile phase A containing acetonitrile and mobile phase B containing acetonitrile:phosphate buffer (33:67) at pH 6.9. The column was equilibrated with 100% mobile phase B and held at this composition for 19 min. Mobile phase A was then increased to 55% after 5 min. The column was washed with 55% A for 2 min, returned to 100% B after 2 min, and then reequilibrated with 100% B for 5 min, for a total run time of 33 min. 10 μL of sample was injected, and the analyte was detected by UV at 375 nm. The prodrug eluted at approximately 3.1 min, while dantrolene eluted at approximately 15.4 min. Peak area changes over time were monitored to determine the conversion of the prodrug to dantrolene. The peak area curve over time is shown in the figure. Figure 1 and Figure 2 .

[0228] Example 5. Conversion of 2a to dantrolene in rat plasma at 22°C

[0229] Warming with blood plasma

[0230] In vitro experiments were performed as follows: 40 μL of approximately 10 mg / mL 2a dissolved in DMF was added to 360 μL of previously frozen rat plasma from male Sprague Dawley rats at 22 °C. The spiked plasma was stored at 22 °C, and 50 μL aliquots were collected at 25 min, 3 h, and 20 h after spiking. Each aliquot was immediately treated with 50 μL of acetonitrile, vortexed, and then centrifuged at 4000 rpm at 25 °C for 5 min. The 50 μL supernatant was diluted 50-fold in acetonitrile:phosphate buffer (33:67, pH 6.9) and transferred to a glass vial for HPLC analysis.

[0231] HPLC analysis of samples

[0232] Analysis was performed using a Waters 2695 Alliance system maintained at 25°C and equipped with a PDA detector and a Restek Ultra C18 column (5 µm, 250 x 4.6 mm). A gradient method was used, with mobile phase A containing acetonitrile and mobile phase B containing acetonitrile:phosphate buffer (33:67) at pH 6.9. The column was equilibrated with 100% mobile phase B and held at this composition for 19 min. Mobile phase A was then increased to 55% after 5 min. The column was washed with 55% A for 2 min, returned to 100% B for 2 min, and then reequilibrated with 100% B for 5 min, for a total run time of 33 min. 10 µL of sample was injected, and the analyte was detected by UV at 375 nm. The prodrug eluted at approximately 3.1 min, while dantrolene eluted at approximately 15.4 min. Peak area changes over time were monitored to determine the conversion of the prodrug to dantrolene. See [link to relevant documentation] Figure 3 .

[0233] Example 6. Conversion of 2a to dantrolene in plasma at 37°C

[0234] Warming with blood plasma

[0235] In vitro experiments were performed by adding approximately 10 mg / mL of a prodrug dissolved in DMF (60 μL) to 690 μL of previously frozen rat plasma from male Sprague-Dawley rats at 37 °C. The spiked plasma was stored at 37 °C, and 50 μL aliquots were collected at 5, 10, 20, 30, 40, 50, 60, 100, 2.5, 3.5, 4, 5, and 6.5 h after spiking. Each aliquot was immediately treated with 50 μL of acetonitrile, vortexed, and then centrifuged at 4000 rpm at 25 °C for 5 min. The 50 μL supernatant was transferred to a glass vial for HPLC analysis.

[0236] HPLC analysis of samples

[0237] Analysis was performed using a Waters 2695 Alliance system maintained at 25°C and equipped with a PDA detector and a Restek Ultra C18 column (5 μm, 250 x 4.6 mm). A gradient method was used, with mobile phase A consisting of water containing 0.1% trifluoroacetic acid and mobile phase B consisting of acetonitrile containing 0.1% trifluoroacetic acid at a flow rate of 1.0 mL / min. The column was equilibrated with 67% mobile phase A / 33% mobile phase B and held at this composition for 19 min. Then, mobile phase B was increased to 70% after 5 min. The column was washed with 70% B for 2 min, returned to 33% B after 2 min, and then reequilibrated with 33% B for 5 min, for a total run time of 47 min. 5 μL of sample was injected, and the analyte was detected by UV at 375 nm. The prodrug eluted at approximately 6.5 min, while dantrolene eluted at approximately 18.5 min. The change in peak area over time was monitored to determine the conversion of the prodrug to dantrolene. The curve of peak area over time is shown in Figure 4 .

[0238] Example 7. Bioavailability of dantrolene in rats after 2 years of administration

[0239] method

[0240] Compound 2a was reconstituted at 8 mg / mL in a 5% mannitol aqueous solution at pH 8.0 (as a tension modifier). The formulation was administered intravenously, sedatively, or intramimetously to cannulated Harlan Sprague Dawley rats (3 rats / group) from Envigo RMS, Inc. (Indianapolis, IN). Each group received 7.5 mg / kg of 2a, which is equivalent to 5 mg / kg dantrolene equivalent (DE). Whole blood (0.1 mL) was collected via a jugular vein catheter at 0, 0.033 (IV only), 0.083, 0.167, 0.33, 0.66, 1, 3, 6, and 9 hours. Immediately after collection, 0.1 mL of whole blood was added to 0.3 mL of acetonitrile to quench the biotransformation of the prodrug. The sample was then placed on wet ice until centrifuged to remove the precipitate. Phenomenex Synergi 4μm PolarRP on a Waters Acquity UPLC system connected to an Applied Biosystems / MDS Sciex API6500LC / MS / MS system. A 75x2 mm column was used to analyze 2a, dantrolene, and the metabolite 5-OH dantrolene in the precipitated whole blood matrix. Samples were quantified based on standard curves prepared using each analyte in the precipitated whole blood matrix.

[0241] Similarly, when Ryanodex was administered at 5 mg / kg...-1 After intravenous administration of a specific dose to rats, plasma concentrations of dantrolene were measured over time. Since Ryanodex is a 3,5-hydrate of dantrolene sodium, this is equivalent to 3.9 mg / kg on a molar basis. -1 The dosage of dantrolene (i.e., 3.9 mg / kg) -1 Dantrolin equivalent (DE)).

[0242] The area under the curve (AUC) was calculated using the trapezoidal rule in SigmaPlot 12.5 software.

[0243] result

[0244] Administration of 2a to rats via IV, IM, and SC routes resulted in the rapid appearance of dantrolene in the blood.

[0245] Example 8. Bioavailability of dantrolene, a compound disclosed herein, in rats.

[0246] method

[0247] The compound of this disclosure was formulated in a 5% mannitol aqueous solution (as a tension modifier). The formulation was administered intravenously, sedatively, or intramuscularly to cannulated Harlan Sprague Dawley rats (3 rats / group) from EnvigoRMS, Inc. (Indianapolis, IN). Each group received an amount equal to 5 mg / kg dantrolene equivalent (DE). Whole blood (0.1 mL) was collected via a jugular vein catheter at 0, 0.033 (IV only), 0.083, 0.167, 0.33, 0.66, 1, 3, 6, and 9 hours. Immediately after collection, 0.1 mL of whole blood was added to 0.3 mL of acetonitrile to quench the biotransformation of the prodrug. The sample was then placed on wet ice until centrifuged to remove the precipitate. The reaction was performed using a Phenomenex Synergi 4 μm Polar RP on a Waters Acquity UPLC system connected to an Applied Biosystems / MDS Sciex API 6500LC / MS / MS system. A 75x2 mm column was used to analyze the parent prodrug, dantrolene, and metabolite 5-OH dantrolene in the precipitated whole blood matrix. Standard curves were prepared based on the precipitated whole blood matrix to quantify the samples.

[0248] Similarly, when Ryanodex was administered at 5 mg / kg... -1 After intravenous administration of a specific dose to rats, plasma concentrations of dantrolene were measured over time. Since Ryanodex is a 3,5-hydrate of dantrolene sodium, this is equivalent to 3.9 mg / kg on a molar basis. -1The dosage of dantrolene (i.e., 3.9 mg / kg) -1 Dantrolin equivalent (DE)).

[0249] The area under the curve (AUC) was calculated using the trapezoidal rule in SigmaPlot 12.5 software.

[0250] result

[0251] Administration of the disclosed compounds to rats via IV, IM, and SC routes results in the rapid appearance of dantrolene in the blood.

[0252] Example 9

[0253] Research Overview

[0254] The objective of this study is to determine whether the compounds of this disclosure (e.g., compound 2a) have neuroprotective effects in survival models of mammals such as dogs, pigs, rabbits, rodents (e.g., rats, mice, guinea pigs), and primates (e.g., monkeys, chimpanzees). An exemplary model is the rat GD (Soman) survival model.

[0255] Following the onset of a nerve agent-induced seizure, a single dose of the disclosed compound is administered. For example, a single dose equivalent to 1 mg / kg to 30 mg / kg dantrolene (e.g., 10 mg / kg or 30 mg / kg) is administered. The disclosed compound can be administered intravenously, subcutaneously, intramuscularly, percutaneously, or intraosseously. For example, the agent can be administered intravenously.

[0256] Treatment with nerve agent antidotes can promote survival. For example, axoxime chloride (HI-6) can be administered before nerve agent exposure (e.g., 30 minutes before a subcutaneous (SQ) soman injection), metronidazole can be administered 1 minute after the SQ soman injection, and midazolam can be administered 20 minutes after a soman-induced seizure occurs and a Racine score of at least 3 is achieved.

[0257] The control group consisted of a group of untreated (native) animals, and another group would receive sterile water after a nerve agent-induced seizure (e.g., 50 minutes after the nerve agent-induced seizure).

[0258] Over a period of time, such as approximately 28 days after a single nerve agent exposure, a series of neurobehavioral tests are performed. The day after the testing period (e.g., day 29), all animals are euthanized under anesthesia, for example by exsanguination and intracardiac perfusion. Brains are collected from each animal for microscopic neuropathological examination, and hearts are collected from each animal for possible pathological examination.

[0259] Material

[0260] Soman (GD) – diluted with 0.9% sodium chloride. Soman is an organophosphate nerve agent that inactivates acetylcholinesterase (AChE) by forming an adduct with it.

[0261]

[0262]

[0263] HI-6: Chemical name: [(E)-[1-[(4-carbamoylpyridin-1-onthiol-1-yl)methoxymethyl]pyridin-2-ylidene]methyl]-oxonium; methanesulfonate (asoxime chloride)

[0264] structure:

[0265]

[0266] Chemical formula: C 14 H 16 Cl2N4O3

[0267] Molecular weight: 359.207

[0268] Metronidazole: Chemical name: (8,8-dimethyl-8-azabicyclo[3.2.1]oct-3-yl)3-hydroxy-2-phenylpropionate; nitrate

[0269] structure:

[0270]

[0271] Chemical formula: C 18 H 26 N2O6

[0272] Molecular weight: 366.414

[0273] Midazolam: Chemical name: 8-chloro-6-(2-fluorophenyl)-1-methyl-4H-imidazo[1,5-a][1,4]benzodiazepine

[0274] structure:

[0275]

[0276] Chemical formula: C 18 H 13 ClFN3

[0277] Molecular weight: 325.771

[0278] dose

[0279] HI-6, Soman, Metronidazole, and Midazolam: Single-dose HI-6 (IP, 125 mg / kg); Soman (SC, 154 μg / kg, 1.4x LD50); Metronidazole (IM, 2 mg / kg); and Midazolam (IM, 2 mg / kg) may be selected. This regimen is expected to induce seizures with a Racine score of at least 3 and a number of eligible survivors for follow-up studies.

[0280] The compounds disclosed herein: any of the compounds of this disclosure or a pharmaceutically acceptable salt thereof may be administered. A preferred compound is compound 2a.

[0281] Dosage preparation

[0282] In those experiments using GD, GD was prepared in ice-cold 0.9% sodium chloride according to SOP MRI-5821, “Preparation of Standards and Samples from Research Development and Testing Evaluation (RDTE) Dilute Solutions”.

[0283] Racine Level

[0284] 1 = Still and motionless & staring

[0285] 2 = Nodding, "shaking like a wet dog"

[0286] 3 = Forelimb clonus

[0287] 4 = Bilateral forelimb clonus

[0288] 5 = Bilateral forelimb clonus, hindlimb rearing, and loss of balance

[0289] Neurobehavioral test

[0290] Brain regions damaged by Soman exposure may include the hippocampus, as well as the entorhinal cortex, frontal cortex, and parietal cortex. These regions contain structures and neural circuits involved in learning, memory formation, information processing, and other cognitive processes. To evaluate the potential neuroprotective effects of the compounds disclosed herein, animals were assessed using a series of behavioral tests requiring learning, memory, sensorimotor integration, and adaptive responses. Examples of such experiments include: 1) a sucrose preference test and 2) a forced swimming test.

[0291] Sucrose Preference Experiment

[0292] The sucrose preference test (SPT) utilizes the natural tendency of rats to prefer sugar water over regular water. This is a well-established test that measures the desire for or lack of pleasurable behavior (pleasure) and requires the animal to adapt to changes in the position of a bottle containing tap water and 1% sucrose water, placed on the left and right sides.

[0293] Prior to SPT, rats were individually housed and given free access to food and water (one water bottle per cage). For the environmental acclimatization portion of SPT, two water bottles were introduced into each rat's cage for 5–6 days. The water bottles were fitted with pipettes to minimize leakage and were weighed approximately every 24 hours. After the acclimatization period, one bottle was filled with approximately 200 mL of 1% sucrose solution, and the other with approximately 200 mL of tap water. Twenty-four hours later, the remaining liquid volume in each bottle was recorded. The bottles were then switched to left / right positions, and the remaining liquid volume in each bottle was recorded again after twenty-four hours. The volume of sucrose solution consumed (mL) was expressed as a percentage of the total volume of liquid consumed (sucrose solution plus water) in each of the two 24-hour periods and compared between groups and days.

[0294] Forced swimming experiment

[0295] The forced swimming test (FST) was developed by Porsolt in the late 1970s as a rapid way to screen the efficacy of antidepressants in rodents. In untreated (“normal”) rodents, the increased immobility observed near the end of the 5-minute FST was interpreted as reflecting “behavioral despair,” and the reversal of this effect by antidepressants was associated with the antidepressant efficacy of these drugs in humans. However, the construct validity of this test has been questioned for several reasons, including: 1) measuring the acute effect of antidepressants in the FST, while in clinically depressed patients, the drug requires 4–6 weeks to achieve clinical improvement; 2) the dependent variable in the FST is the animal’s acute response to the test, rather than a characteristic of the animal; and 3) interpreting floating behavior as 'behavioral despair’ is anthropomorphic. It is now believed that the gradual immobility observed in untreated rats reflects an adaptive response to acute stress from being placed in a container with no possibility of escape.

[0296] In the free-flowing stamina test (FST), swimming activity and immobility were measured in a glass cylindrical chamber (46cm H x 30cm D) filled with water (30cm high, 25°C). The water temperature for all animals was maintained constant at 24–26°C using thermometers. Two swimming time periods were conducted: a preliminary 15-minute 'preliminary experiment' followed by a second 5-minute 'experiment' 24 hours later. Video recordings were made of the experimental time periods. The time spent actively swimming and immobile was recorded per minute during the FST.

[0297] Neuropathology

[0298] A semi-quantitative 6-point scoring system was used to evaluate seven brain slices from each animal under a microscope. Microscopic lesions were graded on a 6-point scale:

[0299] 0 = Normal

[0300] 1 = 1-5 affected cells per 40X microscope field of view

[0301] 2 = 6-20 affected cells per 40X microscope field of view

[0302] 3 = 21-50 affected cells per 40X microscope field of view

[0303] 4 = 50%-80% of the affected cells per 40X microscope field of view

[0304] 5 = >80% of affected cells per 40X microscope field of view

[0305] Preparation of Example 10.2b

[0306]

[0307] At room temperature, a stirred suspension of 2 (100 mg, 0.23 mmol) in water (12 mL, HPLC grade) was added dropwise to Tris (5, 57 mg, 0.47 mmol) dissolved in water (5 mL). The final solution pH was 6.6. The solution was filtered and the filtrate was lyophilized overnight to yield the title compound 2b (150 mg, 95%) as a yellow solid. MS (CI) m / z = 424.9 [M]+. 1 H NMR (300MHz, DMSO-d6): δ8.24(m,2H),7.93(m,2H),7.73(m,1H),7.12(m,1H),6.97(m,1H),5.20(m,2H),4.40(m,2H),3.63(m,15H).

[0308] Example 11.

[0309]

[0310] Step 1: Dry 1a overnight with P2O5. Add glacial acetic acid (340 μL, 5.95 mmol) to a mixture of 1a (1.0 g, 2.97 mmol) and DMF (20 mL) at room temperature. Stir the mixture overnight at room temperature. Pour the mixture onto crushed ice, filter out the solid, and wash with water. Dry the resulting wet solid overnight with anhydrous P2O5 to give target compound 7 (920 mg, 98%) as a yellow solid.

[0311] Step 2: Formalin (4.35 mL, 57.45 mmol, 37% formaldehyde aqueous solution) was added to the suspension of compound 7 (1.35 g, 4.29 mmol) in water (45 mL), followed by K₂CO₃ (51 mg, 0.37 mmol). The mixture was stirred at room temperature for 24 h. The reaction mixture was filtered, and the yellow solid was washed with 3% formaldehyde aqueous solution and dried for 24 h to yield target compound 9 (1.2 g, 82%).

[0312] Step 3: PCl3 (1.2 mL, 13.71 mmol) was slowly added to a solution of compound 9 (615 mg, 1.78 mmol) in DMF:acetone (40 mL, 15:25 mL) at 0 °C. The reaction mixture was stirred at 0 °C for 10 min and then at room temperature for 2 h. The mixture was then poured onto crushed ice, and the resulting yellow solid was filtered, washed with water (3 x 50 mL), and dried under vacuum in P2O5 for 16 h to yield target compound 6 (600 mg, 92%). 1 H NMR (300MHz, DMSO-d6): δ8.33(d,J=8.5Hz,2H),8.03(d,J=8.8Hz,2H),7.87(s ,1H),7.48(d,J=3.3Hz,1H),7.11(d,J=3.6Hz,1H),5.42(s,2H),4.53(s,2H).

[0313] Example 12.

[0314]

[0315] Step 1: Dry 1a overnight with P2O5. Add glacial acetic acid (340 μL, 5.95 mmol) to a mixture of 1a (1.0 g, 2.97 mmol) and DMF (20 mL) at room temperature. Stir the reaction mixture overnight at room temperature. Pour the mixture onto crushed ice, filter the resulting solid, and wash with water. Dry the wet solid overnight with anhydrous P2O5 to give target compound 7 (920 mg, 98%) as a yellow solid.

[0316] Step 2: Formalin (0.29 mL, 3.83 mmol, 37% formaldehyde aqueous solution) was added to the suspension of 7 (90 mg, 0.28 mmol) in water (2.6 mL), followed by K₂CO₃ (3.4 mg, 0.02 mmol). The mixture was stirred at room temperature for 24 h. The reaction mixture was filtered, and the yellow solid was washed with 3% formaldehyde aqueous solution and dried for 24 h to yield target compound 9 (86 mg, 88%). MS (CI) m / z = 343 [M] - . 1H NMR (300MHz, DMSO-d6): δ8.32(d,J=9.0Hz,2H),8.03(d,J=9.1Hz,2H),7.83(s,1H),7.47 (d, J = 3.6 Hz, 1H), 7.08 (d, J = 3.6 Hz, 1H), 6.52 (t, 1H), 4.85 (d, J = 7.1 Hz, 2H), 4.45 (s, 2H).

[0317] Example 13.

[0318]

[0319] Step 1: Dissolve anhydrous DMF (0.8 mL, 10.33 mmol) in anhydrous tetrahydrofuran (13 mL). Add this solution dropwise to a stirred solution of thionyl chloride (0.75 mL, 10.33 mmol) dissolved in tetrahydrofuran (9 mL) and cool in an ice bath. After the addition is complete and the solution has been on ice for 30 minutes, remove the ice bath and immediately add solid N-hydroxysuccinimide (832 mg, 7.23 mmol) (which is completely dissolved), followed by solid morpholine acetic acid (1.0 g, 6.88 mmol) pre-powdered. Morpholine acetic acid dissolves slowly, producing a homogeneous solution that rapidly becomes cloudy. Stir the reaction vigorously overnight at room temperature. Wash the white solid with tetrahydrofuran and dry under vacuum to yield target compound 11 (1.6 g, 96%) as a white solid.

[0320] Step 2: Triethylamine (0.39 mL, 2.8 mmol) was added to a solution of 9 (660 mg, 1.92 mmol) and 11 (928 mg, 3.83 mmol) in anhydrous DMF (12 mL). The resulting mixture was stirred overnight at 60 °C. The reaction mixture was cooled to room temperature and purified by reversed-phase column chromatography using acetonitrile-water as the eluent. The column fraction was analyzed by HPLC, and the fraction containing the product was lyophilized to give a crude compound with a purity of 50%. The crude product was further purified by preparative HPLC using acetonitrile-water. The pure fraction was lyophilized to give title compound 10 (100 mg, 10%) as a yellow solid.

[0321] Step 3: At room temperature, HCl (0.3 mL, 4 N 1,4-diane solution) was added to a stirred solution of 10 (75 mg, 0.16 mmol) in anhydrous 1,4-diane (4 mL), and the resulting mixture was stirred for 2 h. The solvent was evaporated to dryness on a rotary evaporator. The resulting residue was dissolved in water and lyophilized overnight to produce 10c (75 mg, 94%).

[0322] MS(CI)m / z=472.1[M] +. 1 H NMR (300MHz, DMSO-d6): δ8.33(d,J=8.8Hz,2H),8.03(d,J=9.1Hz,2H),7.89(s,1H),7.4 9(d,J=3.6Hz,1H),7.11(d,J=4.1Hz,1H),5.60(s,2H),4.54(s,2H),3.32-3.81(m,10H). 1 H NMR (300MHz, D2O): δ8.17(d,J=8.5Hz,2H),7.81(d,J=8.8Hz,2H),7.60(s,1H),6.93-7.02(m,1H) ,6.88-6.92(m,1H),5.73(s,2H),4.39(s,2H),4.26(s,2H),3.90-4.09(m,4H),3.30-3.52(m,4H).

[0323] Example 14.

[0324]

[0325] Step 1: 13 (2.0 g, 11.48 mmol) was added to a mixture of K₂CO₃ (4.0 g, 28.94 mmol) and TBAHSO₄ (240 mg, 0.70 mmol) in water (8 mL) at 0 °C. The resulting mixture was stirred at 0 °C for 20 min, then 14 (1.3 mL, 12.85 mmol) was added and stirred again for 3 h. The organic layer was separated and washed with water (2 x 5 mL) and saturated brine (5 mL). The CH₂Cl₂ layer was dried over anhydrous Na₂SO₄, filtered, and concentrated under vacuum. The crude residue was purified by rapid chromatography with elution of 0-100% EtOAc / hexane to give target compound 15 (2.2 g, 86%) as a colorless gel.

[0326] Step 2: 1a was dried overnight with P2O5. 1a (1.66 g, 4.93 mmol) was added to a mixture of 15 (2.2 g, 9.87 mmol) in DMF (35 mL) at room temperature. The resulting mixture was stirred at room temperature for 20 h. The mixture was diluted with EtOAc (50 mL) and washed with water (2 x 25 mL) and saturated brine (15 mL). The EtOAc layer was dried over anhydrous Na2SO4, filtered, and evaporated under vacuum. The crude residue was purified by rapid chromatography with 0–100% EtOAc / CH2Cl2 elution (twice), followed by grinding with CH2Cl2-hexane to give target compound 16 (500 mg, 20%) as a yellow solid.

[0327] Step 3: Add TFA (1.8 mL) to the mixture of 16 (340 mg, 0.68 mmol) in CH2Cl2 (18 mL). Stir the resulting mixture overnight at room temperature. Evaporate the solvent to dryness on a rotary evaporator. Grind the resulting residue with hexane for 1 h, filter and dry the yellow solid to yield target compound 12 (300 mg, 99%).

[0328] Step 4: At room temperature, 0.1N NaOH (5.85 mL, 0.58 mmol) was added in 400 μL aliquots to a stirred suspension of 6 (260 mg, 0.58 mmol) in water (36 mL, HPLC grade) and immediately followed by rapid vortexing. The final solution pH was 6.73. The solution was filtered and the filtrate was lyophilized overnight to yield title compound 12a (150 mg, 55%) as a yellow solid. MS (CI) m / z = 445.1 [M] + . 1 H NMR (300MHz, DMSO-d6): δ8.34(d,J=8.8Hz,2H),8.04(d,J=8.8Hz,2H),7.87(s,1H),7.49(d ,J=3.6Hz,1H),7.11(d,J=3.8Hz,1H),5.43(s,2H),4.51(s,2H),2.40(m,2H),2.15(m,2H).

[0329] Example 15.

[0330]

[0331] Step 1: Compound 18 (488 mg, 1.85 mmol) in DMF (2 mL) was added to a solution of compound 9 (500 mg, 1.45 mmol) in anhydrous DMF (10 mL), followed by the addition of TEA (0.3 mL, 2.2 mmol). The resulting mixture was stirred at room temperature for 16 h. The mixture was diluted with EtOAc (50 mL) and washed with water (2 x 15 mL). The organic layer was dried over anhydrous Na2SO4, filtered, and evaporated to dryness. The crude product was purified twice by rapid chromatography with 0–10% MeOH / CH2Cl2 elution to give target compound 19 (350 g, 40%) as a yellow solid.

[0332] Step 2: Add p-toluenesulfonic acid monohydrate (63 mg, 0.32 mmol) to a solution of compound 19 (200 mg, 0.32 mmol) in MeOH:1,4-diane (1:1, 6 mL). Stir the clear solution at room temperature for 16 h. Evaporate the solvent to dryness using a rotary evaporator. Purify the residue by rapid chromatography with 0-10% MeOH / CH2Cl2 to give target compound 20 (125 g, 77%) as a yellow solid.

[0333] Step 3: Add p-toluenesulfonic acid monohydrate (15 mg, 0.07 mmol) to a solution of compound 20 (120 mg, 0.24 mmol) and compound 21 (26.4 mg, 0.26 mmol) in m-xylene:1,4-dioxane (1:1, 16 mL). Molecular sieve (100 mg). The resulting mixture was refluxed for 16 h. The reaction mixture was cooled to room temperature, diluted with 1,4-diane (30 mL), and filtered. The filtrate was evaporated, and the crude residue was purified twice by rapid chromatography with 0-10% MeOH / CH2Cl2 elution to give target compound 17 (16 mg, 11%) as a yellow solid.

[0334] Step 4: 0.1N Tris (90 μL, 8.9 μmol) was added dropwise to a stirred suspension of 17 (5 mg, 8.4 μmol) in water (3 mL, HPLC grade) at room temperature. The mixture was stirred at room temperature for 3 h. The solution was filtered, and the filtrate was lyophilized overnight to yield title compound 17b (6 mg, 100%) as a yellow solid. MS (CI) m / z = 594.1 [M] + . 1 H NMR (300MHz, DMSO-d6): δ9.96(brs,1H),8.34(d,J=8.8Hz,2H),8.05(d,J=8.5Hz,2H),7.89(s,1H),7.5(d,J=3.2Hz,1H),7.46(d,J=8 .8Hz,2H),7.29(d,J=8.5Hz,2H),7.12(d,J=3.5Hz,1H),5.57(s,2H),4.99(s,2H),4.55(s,2H),3.23-3.32(m,9H),2.33-2.36(m,4H).

[0335] Example 16.

[0336]

[0337]

[0338] Step 1: Triethylamine (3.47 mL, 24.93 mmol) was added to a mixture of 23 (2.5 g, 14.28 mmol) and DMF (30 mL) at room temperature, followed by 24 (3.92 mL, 53.9 mmol). The resulting mixture was stirred at room temperature for 40 h. The mixture was diluted with EtOAc (100 mL) and water (50 mL). The EtOAc layer was washed with water (2 x 25 mL), 5% NaHCO3 (25 mL), and saturated brine (15 mL). The EtOAc layer was dried over anhydrous Na2SO4 and concentrated under vacuum. The crude residue was purified by rapid chromatography with elution of 0-100% EtOAc / hexane to give the target compound 25 (657 mg, 21%) as a colorless oil.

[0339] Step 2: 1a was dried overnight with P2O5. 25 (647 mg, 2.89 mmol) was added to the mixture of 1a (647 mg, 1.92 mmol) and DMF (12 mL) at room temperature. The resulting mixture was stirred at room temperature for 110 h. The mixture was diluted with EtOAc (100 mL) and washed with water (2 x 15 mL) and saturated brine (15 mL). The EtOAc layer was dried over anhydrous Na2SO4, filtered, and evaporated under vacuum. The crude residue was purified by rapid chromatography with 0-100% EtOAc / CH2Cl2 elution to give target compound 26 (260 mg, 27%) as a yellow solid.

[0340] Step 3: At room temperature, HCl (4 mL, 4N 1,4-diane solution) was added to a stirred solution of 26 (210 mg, 0.41 mmol) in anhydrous 1,4-diane (4 mL), and the resulting mixture was stirred overnight. The solvent was evaporated to dryness on a rotary evaporator. The resulting residue was ground with hexane for 1 h, and the yellow solid was filtered and dried to yield the title compound 22c (153 mg, 83%). [M] + MS(CI)m / z=402.1[M] + . 1 H NMR (300MHz, DMSO-d6): δ8.47(brs,3H),8.34(d,J=8.8Hz,2H),8.04(d,J=8.8Hz,2H),7.91( s,1H),7.50(d,J=3.6Hz,1H),7.12(d,J=3.6Hz,1H),5.61(s,2H),4.56(s,2H),3.85(s,2H).

[0341] Example 17. General UPLC Materials and Methods

[0342] LC-MS analysis was performed using an Agilent 1290 Infinity II ultra-high performance liquid chromatography system equipped with an Agilent Zorbax Eclipse Plus C18 column (2.1 x 50 mm, 1.8 μm). Elution was monitored at 210 nm and 385 nm using a diode array detector. Mobile phase A was water containing 0.1% (v / v) trifluoroacetic acid, and mobile phase B was acetonitrile containing 0.1% trifluoroacetic acid. 10 μL of each sample was injected, and linear gradient elution was performed, starting at 0.5 mL / min from 25% B and increasing to 43% B within 4 min. The LC system was coupled to an Agilent 6420 triple quadrupole mass spectrometer.

[0343] HPLC standard curve for concentration measurement

[0344] Weigh approximately 1 mg of the target compound and dissolve it to 1 mg / mL using 25% acetonitrile (v / v in water). Prepare a 10-fold dilution of the prodrug using 25% acetonitrile to obtain a 100 μg / mL solution. Perform a series of 2-fold dilutions using 25% acetonitrile to obtain 50, 25, 12.5, and 6.25 μg / mL solutions. For each sample, analyze 10 μL using the equipment and gradient analysis described in the General UPLC Materials and Methods section. A standard curve for the concentration is generated by manually integrating the 385 nm chromatogram and plotting the peak area according to the concentration.

[0345] Pharmacokinetic analysis

[0346] At each time point, 400 μL of blood was collected into K2EDTA tubes and placed on ice before centrifugation. After centrifugation, 100 μL of plasma was combined with 300 μL of acetonitrile and then centrifuged to remove the precipitate. The samples were analyzed using the gradient described in the General UPLC Materials and Methods section. A dantrolene standard curve was created for each experiment as follows: 3.8 mg of dantrolene sodium was dissolved in 14.9 mL of 50% methanol (v / v) aqueous solution. The dantrolene sodium was diluted in a 1:3 supernatant of precipitated rat blood:acetonitrile to generate the sample for the standard curve. The standard curve ranged from 5000 to 5 ng / mL. The absorbance of dantrolene was monitored at 385 nm using a diode array detector and the multiple reaction monitoring function of the mass spectrometer.

[0347] Transformation and pharmacokinetics of compound 2a in plasma

[0348] To measure the conversion of compound 2a to dantrolene in rat plasma, 1 mg of compound 2a was dissolved in water to a concentration of 1 mg / mL. A 100 μL aliquot of compound 2a was combined with 900 μL of plasma from Sprague Dawley rats. The plasma reaction was then incubated in a 37°C water bath. The initial concentration of compound 2a in the plasma was 100 μg / mL. The initial sample was prepared as follows: 100 μL was immediately removed from the plasma reaction and mixed with 100 μL of acetonitrile to quench the reaction, which caused precipitation. The insoluble matter was precipitated into clumps by centrifugation at 15,000 rpm for 5 minutes at 25°C. The sample for LC-MS analysis was prepared by combining 75 μL of the supernatant with 75 μL of water. LC-MS analysis was performed as follows: 10 μL was injected, and the sample was eluted using the equipment and gradient described in the General UPLC Materials and Methods section. Additional samples were prepared after 10, 30, 45, 60, 120, 180, and 240 minutes. The loss of the compound and the increase of dantrolene were quantitatively determined by manually integrating the peaks in the 385 nm chromatogram. Figure 5 The half-life of compound 2a under these conditions was obtained by fitting the data with an exponential function. The half-life of compound 2a was 82 minutes. A negative control was also performed using phosphate-buffered saline instead of rat plasma, and no conversion was observed.

[0349] The plasma pharmacokinetics of dantrolene following administration of compound 2a to live rats were determined by intravenous administration to cannulated Sprague-Dawley rats and analysis of plasma dantrolene concentrations using LC-MS. Figure 6 (See Table 1). Dosing, sample collection, and analysis were performed by MPI Research. Intravenous doses were administered via jugular vein cannulation. Intramuscular doses were administered by injecting half the dose into a large muscle in the left hind limb and the other half into the right hind limb. Five rats were used in each test group. Compound 2a was dissolved in sterile filtered 5% mannitol aqueous solution (w / v) to a concentration of 8 mg / mL to obtain a dose level of 7.5 mg / kg and a dose volume of 0.94 mL / kg. Plasma was collected before administration and at 0.033, 0.083, 0.167, and 0.33, 0.66, 1, 2, 3, 6, and 9 hours after administration.

[0350] In each experiment, only trace amounts of the compound were observed 2 years after administration via IV, IM, or SC. Therefore, pharmacokinetic measurements were performed only at dantrolene concentrations.

[0351] Table 1. Plasma bioavailability of dantrolene after intravenous and intramuscular administration of compound 2a.

[0352]

[0353] In the second analysis, the whole blood pharmacokinetics of compound 2a in live rats were performed. Figure 7 (See Table 2). Each test group consisted of three rats. Compound 2a was dissolved in sterile filtered 5% mannitol aqueous solution (w / v) to a concentration of 8 mg / mL to obtain a dose level of 7.5 mg / kg and a dose volume of 0.94 mL / kg. Blood samples were collected before administration and at 0.033, 0.083, 0.167, and 0.33, 0.66, 1, 2, 3, 6, and 9 hours after administration. Whole blood samples were diluted 4-fold in acetonitrile and centrifuged to remove precipitates.

[0354] Table 2. Whole blood bioavailability of dantrolene after intravenous and intramuscular administration of compound 2a.

[0355]

[0356] Another pharmacokinetic analysis of compound 2a was performed in live rats. Figure 8 and 9 (Table 3). Five rats were used in each test group. Compound 2a was dissolved to 8 mg / mL in sterile filtered 5% mannitol aqueous solution (w / v) to obtain a dose level of 7.5 mg / kg and a dose volume of 0.94 mL / kg. Blood was collected before administration and at 0.033, 0.083, 0.167, and 0.33, 0.66, 1, 2, 3, 6, and 9 hours after administration. Whole blood samples were diluted 4-fold in acetonitrile and centrifuged to remove precipitates. Plasma samples were obtained by placing blood in K2EDTA tubes and then centrifuging. The resulting plasma was then aspirated and diluted 4-fold in acetonitrile, followed by another round of centrifugation to remove precipitates.

[0357] Table 3. Bioavailability of dantrolene after administration of compound 2a via intravenous, intramuscular, and subcutaneous routes.

[0358]

[0359] a Whole blood analysis b plasma analysis

[0360] Transformation and pharmacokinetics of compound 2b in plasma

[0361] To measure the conversion of compound 2b to dantrolene in rat plasma, 1 mg of compound 2b was dissolved in water to a concentration of 1 mg / mL. A 100 μL aliquot of compound 2b was combined with 900 μL of plasma from Sprague Dawley rats. The plasma reaction was then incubated in a 37°C water bath. The initial concentration of compound 2b in the plasma was 100 μg / mL. The initial sample was prepared as follows: 100 μL was immediately removed from the plasma reaction and mixed with 100 μL of acetonitrile to quench the reaction, which caused precipitation. The insoluble matter was precipitated into clumps by centrifugation at 15,000 rpm for 5 minutes at 25°C. The sample for LC-MS analysis was prepared by combining 75 μL of the supernatant with 75 μL of water. LC-MS analysis was performed by injecting 10 μL of the sample, eluting the sample using the equipment and gradient described in the General UPLC Materials and Methods section. Additional samples were prepared after 15, 30, 45, 60, 120, 180, and 240 minutes. The loss of compound 2 and the increase of dantrolene were quantified by manual integration of the peak in the 385 nm chromatogram. Figure 10 The half-life of compound 2b under these conditions was obtained by fitting the data with an exponential function. The half-life of compound 2b was 77 minutes. A negative control was also performed using phosphate-buffered saline instead of rat plasma, and no conversion was observed.

[0362] The pharmacokinetics of dantrolene following administration of compound 2b to live rats were determined by intravenous administration and analysis of blood dantrolene concentrations using LC-MS. Figure 11 (See Table 4). Dosing, sample collection, and analysis were performed by MPI Research. Intravenous doses were administered via jugular vein cannulation. Intramuscular doses were administered by injecting half the dose into a large muscle in the left hind limb and the other half into the right hind limb. Five rats were used in each test group. Compound 2b was dissolved in sterile filtered 5% mannitol aqueous solution (w / v) to a concentration of 11.4 mg / mL to obtain a dose level of 10.6 mg / kg and a dose volume of 0.94 mL / kg. Plasma was collected before administration and at 0.033, 0.083, 0.167, and 0.33, 0.66, 1, 2, 3, 6, and 9 hours after administration.

[0363] Table 4. Plasma bioavailability of dantrolene after intravenous and intramuscular administration of compound 2b.

[0364]

[0365] Example 18. Recirculation of compound 10c in plasma

[0366] To measure the conversion of 10c to dantrolene in rat plasma, 0.9 mg of 10c was dissolved in water to a concentration of 1 mg / mL. 100 μL aliquots of 10c were combined with plasma from Sprague Dawley rats. The plasma reaction was then incubated in a 37°C water bath. The initial concentration of 10c in the plasma was 100 μg / mL. The initial sample was prepared as follows: 100 μL was immediately removed from the plasma reaction and mixed with 100 μL of acetonitrile to quench the reaction, which caused precipitation. The insoluble matter was precipitated into clumps by centrifugation at 15,000 rpm for 5 minutes at 25°C. A sample for LC-MS analysis was prepared by combining 75 μL of the supernatant with 75 μL of water. LC-MS analysis was performed by injecting 10 μL of the sample, eluting the sample using the equipment and gradient described in the General UPLC Materials and Methods section. Additional samples were prepared after 10, 20, 35, 45, and 60 minutes. The loss of 10C and the increase of dantrolene were quantified by manual integration of the peak in the 385 nm chromatogram. Figure 10 The half-life of 10c under these conditions was obtained by fitting the data with an exponential function. Within the first 10 minutes of incubation with plasma, 10c was completely converted to dantrolene. The estimated half-life was 1.9 min. A negative control was also performed using phosphate-buffered saline instead of rat plasma, and a slow conversion to dantrolene was observed. The half-life of 10c in phosphate-buffered saline was 294 min. See also Figure 12 .

[0367] Example 19. Plasma conversion and pharmacokinetics of compound 12a

[0368] To measure the conversion of 12a to dantrolene in rat plasma, a 1 mg / mL solution of 12a was prepared in water. 100 μL aliquots of 12a were combined with plasma from Sprague Dawley rats. The plasma reaction was then incubated in a 37°C water bath. The initial concentration of 12a in the plasma was 100 μg / mL. The initial sample was prepared as follows: 100 μL was immediately removed from the plasma reaction and mixed with 100 μL of acetonitrile to quench the reaction, which caused precipitation. The insoluble matter was precipitated into clumps by centrifugation at 15,000 rpm for 5 minutes at 25°C. A sample for LC-MS analysis was prepared by combining 75 μL of the supernatant with 75 μL of water. LC-MS analysis was performed by injecting 10 μL of the sample, eluting the sample using the equipment and gradient described in the General UPLC Materials and Methods section. Additional samples were prepared after 10, 20, 35, 45, 60, 75, and 150 minutes. The loss of 12a and the increase of dantrolene were quantified by manual integration of the peak in the 385 nm chromatogram. Figure 13The half-life of 12a under these conditions was obtained by fitting the data with an exponential function. The calculated half-life was 12.2 min. A negative control was also performed using phosphate-buffered saline instead of rat plasma, with a half-life greater than 150 min.

[0369] The pharmacokinetic properties of 12a were determined in the same manner as 22c, except that the compound was dissolved in 5% mannitol to a concentration of 5 mg / mL to provide a dose level of 4 mg / kg and a dose volume of 0.8 mL / kg. Figure 14 ).

[0370] Example 20. Plasma conversion of compound 17b

[0371] To generate a standard curve for calculating the 17b concentration, 1 mg of the compound was dissolved in 200 μL of dimethylformamide to obtain a 5 mg / mL solution. This 5 mg / mL solution was diluted 5-fold in water and then centrifuged at 15,000 rpm for 5 min at 25 °C. The supernatant was used for a 10-fold dilution in an aqueous solution of 25% acetonitrile to obtain a 100 μg / mL solution. A series of 2-fold dilutions were performed using 25% acetonitrile to obtain solutions of 50, 25, 12.5, and 6.25 μg / mL. These solutions were analyzed as described in the General UPLC Materials and Methods section, except that the gradient was scaled up to end with 90% acetonitrile after 14.4 min.

[0372] To measure the conversion of 17b to dantrolene in rat plasma, a 1 mg / mL solution of 17b was prepared in water. 100 μL aliquots of 17b were combined with 900 μL of plasma from Sprague Dawley rats. The plasma reaction was then incubated in a 37°C water bath. The initial concentration of 17b in the plasma was 100 μg / mL. The initial sample was prepared as follows: 100 μL was immediately removed from the plasma reaction and mixed with 100 μL of acetonitrile to quench the reaction, which caused precipitation. The insoluble matter was precipitated into clumps by centrifugation at 15,000 rpm for 5 minutes at 25°C. A sample for LC-MS analysis was prepared by combining 75 μL of the supernatant with 75 μL of water. LC-MS analysis was performed by injecting 10 μL of the sample, and the sample was eluted using the apparatus described in the General UPLC Materials and Methods section. Additional samples were prepared after 15, 30, 45, 60, 180, 240, and 300 minutes. The loss of 17b and the increase of dantrolene were quantified by manual integration of the peak in the 385 nm chromatogram. Figure 15 The half-life of 17b under these conditions was obtained by fitting the data with an exponential function. The calculated half-life was 94.5 min. A negative control was also performed using phosphate-buffered saline instead of rat plasma, and the estimated half-life was 447 min.

[0373] Example 21. Plasma conversion and pharmacokinetics of compound 22c

[0374] To measure the conversion of 22c to dantrolene in rat plasma, a 1 mg / mL solution of 22c was prepared in water. 100 μL aliquots of 22c were combined with 900 μL of plasma from Sprague Dawley rats. The plasma reaction was then incubated in a 37°C water bath. The initial concentration of 22c in the plasma was 100 μg / mL. The initial sample was prepared as follows: 100 μL was immediately removed from the plasma reaction and mixed with 100 μL of acetonitrile to quench the reaction, which caused precipitation. The insoluble matter was precipitated into clumps by centrifugation at 15,000 rpm for 5 minutes at 25°C. A sample for LC-MS analysis was prepared by combining 75 μL of the supernatant with 75 μL of water. LC-MS analysis was performed by injecting 10 μL of the sample, eluting the sample using the equipment and gradient described in the General UPLC Materials and Methods section. Additional samples were prepared after 10, 20, 35, 45, 60, 75, and 150 minutes. The loss of 22C and the increase of dantrolene were quantified by manual integration of the peak in the 385 nm chromatogram. Figure 16 The half-life of 22c under these conditions was obtained by fitting the data with an exponential function. The estimated half-life was 3.2 min. A negative control was also performed using phosphate-buffered saline instead of rat plasma, and 85% reactivation was observed after 150 min.

[0375] The pharmacokinetics of 22c in live rats were determined by administering dantrolene to cannulated Sprague-Dawley rats and analyzing blood dantrolene levels using LC-MS. Figure 17 Intravenous doses were administered via jugular vein cannulation. Subcutaneous doses were administered by injection between the skin and the underlying tissue layer of the left hind limb of each animal. Intramuscular doses were administered by injecting half the dose into a large muscle of the left hind limb and the other half into the right hind limb. Three rats were used in each test group. 22c was dissolved in a sterile filtered 5% mannitol aqueous solution (w / v) containing 10% DMSO to a concentration of 3 mg / mL to provide a dose level of 4 mg / g and a dose volume of 1.33 mL / kg. Blood was collected before administration and at 0.167, 0.33, 0.66, and 1.33 hours after administration. At each time point, 100 μL of blood was collected into a vial containing 300 μL of acetonitrile. The blood sample was centrifuged to precipitate any insoluble matter. The supernatant was rapidly frozen and stored on dry ice. Samples were analyzed using the methods described in the General UPLC Materials and Methods and Pharmacokinetic Analysis sections.

Claims

1. A compound of formula I: I Or its pharmaceutically acceptable salt. Where R is -P(O)(OH)2.

2. The compound according to claim 1, wherein it is in the form of a pharmaceutically acceptable salt.

3. The compound according to claim 1, wherein the compound is a diaminobutyrithiol salt of formula IA: 。 4. A pharmaceutical composition comprising the compound of any one of claims 1-3 or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable excipient.

5. Use of the compound of any one of claims 1 to 3 or a pharmaceutically acceptable salt thereof in the preparation of a medicament for treating a condition responsive to dantrolene, wherein the medicament is in a form for intramuscular or subcutaneous administration.

6. The use according to claim 5, wherein the condition is malignant hyperthermia, chronic rigidity, exertional heatstroke, cardiac arrhythmia, cardiac arrest, myocardial infarction, heart failure, myocardial injury, cardiomyopathy, central axillary dysplasia, amyotrophic lateral sclerosis, rhabdomyolysis, Duchenne muscular dystrophy, ataxia, overactive bladder, epilepsy, antipsychotic malignant syndrome, human stress disorder, Alzheimer's disease, Huntington's disease, multiple sclerosis, Parkinson's disease, ischemia-reperfusion injury, neuronal reperfusion injury, hypoxia, cerebral aneurysm, stroke, hyperthermia related to drug abuse or overdose, exposure to neurotoxins, exposure to neurotoxic gases, or acetylcholine accumulation.

7. The use according to claim 5, wherein the condition is tachycardia, atrial fibrillation, detrusor overactivity, epileptic seizure, or subarachnoid hemorrhage.

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